BIOACID Programme - Natural Environment Research Council

BIOACID 

A proposed national ‘Verbundprojekt’ of the Federal Ministry of Education and Research 

Coordinator: 

Prof. Dr. Ulf Riebesell 

Forschungsbereich Marine Biogeochemie 

Leibniz-Institut für Meereswissenschaften an der Universität Kiel 

Düsternbrooker Weg 20 

24105 Kiel 

Tel. 0431-600-4444 

Email: uriebesell@ifm-geomar.de 

Partners: 

Alfred-Wegener-Institut für Polar- und Meeresforschung, Bremerhaven 

Carl von Ossietzky Universität Oldenburg 

Christian-Albrechts-Universität zu Kiel 

Forschungszentrum TERRAMARE, Wilhelmshaven 

GKSS-Forschungszentrum Geesthacht GmbH 

Heinrich-Heine-Universität, Düsseldorf 

Jacobs – University, Bremen 

Leibniz-Institut für Gewässerökologie und Binnenfischerei, Berlin 

Leibniz-Institut für Meereswissenschaften IFM-GEOMAR, Kiel 

Leibniz-Institutes für Ostseeforschung Warnemünde 

Ludwig-Maximilians-Universität München 

Max-Planck-Institut für Marine Mikrobiologie, Bremen 

PreSens Precision Sensing GmbH, Regensburg 

Ruhr-Universität Bochum 

Universität Bremen 

Universität Hamburg 

Universität Rostock 

Westfälische Wilhelms - Universität Münster 

Zentrum für Marine Tropenökologie (ZMT), Bremen

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BIOACID: Biological Impacts of Ocean Acidification 

Zusammenfassung…………………………………………………………………... 4 

Summary……………………………………………………………………………. 5 

1. Project background……………………………………………………………. 6 

2. Scientific and technological objectives………………………………………. . 8 

3. Relevance to National Priorities and the National Funding Policy………….. 9 

4. Structure of the Project and Consortium Building……………………….….. 10 

5. Overview of Themes 1-5………………………………………………………... 13 

5.1. Theme 1: Primary production, microbial processes 

and biogeochemical feedbacks ………………………………..... 13 

5.2. Theme 2: Performance characters: reproduction, growth and 

behaviours in animal species ………………………………….... 16 

. 

5.3. Theme 3: Calcification - sensitivities across phyla and ecosystems....…... 19 

5.4. Theme 4: Species interactions and community structure 

in a changing ocean....................................................................... 22 

5.5. Theme 5: Integrated assessment – sensitivities and uncertainties….….... 25 

6. Management structure and procedures ………………………………….……. 28 

7. Data management and dissemination………………………………………….. 31 

8. Infrastructure development, training and transfer of know-how………….… 32 

9. International and National Cooperation……………………………………..... 33 

10. Summary Budget………………………………………………………………... 38 

11. Detailed descriptions of Themes and Projects……………………………….... 39 

11.1: Theme 0: Overarching activities……………………………………….... 41 

0.1 Project coordination……………………………..………………………….. 41 

0.2 BIOACID data management…………………………………………….….. 44 

0.3 Infrastructure development……………………………………..................... 49 

0.4 Training and transfer of knowhow……………………………........……..… 63 

11.2: Theme 1: Primary production, microbial processes 

and biogeochemical feedbacks…………………………….….. 67 

1.1: Acclimation versus adaptation in autotrophs………………………………. 70 

1.2: Turnover of organic matter……………………………………………....…. 85 

1.3: Modelling biogeochemical feedbacks of the organic carbon pump……..…. 98 

11.3: Theme 2: Performance characters: reproduction, growth and 

behaviours in animal species………………………………..... 105 

2.1: Effects on grazers and filtrators……………………………………….….... 108 

2.2: Long-term physiological effects on different life stages 

of benthic crustaceans.................................................................................... 121 

2.3: Effects on top predators (fishes, cephalopods)…………………………….. 134


11.4: Theme 3: Calcification - sensitivities across phyla and ecosystems….. 145 

3.1: Cellular mechanisms of calcification……………………………………….. 148 

3.2: Calcification under pH-stress: Impacts on ecosystem and 

organismal levels………………………………………………………….... 168 

3.3: Ultra-structural changes and trace element / isotope partitioning in 

calcifying organisms…………………………………………………… …... 183 

3.4: Microenvironmentally controlled (de-)calcification mechanisms…………... 193 

3.5: Impact of present and past ocean acidification on metabolism, 

biomineralization and biodiversity of pelagic and neritic calcifiers………… 207 

11.5: Theme 4: Species interactions and community structure in 

a changing ocean........................................................................ 219 

4.1: OA impacts on interactions in and structure of benthic communities………. 224 

4.2: OA effects on food webs and competitive interactions 

in pelagic ecosystems ……………………………………………………... 236 

11.6: Theme 5: Integrated assessment – sensitivities and uncertainties……. 247 

5.1: Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle 

and the primary production in the North Sea………………………………… 249 

5.2: Evaluating and optimising parameterisations of pelagic calcium 

carbonate production in global biogeochemical ocean models………………. 255 

5.3: Viability-method for the impact assessment of ocean acidification 

under uncertainty……………………………………………….………….…. 261 

12. Appendices………………………………………………………………………. 264 

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Zusammenfassung 

4 


Neben der Atmosphäre ist der Ozean die zweitgrößte Senke für anthropogenes Kohlendioxid. 

Das vom Ozean aufgenommene CO2 reagiert mit dem Meerwasser und bildet Hydrogenkarbonat 

und Säure, bei gleichzeitiger Zehrung von Karbonationen. Das Ergebnis dieses Prozesses, der als 

Ozeanversauerung bezeichnet wird, ist ein Anstieg der Konzentrationen von CO2 und 

Hydrogenkarbonat und eine Abnahme des pH Wertes (Zunahme des Säuregrades) und der 

Karbonationenkonzentration. Bei ungebremst fortschreitenden CO2 Emissionen wird sich die 

Chemie des Meerwassers bis zum Ende dieses Jahrhundert in einer Weise verändern, wie es die 

meisten der heute im Ozean lebenden Organismen während ihrer jüngeren Evolution nicht erlebt 

haben. Dies könnte Folgen für die Konkurrenzfähigkeit pH/CO2 sensibler Arten haben. Bei 

zunehmender Ozeanversauerung droht dadurch der Verlust an Biodiversität, ökologische und 

funktionale Veränderungen in den marinen Lebensgemeinschaften und eine reduzierte Kapazität 

des Ozeans zur weiteren Aufnahme von anthropogenem CO2. 

Trotz der Risiken, die diese Entwicklung in sich birgt, fehlt es uns an einem grundlegenden 

Verständnis der möglichen Konsequenzen einer Ozeanversauerung. Um diese Lücke zu schließen 

und eine System basierte Abschätzung der hiermit verbundenen Risiken und Unsicherheiten zu 

erlangen, wird BIOACID die Expertise von Molekular- und Zellbiologen, Biochemikern, 

Pflanzen- und Tierphysiologen, Meeresökologen, marinen Biogeochemikern und 

Ökosystemmodellierern in einem integrierenden Ansatz kombinieren. Über Disziplin-, Themen- 

und Projektgrenzen hinweg werden die BIOACID Partner gemeinschaftlich Experimente 

durchführen, Labor übergreifend Ausrüstung und Messkapazitäten nutzen, Probenmaterial und 

Fachkompetenz austauschen und die umfangreichen Datensätze in Richtung auf ökosystemare 

Modelle der Ozeanversauerung analysieren und synthetisieren. Dies wird ergänzt durch 

Trainingsworkshops zu zentralen Forschungsinhalten und -methoden von BIOACID Experten für 

alle Mitglieder des Konsortiums. Die übergeordneten Fragestellungen des Projektes sind: 

Welche Auswirkungen hat die Ozeanversauerung auf die marinen Organismen und ihre 

Lebensräume, was sind die zu Grunde liegenden Mechanismen und möglichen 

Anpassungen auf der Ebene von Populationen und Gemeinschaften, in welchem Maße 

werden die Auswirkungen durch andere Stressfaktoren beeinflusst und welche 

Konsequenzen ergeben sich daraus für die marinen Ökosysteme, biogeochemischen 

Kreisläufe und mögliche Rückkopplungen auf das Klimasystem? 

Um diese Fragen über einen weiten Bereich von potentiell sensitiven biologischen Prozessen 

anzuwenden, sind die in BIOACID geplanten Forschungsaktivitäten nach Schlüsselkomponenten 

und funktionalen Gruppen der marinen Lebensgemeinschaften strukturiert. Aus den diversen 

experimentellen Ansätzen und Feldbeobachtungen gewonnene Erkenntnisse sollen auf der Basis 

einer integrierenden Datensynthese dazu beitragen, Unsicherheiten bezüglich der 

prognostizierten Entwicklung zu identifizieren und mögliche Schwellenwerte unumkehrbarer 

Veränderungen zu erkennen. Ein zusätzlicher Mehrwert für das Projekt soll durch enge 

Kooperation mit verwandten nationalen und internationalen Projekten mit Schwerpunkt auf 

Ozeanversauerung und Ozeanerwärmung erzielt werden.

Summary 


Next to the atmosphere, the ocean is the second largest sink for anthropogenic carbon dioxide. As 

fossil fuel CO2 enters the surface ocean, it reacts with seawater to form bicarbonate and protons, 

thereby consuming carbonate ions. The net result of this process, termed ocean acidification, is 

an increase in CO2 and bicarbonate concentrations and a decrease in seawater pH and carbon ion 

concentration. If CO2 emissions continue to rise at current rates, before the end of this century the 

resulting changes in seawater chemistry will expose marine organisms to conditions which they 

have not experienced during their recent evolutionary history and which may pose a threat to the 

competitive fitness of pH/CO2 sensitive species and groups. Thus, as the ocean continues to 

acidify, there is an increasing risk of biodiversity losses, of profound ecological and functional 

shifts, and of a reduced capacity of the ocean to buffer further CO2 increase. 

Despite this emerging problem and the risks it may involve, surprising little is know about the 

possible impacts of ocean acidification. To close the many gaps in our understanding and to 

allow a systems-based assessment of the risks and uncertainties associated with ocean 

acidification, BIOACID will take an integrated approach combining the expertise of molecular 

and cell biologists, biochemists, plant and animal physiologists, marine ecologists, ocean 

biogeochemists and ecosystem modellers. The interaction between BIOACID scientists across 

disciplines, research themes and projects will include joint experiments, collaborative use of 

equipment and measurement capacity, exchange of samples and expertise, and the analysis and 

synthesis of comprehensive data sets towards an ecosystem model of ocean acidification. These 

activities will be complemented by training workshop offered by BIOACID experts to all 

members of the consortium. Following this approach, the overarching questions of BIOACID 

are: 

What are the effects of ocean acidification on marine organisms and their habitat, what are 

the underlying mechanisms of responses and possible adaptations on the level of 

populations and communities, how are they modulated by other environmental stressors, 

and what are the consequences for marine ecosystems, ocean biogeochemical cycles, and 

possible feedbacks to the climate system? 

To address these questions for a wide range of potentially sensitive biological processes, research 

activities will be structured according to key ecosystem components and functional groups. 

Information gained in a variety of experimental approaches and field assays will be synthesized 

to obtain an integrated assessment of sensitivities and uncertainties and to identify the potential 

thresholds associated with ocean acidification. BIOACID will benefit from close interactions 

with related national and international project focussing on ocean acidification and ocean 

warming. 

5

1. Project background 

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The world’s oceans help moderating climate change thanks to their extensive capacity to store 

anthropogenic carbon dioxide. Since pre-industrial times, the oceans have sequestered nearly half 

of the fossil fuel CO2 released into the atmosphere and presently take up approximately 30% of 

current CO2 emissions (Sabine et al. 2004). As CO2 enters the surface ocean it reacts with 

seawater and generates changes in carbonate chemistry already measurable today (Figure 1). In 

case of unabated CO2 emissions, the resulting changes in seawater chemistry will, in the course 

of this century, expose marine organisms to conditions which they may not have experienced 

during their recent evolutionary history (Raven et al., 2005). This raises concerns regarding the 

biological, ecological, biogeochemical, and societal implications of ocean acidification. 

Fig. 1: Measured changes in surface ocean CO 2 partial pressure (pCO 2) and pH at the European Station for 

Time-series in the Ocean off the Canary Islands (ESTOC), the Hawaii Ocean Time-series Station 

(HOT) and the Bermuda Atlantic Times Series station (BATS). Since 1750, surface ocean pH has 

decreased by 0.12 units (calculated). Since 1980, pH has decreased by 0.02 units per decade (measured). 

Source: IPCC 4 th Assessment Report (2007) 

In its Special Report “The Future Oceans – Warming up, Rising High, Turning Sour”, the 

German Advisory Council on Global Change (WBGU, Berlin 2006) states: “Because of the 

importance of the consequences of ocean acidification, research in this area should be intensified 

considerably. As long as there is no general scientific consensus about the tolerable limit for the 

effects of acidification, a margin of safety according to the precautionary principle should be 

observed. The suggestion of the WBGU to prevent a pH decrease of more than 0.2 is oriented 

toward the goal of avoiding an aragonite undersaturation in the ocean surface layer. If it is found 

that other intolerable damages already occur before reaching aragonite undersaturation, then the 

guard rail will have to be adjusted accordingly.” (see Fig. 2)


Fig. 2: Mean surface ocean pH values during glacial a pre-industrial times (calculated from reconstructed 

atmospheric pCO2), measured pH values in the present ocean, and projected future seawater pH for an 

atmospheric CO 2 concentration of approx. 750 ppm. The red line indicates the WBGU guard rail. Source: 

WBGU Special Report (2006) after IMBER (2005) 

As stated in the report, the tolerable window for ocean acidification defined by WBGU presently 

relies on an extremely small data base. In fact, rather than using the limited data on observed 

biological consequences of ocean acidification, the WBGU reaches its recommendation on the 

basis of projected changes in water chemistry (aragonite saturation state). While this is an 

appropriate approach in view of the scarcity of biological information, there is a clear need to 

establish a reliable data base on tolerance levels for ocean acidification in key groups of pH/CO2 

sensitive marine organisms in order to reach a more informed recommendation. 

Our present knowledge on the effects of ocean acidification (OA) on the marine biota is largely 

based on experimental work with single organisms or strains maintained in short-term 

incubations often exposed to abrupt and extreme changes in carbonate chemistry. Based on 

presently available data little is known about OA induced habitat change, synergetic effects from 

other stressors, such as ocean warming, the sensitivity of genetically diverse populations, and the 

ability of organisms to undergo long-term physiological and genetic adaptations. Hence, it is 

difficult to predict how the responses of key species will affect the population, community and 

ecosystem level, for example by the replacement of OA-sensitive with OA-tolerant species. In 

view of these uncertainties, it is presently impossible to define critical thresholds (tipping points) 

for tolerable pH decline or to predict the pathways of ecosystem changes where threshold levels 

have been surpassed. 

BIOACID will take the challenge to address the substantial gaps in the understanding of the 

consequences of ocean acidification. To achieve this, BIOACID will take an integrated approach 

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combining the expertise of molecular and cell biologists, biochemists, plant and animal 

physiologists, marine ecologists, ocean biogeochemists and ecosystem modellers. Close 

collaboration will be established with other European national and EU projects on ocean 

acidification to complement the expertise and research capacities of the marine science 

community in Germany and to benefit from the emerging synergies. Most notably, BIOACID 

will profit from existing links to the European Project on Ocean Acidification (EPOCA) and 

intends to develop close interactions and possibly joint activities with the UK programme on 

ocean acidification presently developed in the framework of the Natural Environment 

Research Council’s (NERC) new five-year strategy. 

References 

Fabry VJ, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES J Mar Sci 65: 

414–432. 

Feely RA, Sabine CL, Hernandez-Ayon JM, Ianson D, Hales B (2008) Evidence for upwelling of corrosive "acidified" water onto the 

continental shelf. Science 320: 1490 – 1492. 

Raven et al. Royal Society, Ocean Acidification Due to Increasing Atmospheric Carbon Dioxide (Policy Document 12/05, Royal Society, 

London, 2005). 

Sabine, C.L., R.A. Feely, N. Gruber, R.M. Key, K. Lee, J.L. Bullister, R. Wanninkhof, C.S. Wong, D.W.R. Wallace, B. Tilbrook, F.J. Millero, 

T.-H. Peng, A. Kozyr, T. Ono, and A.F. Rios (2004), The Oceanic sink for Anthropogenic CO2, Science, 305, 367-371. 

WBGU Special Report “The Future Oceans – Warming up, Rising High, Turning Sour”, the German Advisory Council on Global Change, 

Berlin 2006 

2. Scientific and technological objectives 

BIOACID will assess uncertainties, risks and thresholds related to the emerging problem of 

ocean acidification (OA) at molecular, cellular, organismal, population, community and 

ecosystem scales. It will 

contribute to determining the impacts of OA on marine biota and their potential for 

acclimation and adaptation, 

improve our understanding of OA-related ecosystem changes, including both pelagic and 

benthic habitats, synthesize information for ecosystem modelling and determine critical 

thresholds, 

assess the consequences of OA-induced biological responses on elemental cycling and 

biogeochemical feedbacks to the climate system. 

To achieve this goal, BIOACID will (i) employ a suite of observational, experimental and 

modelling approaches leading to an integrated assessment of potential risks and thresholds 

associated with ocean acidification (see Section 4 and 10), and (ii) coordinate with major national 

and international projects and programmes (see Section 8). The project aims to deliver scientific 

information and knowledge that directly addresses important global environmental issues and 

that is relevant to national policy issues relating to energy policy, the biodiversity agenda, and 

adaptation to global change. Major deliverables will include:


high-profile scientific contributions to IGBP’s international science programmes of IMBER 

(Integrated Marine Biogeochemical and Ecosystem Research) and SOLAS (Surface Ocean 

Lower Atmosphere Study) based on Germany’s national marine expertise and infrastructure 

strengthened scientific basis for policy-relevant assessments, including input into the IPCC 

(Intergovernmental Panel on Climate Change) 5 th Assessment Report and providing the 

scientific foundation for assessments of changes in marine biodiversity, e.g. in the framework 

of DIVERSITAS. 

3. Relevance to National Priorities and the National Funding Policy 

BIOACID addresses topics of global change research that are central to the BMBF’s mission 

(see below) and which are of immediate relevance with regard to the “Positionspapier für eine 

kohärente deutsche Forschungsstrategie zum Globalen Wandel” developed by the Nationales 

Komitee für Global Change Forschung to guide national research over the next years. BIOACID 

will take the challenge of providing a sound data base on which to make recommendations 

regarding critical thresholds for ocean acidification as requested in the Special Report “The 

Future Oceans – Warming up, Rising High, Turning Sour” by the German Advisory Council on 

Global Change (WBGU, Berlin 2006). BIOACID’s combination of experimental approaches, 

field studies and modelling activities to assess different scenarios of future CO2 levels, targeting 

in a coordinated manner all relevant entities from the cell to the ecosystem and from short to 

long-term effects, is a unique and ambitious approach to the emerging problem of ocean 

acidification. Research carried out under BIOACID is also relevant as a basis for informed 

decisions on adaptation and mitigation strategies developed in the context of the 

“Aktionsprogramm Forschung zum Klimawandel”. 

In particular, BIOACID directly addresses goals of the BMBF’s Earth System and Marine 

Research programmes by focussing on the response of marine organisms and ecosystems to a 

changing global environment. The project is designed to assess the sensitivity of key species, 

communities, and habitats to variable forcing including anthropogenic change such as 

elevated CO2 levels. In this way, the project will improve the understanding of mechanisms by 

which the ocean and its ecosystems react to human and natural impacts, including feedbacks of 

such changes on biogeochemical cycles, the atmospheric composition and climate. The project 

will therefore contribute to a better understanding of the role of the ocean in the climate 

system and an improved description of the effects of global change on sensitive marine 

ecosystems. Such understanding is a pre-requisite for prediction and assessment of the impacts of 

human activity including climate-protection measures, on climate, global ecosystems and marine 

natural resources. 

Due to its multi-disciplinary nature, BIOACID also contributes directly to at least three Priority 

Research Themes of the BMBF’s Geotechnology Programme, namely: (1) The Coupled System 

Earth – Life, (2) Global Climate Change: Causes and Effects and (3) Biogeochemical 

Cycles: Links between Geosphere and Biosphere. Furthermore, BIOACID research is of 

immediate relevance to the new BMBF framework programme “Research for Sustainability” 

and will make significant contributions to risk assessments on the loss of marine biodiversity, 

including input to the Global Biodiversity Information Facility. 

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4. Structure of the Project and Consortium Building 

4.1. Scientific programme 

In order to achieve the objectives of BIOACID, five thematic areas have been identified which 

cover the range of processes from the base of the marine food chain to the community and 

ecosystem level, and of mechanisms from the sub-cellular to the whole organism level. In view 

of their distinct sensitivities to ocean acidification, calcification and carbonate dissolution 

processes will be the focal point of a separate theme. According to these research priorities the 

scientific programme of BIOACID is structured under the following five themes: 

Theme 1: Primary production, microbial processes 

& biogeochemical feedbacks 

Theme 2: Performance characters: 

Reproduction, growth & behaviours in animal species 

Theme 3: Calcification: Sensitivities across phyla and ecosystems 

Theme 4: Species interactions and community structure in a changing ocean 

Theme 5: Integrated assessment: Sensitivities and uncertainties 

BIOACID will pursue a multidisciplinary approach, involving scientists with expertise in cell and 

molecular biology, microbiology, physiology, evolutionary biology, marine ecology, marine 

chemistry, biogeochemistry and numerical modelling. BIOACID will employ a wide range of 

scientific approaches and methodologies, extending from field monitoring of OA-sensitive areas 

and ecosystems to joint mesocosm perturbation experiments, combined with closely coordinated 

ecosystem and biogeochemical modelling activities using parameterizations of observed 

biological responses and their biogeochemical implications.


Project coordination 

Data management 

Infrastructure 

development 

Training and transfer 

of knowhow 

Theme 1 

Primary production, 

microbial processes 

and biogeochemical 

feedbacks 

Theme 3 

Calcification: 

Sensitivities across 

phyla and ecosystems 

Theme 5 

Integrated 

assessment: 

Sensitivities and 

uncertainties 

Theme 2 

Performance characters: 

reproduction, growth 

and behaviours in 

animal species 

Theme 4 

Species interactions 

and community 

structure in a 

changing ocean 

Fig. 3: Structure of the project: research themes and overarching activities. Joint activities and the multiple 

links between themes and projects are described in detail in section 11. 

During the development of the research programme strong emphasis was put on linking the subprojects 

and projects, both within and between themes. These links range from joint experiments, 

joint use of equipment and measurement capacity, to exchange of samples, exchange of expertise, 

and training workshop offered by BIOACID experts to all members of the consortium (see 8.2. 

Training and transfer of know how). For detailed information on the multiple linkages developed 

in BIOACID see Table 0.1 under 11. Detailed Description of Themes and Projects and the 

specific links described in each of the individual projects. Over-arching activities include not 

only project coordination and logistics, outreach and data management but also the development 

and application of joint infrastructure (CO2 sensors, benthic and pelagic mesocosm systems, 

NMR, NanoSIMS). 

4.2. Consortium building 

The BIOACID consortium was built based on a bottom-up, open competition approach among all 

interested German institutes and universities conducting marine-oriented research. Starting with a 

presentation of the project idea to representatives of the Projektträger Jülich (PTJ) in 

Warnemünde in May 2007, followed by a positive signal from the PTJ, a planning group (Table 

1) was formed and first met at the MPI in Bremen July 2007. 

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Table 1: Composition of the planning group. 

Planning group Members 

Chairs: Ulf Riebesell (IFM-GEOMAR), Hans-Otto Pörtner (AWI) 

Members: 

Antje Boetius (MPI Bremen), Gerd Graf* (U Rostock), Thorsten Reusch 

(U Münster), Maren Voss (IOW) 

* during the pre-proposal writing stage Gerd Graf (U Rostock) asked to withdraw from the 

planning group; Maren Voss (IOW) was invited as a replacement 

In the following months the planning group developed a tentative structure for a German national 

programme on ocean acidification and prepared a core proposal which was submitted to the 

German Ministry for Education and Research (BMBF) in November 2007. An oral presentation 

of the core proposal to representatives of the BMBF (Referat 725 System Erde and Referat 723 

Globaler Wandel) and of the PTJ took place in Bonn in December 2007. Based on this 

presentation and the follow-up discussions the planning group was invited to prepare a full 

proposal for international peer review. 

In January 2008 a call for project contributions was made public and sent to potential partner 

universities/institutes, requesting a 2-page description of proposed contributions to the 

programme by February 15, 2008. Over 50 pre-proposals were received and screened by the 

planning group during a group meeting in Kiel in February 2008 based on quality and 

complementarity of the proposed research. The principle investigators (PI) of all selected preproposals 

were invited to participate in a workshop and at the same time informed about the 

tentative budget allocated to their project contributions, as recommended by the planning group. 

A 2-day workshop of all invited PIs was held in Kiel in April 2008, during which the consortium 

was founded and the structure of the programme, including the themes, projects and sub-projects 

were developed. During the workshop theme and project leaders (see Tables 2 and 3) were 

chosen and responsibilities for the preparation of the full-proposal were assigned. It followed a 

10-week period of intense communication and writing activities by all partners involved. The full 

proposal was completed in June and proudly handed over to the PTJ on July 2, 2008. 

Time line of consortium building and proposal preparation: 

(1) Presentation of project idea at PTJ Warnemünde May 2007 

(2) Planning group established July 2007 

(3) Planning group finalizes core proposal Nov. 2007 

(4) Presentation of project idea to BMBF and PTJ in Bonn Dec. 2007 

(5) Call for project contributions to potential partner universities/institutes Jan. 

(deadline for submission: February 15, 2008) 

2008 

(6) Screening of proposed contributions based on quality and 

complementarity, invitation of project partners 

Febr. 2008


(7) joint workshop of all partners, revision of core proposal, Apr. 2008 

development and structuring of themes, projects, and sub-projects 

(8) preparation of full proposal by all project partners April - June 2008 

(9) submission of full proposal July 2008 

5. Overview of Themes 1-5 

5.1. Theme 1: Primary production, microbial processes and 

biogeochemical feedbacks 

5.1.1. Theme summary 

Overarching questions 

1. How do marine primary producers and heterotrophic bacteria of diverse taxonomic 

groups respond to ocean acidification (OA) and increase in CO2 concentration on? 

Which groups/species (e.g. calcifying vs. non calcifying species) are negatively impacted 

and which benefit? 

2. To what extent will key phytoplankton species and bacteria be able to acclimate to OA? 

What is the potential for evolutionary adaptation and concomitant genetic changes 

within 100s to 1000s of generations? 

3. What are the consequences of ocean acidification for the turn-over and export of 

organic matter? 

4. What are the combined effects of CO2 and temperature changes on the marine soft 

tissue pump and DOC export and the air-sea exchange of CO2? 

Primary producers in the marine realm encompass phylogenetically very diverse groups 

(Falkowski et al. 2004), differing widely in their photosynthetic apparatus and carbon enrichment 

systems (Giordano et al. 2005). Preliminary data reveal that species with effective carbon 

concentration mechanisms (CCMs) are less sensitive to increased CO2 levels than those lacking 

efficient CCMs, analogous to findings in terrestrial vegetation. Currently, our ignorance of the 

metabolic diversity of oceanic autotrophy and microbial heterotrophs hampers any projection of 

total marine primary production and regeneration in response to increased carbonation. A focus 

of Theme 1 will therefore be to identify critical physiological traits that determine the sensitivity 

of key groups of primary producers and bacteria to increases ocean acidification and carbonation. 

Most of the biological oceanic carbon uptake is driven by regenerated production and only ~20% 

by new nitrogen input to surface waters (Laws et al., 2000) which drives the export of organic 

carbon from surface waters (Eppley and Peterson 1979). While the export of carbon, nitrogen, 

and phosphorus is generally considered to be closely coupled in marine biogeochemical cycling, 

recent work in mesocosms shows increasing C:N and C:P ratios during primary production with 

increasing CO2 concentrations (Riebesell et al., 2007). Excess carbon assimilation partially ends 

up as dissolved organic carbon and may increase the export of organic matter through the 

formation of gel particles that enhance particles aggregation, such as transparent exopolymer 

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particles (Arrigo, 2007; Engel et al., 2004). These processes, if representative for pelagic 

autotrophic communities, could give rise to a biologically driven feedback to the climate system. 

What controls the release of dissolved organic matter by either phytoplankton or bacteria, and 

how these substances affect the nutrition and aggregation of pelagic organisms needs further 

investigation in order to improve the description of biogeochemical turnover processes and their 

sensitivities to increasing pCO2 (Figure 4). 

Theme 1 will study plankton communities in controlled lab experiments on several relevant time 

scales, from short-term physiological adjustment, to acclimation, to longer-lasting evolutionary 

adaptation. Treatment levels, experimental set-ups and response variables were chosen 

concordantly among diverse target groups in order to ensure full comparability of results in the 

subprojects. 

Fig. 4: Compartments and interactions studied in Theme 1 

Theme 1 will combine laboratory-based work with field campaigns to assess the responses of 

natural plankton communities. Given the importance of coastal areas to fisheries and other 

marine resources and services, the coastal ecosystems constitute an important target region. Insitu 

experiments and sampling will be carried out in the Baltic Sea, an enclosed sea with high 

nutrient input and anthropogenic pressures.


An improved implementation of possible impacts of ocean acidification and sea surface warming 

influence on the marine soft-tissue pump and the cycling (and export) of dissolved organic 

carbon in global marine carbon cycle models, like the model PICES, is urgently needed. As part 

of the modeling component of Theme 1 new parameterizations will be incorporated based on 

empirical results generated under this theme. 

5.1.2. Progress expected 

Theme 1 aims at a better understanding of complex species interactions in communities 

comprising primary producers and bacteria and associated biogeochemical dynamics under 

present pCO2 concentrations, twice (projected for 2100) and three times the present 

concentration. Laboratory and field experiments will deliver insight into the sensitivity of key 

plankton species to high CO2 concentrations. Synergistic effects due to temperature increase and 

changes in nutrient concentrations will be considered in some experimental setups. 

Expected changes in the formation, sedimentation, and export of biogenic particles due to 

changing mineral ballast will be assessed by means of experiments in pressure controlled 

chambers. This will be complemented by determining CO2 effects on the regulation of DOM 

release from primary producers and its impact on the bacterial community. 

Acclimation will be described in terms of physiological responses. The genomic changes in 

response to long-term exposure will be investigated to identify the time scales on which 

adaptations can be achieved. 

Data on responses of organisms will be built into models. This involves (1) an improvement of 

the description of the soft-tissue pump in a global biogeochemical model and (2) integrating 

ocean acidification sensitivities at the organism level into ecosystem modelling. 

5.1.3. Projects under this Theme 

Project 1.1: Acclimation versus adaptation in autotrophs (Thorsten Reusch) 

Project 1.2: Turnover of organic matter (Anja Engel) 

Project 1.3: Modelling biogeochemical feedbacks of the organic carbon pump (Birgit 

Schneider) 

References 

Arrigo KR, (2007) Carbon Cycle Marine manipulations, Nature, 450: 491-492. 

Engel A, Thoms S, Riebesell U, Rochelle-Newall E,Zondervan I, (2004) Polysaccharide aggregation as a potential sink of marine dissolved 

organic carbon, Nature, 428: 929-932. 

Eppley RW, Peterson BJ, (1979) Particulate organic matter flux and planktonic new production in the deep ocean, Nature, 282: 677-680. 

Falkowski P, Barber RT, Smetacek V, (1998) Biogeochemical Controls and Feedbacks on Ocean Primary Production, Science, 281,(5374): 

200-205; DOI: 10.1126/science.281.5374.200. 

Giordano M, Beardall J, Raven JA (2005) CO2 concentrating mechanisms in algae: mechanisms, environmental modulation, and evolution. 

Annu Rev Plant Biol 56: 99-131 

Laws EA, Falkowski PG, Smith WO, Ducklow H, McCarthy JJ, (2000) Temperature effects on export production in the open ocean, Global 

Biogeochem. Cycles, 14,(4): 1231-1246. 

Riebesell U, Schulz KG, Bellerby RGJ, Botros M, Fritsche P, Meyerhöfer M, Neill C, Nondal G, Oschlies A, Wohlers J, Zöllner E, (2007) 

Enhanced biological carbon consumption in a high CO2 ocean, Nature 450: 545-548 

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5.2. Theme 2 Performance characters: Reproduction, growth and 

behaviours in animal species 



1. Which physiological mechanisms define sensitivity or tolerance of marine 

animals to ocean acidification and how do they set or modify performance levels 

and fitness? 

2. Can acclimation capacity (gene expression capacity) for such mechanisms 

explain physiological plasticity? 

3. How does acclimation or adaptation to new levels of CO2 and temperature 

affect organism performance? 

4. In a comparison of species and their populations from temperate to polar 

climates, do they differ in their sensitivity or capacity to resist ocean 

acidification through acclimation or evolutionary adaptation? How do these 

findings relate to differences in temperature and associated ocean 

physicochemistry? 

5. Which life stages of functionally important marine organisms are most sensitive 

to ocean acidification and how does the level of sensitivity relate to the ontogeny 

of physiological mechanisms? 

Ecosystem effects of ocean acidification include those on metazoan life. However, while 

ecosystem effects of warming trends have clearly been identified, those of ocean acidification are 

still equivocal. Within the next decades, elevated CO2 levels are expected to affect marine water 

breathing animals directly through effects on the physiology and performance of the individual 

organism and indirectly through changes in food web structure. Emerging knowledge indicates 

that sensitivity to elevated CO2 levels differs between animal phyla and species. It may also differ 

depending on geographical latitude and associated climate conditions. Effects may be large and 

potentially detrimental especially in life forms with a low metabolic rate, for examples among 

calcifying benthic macroorganisms (Wood et al., 2008) or in the deep sea. This hypothesis is in 

line with recent observations in habitats contaminated by natural CO2 emissions, e.g. in volcanic 

areas around Ischia (Hall-Spencer et al., 2008). Initial findings suggest decreased growth and 

enhanced mortality of sensitive species such as among molluscs or echinoderms in response to a 

doubling of CO2 from pre-industrial levels to 560 ppm (Shirayama and Thornton 2005), a value 

which is likely surpassed during this century. As effects of ocean acidification are expected on 

top of those of ocean warming, studying ecosystem effects of OA will thus need to consider both, 

the direct influence of ocean physicochemistry on individual organisms and species, and also the 

CO2 dependent modulation of responses to temperature in particular. 

For an in-depth cause and effect understanding, it is essential to unravel the physiological 

mechanisms that define whole organism sensitivity to ocean acidification (e.g. Pörtner et al. 

2004, Fabry et al. 2008) and especially those, which synergistically interact with temperature 

effects on marine organisms (cf. Pörtner et al. 2005). A current hypothesis emphasizes a key role 

for the capacity of acid-base regulation in defining sensitivity (Pörtner, 2008 for review).


Available data indicate that deviations of extracellular pH from its setpoint mediate several of the 

observed whole organism effects. Theme 2 will investigate how and to what extent these 

disturbances affect whole animal performance and how acclimation to various CO2 levels can 

alleviate some of these effects. It will also address to what extent sensitivity to ocean 

acidification interacts with thermal stresses and is shaped by the specialization of organisms on 

ambient climate conditions according to latitude. 

Fig. 5: Effects of ocean acidification and warming on marine ectotherms 

– Theme 2 concepts and their implementation 

Effects of anthropogenic ocean acidification on animal communities are expected on medium to 

long time scales, due to the progressive accumulation of CO2 and due to long generation times. In 

variable environments (e.g. upwelling areas, Feely et al., 2008) not only the drift in mean 

physicochemical parameters but also enhanced amplitudes will require consideration in analyses 

of CO2 effects. For an analysis of the interaction between specialization on various climates on 

the one hand and sensitivity to ocean acidification on the other hand, physiological studies (of 

e.g. performance and acid-base regulation) as well as investigations of gene expression patterns 

and population structure will be carried out in species and populations living in a climate gradient 

across latitudinal clines. Comparisons of fertilized eggs, juvenile and adult life stages will be 

essential to identify the bottlenecks of sensitivity throughout ontogeny as well as their 

physiological background. The physiological principles shaping performance may also control 

calcification, and thus the shell growth of bivalves and other calcifiers over time. Performance 

has been shown to link climate change to ecosystem effects of warming (Pörtner and Knust 

2007). Performance characters like growth or foraging capacity are likely also involved in multistep 

processes affecting marine food webs. Here, species-specific responses and sensitivities 

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cause various species of an ecosystem to be affected differently, resulting in changes in species 

interactions, population adaptability, food web structure and associated carbon fluxes. The work 

will include species of coastal areas and relevant to fisheries and other marine services. Theme 2 

also includes approaches to test and further develop mechanistic concepts and models of effects 

of ocean acidification. This includes kinetic modelling of the mechanisms of ion and acid-base 

regulation for an improved quantitative understanding of effects, to generate a basis for a more 

comprehensive, mechanism based modelling approach. 


Theme 2 aims at a better understanding of the physiological underpinning of specific and 

combined CO2 and temperature effects on marine water breathing animals and on ecosystem 

function. Lab and mesocosm experiments will provide insight into specific sensitivities, 

acclimation and adaptation of marine invertebrates and fish to CO2 and temperature under present 

CO2 concentrations as well as under levels twice (as projected for 2100) and three times the 

present concentration. We expect to complement our knowledge of the physiological mechanisms 

responding to changing CO2 levels and to changing body fluid and tissue acid-base variables, 

considering the specific patterns of tissue functioning and its regulation. As a result, a more 

comprehensive picture is expected of the mechanisms involved in shaping sensitivity, 

acclimation and adaptation to various CO2 accumulation scenarios. Furthermore, consideration of 

the projected temperature increase in our experiments will lead to a better understanding of the 

synergistic effects of warming and acidification as well as the underlying mechanisms. The 

generalized view of physiological principles should also help to identify the master variables 

mediating the effects of ocean acidification (such as e.g. extracellular pH) and to comprehend 

their integrating role at various levels of biological organization. 

Mechanisms contributing to the set-points of acid-base regulation will be identified and their 

contribution quantified in experimental and kinetic modelling studies. Acclimation will be 

described in terms of the longer term adjustments in the functioning of mechanisms and 

processes as well as in terms of the underlying genomic changes involving transcriptional and 

translational processes. The time scales of acclimation and adaptation should become accessible 

from long-term exposures and from comparisons of species and populations in various 

temperatures and thus CO2 concentration regimes along a latitudinal cline between temperate and 

polar areas. 

Insight in the responses at organism level throughout ontogeny as well as at population level will 

build the basis for identifying the CO2 and temperature induced modulation of key processes such 

as recruitment and calcification which are crucial for a population’s role in ecosystem 

functioning. The compilation of CO2 and temperature impacts on animal performance at different 

trophic levels, e.g. from herbivores to top predators will facilitate the development of scenarios of 

future marine ecosystem functioning. Such integrated assessments of sensitivities to ocean 

acidification would also become available for future ecosystem modelling. 


Project 2.1: Effects on grazers and filtrators (Thomas Brey) 

Project 2.2: Long-term physiological effects on different life stages of benthic crustaceans 

(Felix Mark) 

Project 2.3: Effects on top predators (fishes, cephalopods) (Catriona Clemmesen)


References 

Fabry VJ, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES J Mar Sci 65: 

414–432. 

Feely RA, Sabine CL, Hernandez-Ayon JM, Ianson D, Hales B (2008) Evidence for upwelling of corrosive "acidified" water onto the 

continental shelf. Science 320: 1490 – 1492. 

Hall-Spencer J.M., Rodolfo-Metalpa R, Martin S, Ransome E, Fine M, Turner SM, Rowley SJ, Tedesco D, Buia MC (2008) Volcanic carbon 

dioxide vents show ecosystem effects of ocean acidification. Nature, doi:10.1038/nature07051. 

Pörtner HO (2008) Ecosystem effects of ocean acidification in times of ocean warming: a physiologist’s view. Mar. Ecol. Progr. Ser. in review 

Pörtner HO, Langenbuch M, Reipschläger A (2004) Biological impact of elevated ocean CO2 concentrations: lessons from animal physiology 

and earth history. J. Oceanogr. 60: 705-718. 

Pörtner HO, Langenbuch M and Michaelidis B (2005) Synergistic effects of temperature extremes, hypoxia, and increases in CO2 on marine 

animals: From Earth history to global change, J. Geophys. Res. 110: C09S10 

Pörtner HO, and Knust R (2007) Climate change affects marine fishes through the oxygen limitation of thermal tolerance. Science 315: 95 - 

97. 

Shirayama, Y., and H. Thornton (2005), Effect of increased atmospheric CO2 on shallow water marine benthos, J. Geophys. Res., 110, 

C09S08, doi:10.1029/2004JC002618. 

Wood HL, Spicer JI, Widdicombe S (2008) Ocean acidification may increase calcification rates, but at a cost. Proc R Soc B 275:1767-1773. 

5.3. Theme 3 Calcification: Sensitivities across phyla and ecosystems 



1. What are the cellular mechanisms of calcification and decalcification in different marine 

organisms, particularly with regard to ion transport to and from calcification sites? 

2. How will OA and pH stress affect calcification at the level of organisms, communities 

and ecosystems? Will concurrent temperature change modulate these effects? 

3. What changes will occur in the ultra-structure, trace element partitioning, and isotopic 

signature of the shells and skeletons of calcifyers in response to pH stress? 

4. How does the changing water column chemistry influence carbonate dissolution and 

deposition in sedimentary systems? 

5. Are past OA events (e.g. in the Cenozoic) useful analogues for projected future OA in 

their effects on calcifying organisms? Is the performance and sensitivity of past and 

present calcifyers comparable, and does this information help to assess the future? 

Calcification, the precipitation of calcium with carbonate, is influenced by the current increase in 

atmospheric CO2 levels and its concurrent gradual decrease in ocean pH. Since acidification leads 

to a decrease of the carbonate ion concentration, ocean acidification causes a decrease of the 

carbonate saturation sate. Hence we can expect a future reduction in calcification rates, or even 

net decalcification. Indeed reduction of calcification rates have been observed upon acidification 

in several marine calcifying groups, but not in all. This is indicative for a variety of calcification 

mechanisms. Calcification is a proton-generating process, and decalcification is proton 

consuming. Thus, these processes are typically coupled to either proton consuming (calcification) 

or proton producing (decalcification) processes. Conversely, calcification and decalcification can 

buffer other pH changing processes, and will thus to some extend buffer the oceanic pH. 

Biological calcification always occurs in more or less isolated microenvironments, in which the 

carbonate and/or calcium concentrations are changed by biological activity. These changes can 

be driven by ion pumps in specialized transport tissue, e.g. for Ca 2+ or H + , which is typical for the 

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highly controlled calcification in corals, foraminifera, bivalves and coccolithophores. 

Alternatively, calcification can be a side effect of metabolic activities such as photosynthesis, 

occurring in a matrix with mass transfer limitation (a sediment or microbial mat). We will 

investigate if some calcification mechanisms are more sensitive than others, and the extent to 

which decalcification can buffer the decrease in oceanic pH. We will further investigate if 

decalcification can contribute to pH buffering of the oceans. We will finally investigate if the 

oceanic pH may leave signatures in the biogenic carbonates, and learn from acidic events in the 

past what the effect is on biodiversity on calcifying nano-plankton. 

Differences in calcification mechanisms 

may be responsible for observed 

differences in CO2/pH sensitivities of 

marine calcifiers. Left is a possible 

calcification scheme by actively 

calcifying organisms, which depend on 

transmembrane pumping of ions to and 

from a calcification site, such as shells 

and skeletons. Right is a scheme of 

calcification by diffusion-controlled 

calcification. A calcifying microenvironment 

is created by the combined 

effect of diffusion resistance and 

metabolic activity, like photosynthesis. 

Fig. 6: Groups of calcifying organisms and processes studied in Theme 3 


Within this theme we will try to elucidate how pH affects calcification rates of two principally 

different mechanisms. We will study (1) calcification driven by membrane transport processes, 

i.e. fully organism-controlled, and (2) calcification as a side reaction of photosynthesis, i.e. 

environmentally controlled. In both cases the calcification occurs in a more or less shielded 

space, but in the first case it is shielded by membranes and tissue within the calcifying organisms, 

in the second case calcification is rather an encrustation outside the organism, and the transport 

limitation is by diffusion.


Hypotheses: As the first mechanism involves considerable energy input from the actively 

calcifying organisms, which will increase upon acidification, we expect significant effects on the 

ecological fitness of calcifiers and on the biodiversity of ecosystems with calcifying 

communities. In the second case calcification does not require metabolic energy, and reduced 

calcification does not present an ecological disadvantage for the organisms that drive 

calcification, and will not have an effect on their biodiversity. We expect to have a better 

understanding on this issue through the projects 3.1 and 3.2. Secondly, we will investigate if pH 

changes leave a signature in the biogenic carbonates, and whether the isotope and trace metal 

signatures can be used to better understand the calcification process. More specifically, it will be 

tested in project 3.3 whether calcification in corals and foraminifera is indeed a result of 

enveloping seawater parcels in vacuoles in which subsequently the chemistry is shifted towards 

an environment where carbonates precipitate. This will enhance understanding of the chemical 

signatures of biogenic carbonates. We will investigate in project 3.4 in how far calcification in 

sediments and other matrices is resilient against ocean acidification, and construct a diffusionconversion 

model describing calcification and decalcification. This model will include the 

carbonate system, pH, photosynthesis and respiration. In project 3.5 we will investigate if several 

acidic events in the past have left traces in the carbonate skeletons. The calcification driven by 

photosynthesis occurs in sediments and mats where transport is diffusionally limited. This to 

some extend uncouples the microenvironment from the seawater. 


Project 3.1: Cellular mechanisms of calcification (Frank Melzner) 

Project 3.2: Calcification under pH-stress: Impacts on ecosystem and organismal levels 

(Ralph Tollrian) 

Project 3.3: Ultra-structural changes and trace element / isotope partitioning in calcifying 

organisms (foraminifera, corals) (Jelle Bijma) 

Project 3.4: Micro-environmentally controlled (de-)calcification mechanisms 

(Michael Böttcher) 

Project 3.5: Impact of present and past ocean acidification on metabolism, 

biomineralization and biodiversity of pelagic and neritic calcifiers 

(Adrian Immenhauser) 

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5.4. Theme 4 Species interactions and community structure in a 




1. What is the role of differential sensitivities to OA at the community, species and intraspecific 

(ontogenetic stages, genotypes) level? Are there emerging properties resulting 

from organism interactions that are not visible from single-species investigations 

alone? 

2. Does the community structure change as a consequence of OA and how do shifts in 

competitive abilities of benthic and pelagic organisms affect community structure? 

3. What is the role of OA induced changes in food quality and quantity in primary 

producers to higher trophic levels? 

4. To which extent will energy transfer between lower and higher trophic levels change as 

a result of changing competitive interactions and/or changes in the feeding 

environment? 

5. Do different types of communities (benthic – pelagic, microbial – macrobial) react 

differently to acidification/warming stress? 

Observed effects of low pH and/or high CO2 conditions are mostly based on experiments with 

single species. Hence, not much is known about how these factors affect interactions between 

species and, through that, communities. Indeed, the projected shifts in pCO2 and pH in many 

species often only slightly impact performance and fitness of a given species, and many studies 

find reactions in single species experiments only with very high CO2 concentrations or at 

unrealistically low pH (e.g. Mayor et al. 2007). However, as shown for other stressors 

(Christensen et al. 2006) ensuing modifications of species interactions may substantially amplify 

or buffer the original stress (Wahl 2008). How environmental stress spreads through a 

community via shifts in composition and interaction is still very much an open question. 

Theme 4 will therefore address the pivotal role of interaction modulation in close cooperation 

with a number of projects from other themes which, in contrast, focus on single species reactions 

to ocean acidification. Thus, Theme 4 is the logical extension of these projects, placing the 

responses of individual organisms to OA into a community and ecosystem context. We expect to 

find effects of OA on community structure and interaction that are not visible when studying 

single organism reactions. Hence, we will focus on shifts in competitive and trophic interactions 

as well as on community structure. Organisms will vary considerably in their reaction to ocean 

acidification. As a consequence, formerly superior competitors may be weakened, as is the case 

in interactions between calcifying and non-calcifying species (e.g. Kuffner et al. 2007), or 

relative susceptibility to predation may change (Swanson & Fox 2007). Furthermore, there may 

be strong selective pressures within species, if susceptibility to stress differs between genotypes 

or between ontogenetic stages. At the unicellular level, less sensitive clones or strains may 

become more dominant under ocean acidification, and sensitive rare populations may disappear. 

In multicellular species, the most sensitive ontogenetic phase will determine survival, and genetic 

diversity may determine the fate of a population under OA.


The changes in species composition on one trophic level will obviously affect the transfer of 

energy and matter to higher trophic levels. Shifting species composition at the base of the food 

chain may represent different quality feeds for predators, which may result in a complete 

restructuring of the trophic web. Also, direct effects of increased CO2 availability may change the 

quality of organisms as food for higher trophic levels. Higher carbon availability will typically 

result in higher carbon to nutrient ratios in primary producers. This has on the one hand been 

linked to decreased toxicity of some dinoflagellates (Parkhill & Cembella 1999), with the 

potential of higher palatability of previously noxious algae, but on the other hand to a decrease in 

the quality as food for higher trophic levels (Malzahn et al. 2007). 

So, if we are to understand the effects of OA on ecosystem structure and function, it is essential 

that we understand the shifts of interaction mode or strength (Tortell et al. 2002), because the 

typical non-linearity of the ensuing effects has the potential to cause regime shifts in marine 

ecosystems. To date, regime shifts have mainly been linked to climate forcing, but we expect 

ocean acidification – amplified by interaction modulation - to have the same potential. 

Approach: 

Fig. 7: Compartments and interactions studied in Theme 4 

In Theme 4, we will investigate how interactions between and within species in benthic [4.1.1.- 

4.1.4] and pelagic [4.2.1-4.2.2] communities shift under the influence of OA. We investigate 

changes in trophic grazer- alga interactions in the benthos [4.1.1], and competitive interactions 

between sessile organisms [animals 4.1.2 and plants 4.1.3]. Moreover, we will investigate the 

effects of ocean acidification on bacterial communities, both directly, as well as a result of altered 

excretion products of algae and herbivores faced with resources of different quality [4.1.4 for the 

benthos, 4.2.1 for the pelagic zone]. Competition between pelagic microalgae will be studied in 

4.2.2, and the resulting prey community will be fed to pelagic herbivorous grazers in 4.2.1 

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Theme 4 will further our understanding of between- and within-species interactions under OA 

stress. We expect that after a successful completion of the project we will be able to define the 

expected impacts of ocean acidification that go beyond single species reactions but are a result of 

changing interactions between and within species. As stated above, only by taking these 

interactions into account we will be able to truly appreciate the potential impact of OA. More 

specifically we expect progress in the following areas: 

1. Assessment of the role of differential sensitivities to OA at the intra-species (ontogenetic 

stages, genotypes), and species level. Populations and communities can only re-structure and, 

thus, adapt if their respective components differ in sensitivity towards the stressor(s). In close 

cooperation with appropriate projects in the other themes we will quantify how the responses 

to stress varies within species between life stages (not applicable for unicellular organisms), 

genotypes/strains, and between species in the focal communities. 

2. Shifts in competitive abilities of model organisms. Co-culturing of organisms exhibiting 

differing stress sensitivity (at the ontogenetic, genetic, species level) allows the quantification 

of the stress-induced shifts in competitiveness as compared to a low-stress situation. Strong 

shifts will lead to fast changes in relative abundance of the components in the focal 

communities: benthic and pelagic microbes, microalgae, macroalgae, macroalgae/sea grass, 

sessile invertebrates. This issue will highlight the role of genetic and functional diversity for 

the adaptability of populations and communities. Further, prolonged co-culturing of simple 

communities in micro- and mesocosms will produce re-structured communities adapted to 

the OA scenario applied. Since such a re-organization depends on the gene and species pool 

available, and on the number of generations allowed, and both these conditions can not be 

simulated realistically in experiments we do not pretend to simulate the ecosystem changes 

which will happen in the future. However, the kind of shifts among functional groups – e.g. 

macroalgae vs. seagrass or calcifiers vs. non-calcifiers – will improve our capacity for 

plausible projections. 

3. Quantification of the changes in community structure and community services. The structural 

shifts driven by OA will lead to changes in the functions within communities. Thus, an 

altered prey community will differ with regard to food quality and quantity. How this affects 

structure and dynamics of primary or secondary consumers, and storage in or flow of 

energy/matter between different strata of the system, will be a further outcome of Theme 4. 

Ultimately, we aim to make predictions of how OA is likely to affect issues such as surface 

water quality or the yield of economically relevant species. 

4. We will strive to incorporate the findings on direct and indirect OA impacts at the different 

organisational into descriptive and/or predictive models. 


Project 4.1: OA impacts on interactions in and structure of benthic communities 

(Martin Wahl) 

Project 4.2: OA effects on food webs and competitive interactions in pelagic ecosystems 

(Maarten Boersma)


References 

Christensen MR, Graham MD, Vinebrooke RD, Findlay DL, Paterson MJ, Turner MA (2006) Multiple anthropogenic stressors cause 

ecological surprises in boreal lakes. Glob Chan Biol 12:2316-2322 

Kuffner IB, Andersson AJ, Jokiel PL, Rodgers KS, Mackenzie FT (2007) Decreased abundance of crustose coralline algae due to ocean 

acidification. Nature Geosci 1:114-117 

Malzahn AM, Aberle N, Clemmesen C, Boersma M (2007) Nutrient limitation of primary producers affects planktivorous fish condition. 

Limnol Oceanogr 52:2062-2071 

Mayor DJ, Matthews C, Cook K, Zuur AF, Hay S (2007) CO2-induced acidification affects hatching success in Calanus finmarchicus. Mar 

Ecol Prog Ser 350:91-97 

Parkhill J, Cembella A (1999) Effects of salinity, light and inorganic nitrogen on growth and toxigenicity of the marine dinoflagellate 

Alexandrium tamarense from northeastern Canada. J Plankton Res 21:939-955 

Swanson AK, Fox CH (2007) Altered kelp (Laminariales) phlorotannins and growth under elevated carbon dioxide and ultraviolet-B 

treatments can influence associated intertidal food webs. Glob Chan Biol 13:1696-1709 

Tortell PD, DiTullio GR, Sigman DM, Morel FMM (2002) CO2 effects on taxonomic composition and nutrient utilization in an Equatorial 

Pacific phytoplankton assemblage. Mar Ecol Prog Ser 236:37-43 

Wahl M (2008) Ecological modulation of environmental stress: interactions between UV radiation, epibiotic snail embryos, plants and 

herbivores. J Anim Ecol 77:549-557 

5.5. Theme 5 Integrated assessment: Sensitivities and uncertainties 


Objectives and overarching questions 

1. Synthesize information obtained in Themes 1 to 4 in order to achieve an integrated 

understanding of biological responses to ocean change. 

2. Develop a framework for integrating ocean acidification sensitivities at the organism 

level into ecosystem models. 

3. What are the integrated effects of ocean acidification and warming on ecosystem to 

global ocean scales? 

4. What are the critical threshold levels (‘tipping points’) of ocean acidification for 

irreversible ecosystem changes? 

5. What is the most suitable definition of dangerous ocean acidification in terms of the 

goods and ecosystem services lost due to OA? 


German Advisory Council on Global Change (WBGU, Berlin 2006) recommends a guard rail for 

future ocean pH decrease of 0.2 units as a margin of safety according to the precautionary 

principle. This suggestion is motivated by the intention of avoiding an aragonite undersaturation 

in the ocean surface layer. As stated in the report, the tolerable window for ocean acidification 

defined by WBGU presently relies on an extremely small data base. In fact, rather than using the 

limited data on observed biological consequences of ocean acidification, the WBGU reaches its 

recommendation on the basis of projected changes in water chemistry (aragonite saturation state). 

While this is an appropriate approach in view of the scarcity of biological information, there is a 

clear need to establish a reliable data base on tolerance levels for ocean acidification in key 

groups of ocean-acidification sensitive marine organisms in order to reach a more informed 

recommendation. 

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Fig. 8: Schematic diagram of Theme 5. Proposed projects are indicated (5.1, 5.2, 5.3). 

Theme 5 of BIOACID will take the challenge of integrating the information gained under 

Themes 1 to 4 in order to identify the potential thresholds associated with ocean acidification. 

Uncertainties, probabilities and risks to the marine environment have to be assessed as well as 

their feedback to climate system. This will be achieved through a meta-analysis of process 

studies and process parameterisations, and by combining models and data in a data-assimilative 

framework. In return, feedback from the modelling work will inform the experimental work in 

BIOACID about uncertainties in models and the relevant process parameterisations. 

During the first 3-year phase of BIOACID, our main aim is to develop and establish the tools that 

will allow us to fulfil the BIOACID synthesis needs. For the three subprojects proposed here, the 

synthesis tools to be established within BIOACID range from meta-analysis techniques over 

regional and global numerical ecosystem models to economic methods of integrated assessment. 

These tools will help to better understand ongoing changes in chemical and biological state of the 

North Sea from alkalinity fluxes originating from the Wadden Sea over a synthesis model that 

integrates OA sensitivities at organism level into a North Sea ecosystem model (5.1) to an 

economical impact assessment. (5.3). Newly developed assessment tools will also be used to 

improve parameterisations of calcium carbonate production in global biogeochemical climate 

models (5.2). By investigating the combined effects of variations in temperature and ocean 

acidity, such parameterisations will allow to put better constraints on possible threshold levels on 

ocean acidification in a warming world. 


We subdivide expected progress into three categories, development of synthesis tools, synthesis 

of research results, and large scale modelling ocean acidification impacts and biogeochemical


feedbacks to carbon cycling and climate. Categories which reflect the major research aspects 

covered by projects of Theme 5 as well as collaborative research carried out together with 

projects from Themes 1-4 (see table 0.1, Section 11.1 for details). 

Development of synthesis tools 

This theme will 

• carry out supplementary model development and evaluation of a shelf sea ecosystem 

model (ECOHAM) for studying ocean acidification effects on the ecosystem level as 

well as alkalinity exchange of the North Sea with Wadden Sea and open ocean, 

• implement proposed parameterization of open ocean calcium carbonate production, 

export and dissolution into a tracer transport matrix rapid spin-up model system, 

• initiate a Bayesian meta-analysis of BIOACID experimental findings 

• further develop the ecological-economic viability-method towards a general approach for 

integrated assessment of human actions influencing ocean acidification and the 

consequences for human well-being that takes uncertainties about future development 

into account 

Synthesis of research results 


• critically review existing parameterisations of calcification currently used in large-scale 

biogeochemical climate models, 

• provide a review of experimental findings on temperature and pCO2 sensitivity of 

calcium carbonate production, its export and dissolution to be put in perspective with 

model parameterisations. 

Large scale modelling ocean acidification impacts and biogeochemical feedbacks to carbon 

cycling and climate 


• provide an evaluation and quantification of alkalinity fluxes from the Wadden Sea to the 

North Sea and their role in buffering ocean acidification from anthropogenic CO2 

invasion. Response of calcifying and non-calcifying primary producers to pH changes 

will be addressed by extrapolating results from mesocosm studies and future scenario 

model simulations (pCO2=1000µatm) studying critical ecological and biogeochemical 

aspects, 

• quantitatively assess the ability of current parameterisations of calcification in global 

biogeochemical models to reproduce observed alkalinity fields, and to suggest improved 

parameterisations that can reliably predict the response of pelagic calcium carbonate 

production to variations in both temperature and ocean carbonate chemistry 

• by means of the viability-method, assess the impacts of different human actions on ocean 

acidification, taking into account the uncertainties about the future development of 

acidification and the exact impact on cod recruitment. This modelling approach will take 

into account measures to mitigate acidification or to adapt the management of the North 

Sea cod fishery. 

27

28 



Project 5.1: Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle and the 

primary production in the North Sea, (Johannes Pätsch) 

Project 5.2: Evaluating and optimising parameterisations of pelagic calcium carbonate 

production in global biogeochemical ocean models, (Andreas Oschlies) 

Project 5.3: Viability-method for the impact assessment of ocean acidification under 

uncertainty (Martin Quaas) 

6. Management structure and procedures 

A good organizational structure and an effective labour division based on clearly identified tasks 

and accompanied with a well-defined decision-making system will be put in place in order to 

fulfil the scientific objectives of BIOACID, to ensure an efficient work-flow and to reduce risks 

of failure. An overall organizational chart depicting the communication links is depicted in 

Figure 9 and is described in further detail below. The management procedures clearly define the 

consortium tasks, responsibilities and lines of command in a transparent decision-making 

structure. 

Fig. 9: Management structure and executive and advisory bodies.


6.1. Project Coordinator and Project Office 

The Coordinator of BIOACID will be Ulf Riebesell at the Leibniz Institute of Marine Sciences, 

Kiel. Hans-Otto Pörtner at the Alfred-Wegener Institute, Bremerhaven will act as Deputy 

Coordinator. The coordinator (or in the case of his absence the Deputy Coordinator) will be 

responsible for the day-to-day management of BIOACID and contact with the Ministry of 

Education and Research (BMBF) and the Projektträger Jülich (PTJ). He will ensure that the 

project runs in compliance with the requirements of the contract (administrative, operational and 

scientific aspects). The Project Management Office will be located at IFM-GEOMAR. It will 

act upon decisions taken by the Executive Board (EB), the Scientific Steering Committee 

(SSC) and the Member’s General Assembly (MGA). 

6.2. Executive Board (EB) 

The Executive Board (Table 2), chaired by the Project Coordinator, will prepare decisions to be 

approved by the SSC and MGA and implement decisions on executive management. It will meet 

annually at the consortium meeting and will communicate regularly by phone and/or e-mail. 

Major decisions and recommendations will be made at the meetings with one vote per member of 

the EB. In addition to the meetings, the Executive Board will communicate via the project 

website. All communication with other Committees will be through the Project Office. 

Table 2: Composition of the executive board. 

Executive board Members 

Coordinator: Ulf Riebesell (IFM-GEOMAR) 

Theme leaders: Maren Voss (IOW), Hans-Otto Pörtner (AWI), Dirk de Beer (MPI Bremen), 

Maarten Boersma (AWI), Andreas Oschlies (IFM-GEOMAR) 

The responsibility of the EB will include: 

• ensuring that sufficient management is in place for monitoring the scientific work in 

project and perform quality control 

• enforcing decisions of the SSC and MGA 

• ensuring the preparation of reports and work plans 

• informing the SSC about project progress, any problems and risks encountered and any 

change in strategy 

6.3. Scientific Steering Committee (SSC) 

The SSC (Table 3) will advise on the overall scientific policy, direction and management of the 

project to be decided by the MGA. It will meet annually at Consortium meetings and be chaired 

by the Coordinator. All communication with the MGA will normally be through the Project 

Office. 

29

30 


Table 3: Composition of the scientific steering committee (SSC). 

SSC Members 

Coordinator: Ulf Riebesell (IFM-GEOMAR) 

Deputy coordinator: Hans-Otto Pörtner (AWI) 

Theme leaders: Maren Voss (IOW), Hans-Otto Pörtner (AWI), Dirk de Beer (MPI 

Bremen), Maarten Boersma (AWI), Andreas Oschlies (IFM-GEOMAR) 

Project leaders: Michael Diepenbroek (U Bremen), Magnus Lucassen (AWI), Thorsten 

Reusch (IFM-GEOMAR), Anja Engel (AWI), Birgit Schneider (CAU 

Kiel), Thomas Brey (AWI), Felix Mark (AWI), Catriona Clemmesen 

(IFM-GEOMAR), Frank Melzner (IFM-GEOMAR), Ralph Tollrian (U 

Bochum), Jelle Bijma (AWI), Martin Wahl (IFM-GEOMAR), Maarten 

Boersma (AWI), Johannes Pätsch (U Hamburg), Andreas Oschlies (IFM- 

GEOMAR), Martin Quaas (CAU Kiel) 

Additional member: Antje Boetius (MPI Bremen), as member of the original planning group 

The responsibilities of the SSC will include: 

• acting on the initiative of the Executive Board (EB) on issues or problems relating to the 

progress towards the fulfilment of the scientific objectives 

• assessing scientific progress against the objectives and, where necessary, make 

recommendations to the EB 

• providing advice on any call for participants or partners that might be needed to finalize 

the project’s objectives 

• giving recommendations to the Executive Board on any scientific aspects it foresees as 

requiring ethical considerations 

6.4. Members’ General Assembly (MGA). 

The MGA consists of all project and sub-project PIs and will be the overall decisive body of the 

Consortium. At annual meetings, it will give scientific advice to the Steering Committee on the 

most important issues of the project. If needed, extraordinary meetings will be held. The MGA 

will be responsible for all major formal decisions regarding project strategy and any change to 

the consortium. Decisions in the MGA need two thirds of the votes present at the meeting. The 

MGA will act upon proposals from the Executive Board and decide on the following issues: 

• any major change in the scientific plans 

• any major budget reallocation between partners 

• any alteration of the management structure and management procedures 

• any change in the advisory bodies


7. Data management and dissemination 

7.1. Data management 

The data management in BIOACID will be carried out by the World Data Center for Marine 

Environmental Sciences WDC-MARE (www.wdc-mare.org) at the University of Bremen. WDC- 

MARE uses the information system PANGAEA (Publishing Network for Geoscientific & 

Environmental Data – www.pangaea.de), which is a system for acquisition, processing, long-term 

storage, and publication of geo-referenced data related to all earth science fields. Specifically, 

WDC-MARE will be responsible for 

1. Coordination of data capture, integration and quality control activities for the five 

BIOACID Themes. 

2. Archiving and publishing data sets and data collections online and as offline products 

(DVD) using persistent Digital Objects Identifiers (DOI). 

3. Implementation of the BIOACID data infrastructure 

- enabling a distributed storage of observational and model simulation data within a 

common networked structure, and 

- establishing a robust and long lasting data network which can be extended by or 

integrated into ongoing projects and programmes. 

4. Maintain the website and data portal for BIOACID. 

(For details on data managing see Theme 0.2 below). A half-time data manager position will be 

established by WDC-MARE at the University of Bremen. 

7.2 Dissemination 

Dissemination will include a multifunctional web service comprising a publicly accessible area to 

present the activities, techniques, and results of the project and a secure (password protected) area 

for internal project communication. This website will be maintained and administrated by the 

data manager (see project 0.2). 

Jointly with the administration of the Coordinator Project Office, all BIOACID partners 

contribute to 

• preparing a series of fact/information sheets and high level presentation packages aimed 

at policy makers’ needs 

• generate TV-quality video material on BIOACID research activities for distribution to 

interested TV channels and to produce a 8-10 minutes video clip on the BIOACID 

project and the emerging problem of ocean acidification (see project 0.1) 

• communicate with society through on-line debates, web site, media interviews, press 

releases, events and articles in popular magazines, and consortium consensus responses 

to frequently asked questions 

• scientific publications in international refereed journals and presentation at conferences, 

short reports and flyers to authorities, public bodies, agencies and organisations 

31

32 


Flyers and posters will be available on the BIOACID web site for members to download and 

distribute or display. Synergies with other national and EU projects (see 9 National and 

international Cooperations) will be formed to ensure effective exchange of information. 

8. Infrastructure development, training and transfer of know how 

8.1. Infrastructure development 

BIOACID will support the development of instrumentation and infrastructure urgently needed in 

ocean acidification research which will become of direct and general use to all BIOACID 

partners. In particular, two activities are proposed in this regard facilitating measurements of CO2 

partial pressure and improving CO2 perturbation-based experimentation. 

1. Development and maintenance of aquarium culture facilities and research infrastructure 

needed for long-term incubations in the form of a re-circulating system. The proposed 

system, to be developed and installed at the AWI in Bremerhaven, is expected to 

establish and maintain baseline conditions in larger experimental set-ups at pre-industrial 

levels and various CO2 scenarios (for details see project 0.3.1.). Time permitting the 

system will be open access to all BIOACID partners. 

2. Development of new chemical optical sensors and instrumentation for the determination 

of carbon dioxide partial pressure (pCO2) in fluids of marine organisms and in marine 

environments based on the dual lifetime referencing (DLR) method patented by PreSens. 

Emphasis will be put on the development of pCO2 sensors with appropriate resolution in 

the trace concentration range (380-1900 ppm in seawater, 380-3000 ppm in body fluids). 

The performance of the newly developed instrumentation will be optimised according to 

the experimental applications provided by project partners with prototype pCO2 sensors 

(for details see project 0.3.2.). 

8.2. Training and transfer of know how 

BIOACID will embrace a wide range of scientific disciplines, many with different approaches to 

monitoring the carbonate system and designing perturbation experiments. It is of vital importance 

for a large integrating project such as BIOACID to have common understanding on 

methodological and reporting procedures to facilitate comparison between investigators. 

Moreover, several BIOACID partners are international leaders for specific techniques which are 

of direct relevance to ocean acidification research. To this end, a series of training activities will 

be designed to ensure high data quality and inter-comparability and open up state-of-the-art 

technology to BIOACID partners. All workshops are intended to also facilitate the exchange and 

collaboration of young researchers employed in BIOACID and – space permitting – from outside 

the consortium. In this respect, close collaboration with interdisciplinary graduate schools such as 

the Integrated School for Ocean Sciences (ISOS) of the Excellence Cluster “The Future Ocean” 

and the Excellence Cluster “GLOMAR” (Global Change in the Ocean Realm) is envisioned. 

With over 30 Ph.D. positions and several Post-Doc positions offered in BIOACID, the project 

will also make a major contribution to the education of young scientists in a wide range of marine 

disciplines, from molecular biology, physiology, ecology, to biogeochemistry and 

palaeoceanography.

Table 4: Training workshops 


Organizer Institution Titel Duration 

Dirk de Beer MPI Bremen Microsensor applications 10 days 

Kai Schulz 

Anton 

Eisenhauer 

Martin Wahl 

IFM-GEOMAR 

IFM-GEOMAR 

IFM-GEOMAR 

Seawater carbonate chemistry 

Isotope geochemistry and laser-ablationtechniques 

Experimental design 

Franz Sartoris AWI Physiological approaches to body fluid 

physicochemistry and acid-base regulation 

9. International and National Cooperation 

2 days 

4 days 

3 days 

5 days 

German scientists, including members of the BIOACID planning group, have played a key role 

in recent reports or working groups related to ocean acidification such as: The Royal Society 

Working Group on Ocean Acidification "Ocean acidification due to increasing atmospheric 

carbon dioxide" (London 2005), the OSPAR intercessional working group on ocean acidification 

(2006), IPCC's fourth Assessment Report, Special IPCC report on Carbon Dioxide Capture and 

Storage (2006), Special Report of the German Advisory Council on Global Change "The Future 

Oceans – Warming Up, Rising High, Turning Sour" (2006), NSF-NOAA-USGS Report "Impacts 

of Ocean Acidification on Coral Reefs and Other Marine Calcifiers" (2006), the IGBP-SCOR 

Fast Track Initiative on “Ocean acidification, atmospheric CO2 and ocean biogeochemistry: 

modern observations and past experiences“(Lamont, 2006), and related to ocean biodiversity 

such as the DIVERSITAS Marine Biodiversity Cross Cutting Network and Census of Marine 

Life programmes. 

Members of the BIOACID planning group have also organized or co-organized meetings, 

workshops, special sessions or edited special issues of journals on ocean acidification. For 

example: The Ocean in a high CO2 World I and II, (I: Paris, May 2004, II: Monaco, October 

2008), ocean acidification sessions at the Ocean Sciences Meeting of the American Society of 

Limnology and Oceanography (ASLO) and at the annual conference of the European 

Geophysical Union (EGU) (Vienna, April 2006), a press conference on ocean acidification at the 

33

34 


EGU conference (Vienna, April 2006), a special issue of the Journal of Geophysical Research 

"The Ocean in a High-CO2 World" (2005) and a special issue of the journal Biogeosciences 

“PeECE: Pelagic ecosystem CO2 Enrichment Studies” (2007). Members of the BIOACID 

planning group were invited to attend and contribute to the planning of U.S. and UK national 

programmes on ocean acidification, and will remain to be involved in further developments of 

OA programmes in these countries. 

9.1. International Cooperation 

Close coordination of BIOACID activities with European research on ocean acidification will 

be ensured through high level participation of members of the BIOACID planning group in 

relevant EU projects, e.g. as co-coordinator, theme and work package leaders of the European 

Project on Ocean Acidification (EPOCA) and as leading scientists of the EU coordinated project 

Marine Carbon Sources and Sinks (CARBOOCEAN). As outlined by the coordinator of EPOCA, 

Dr. Jean-Pierre Gattuso (see p. 36): “The contribution from the European Commission (6.5 M€) 

is relatively limited for a 4-year project comprising 29 laboratories and more than 100 permanent 

researchers. Hence, its partners must complement the EU contribution with national funding in 

order to fulffill the objectives of the project and maintain the leadership that the EU currently has 

in the field of ocean acidification.” 

UK’s Natural Environment Research Council (NERC) has just launched its new five-year 

strategy which includes seven science themes and within one of these (Earth System Science) is a 

specific challenge “to understand changes in ocean ecosystems in response to ocean 

acidification”. During a preparation workshop of NERC’s five-year strategy, initial steps have 

been taken to aim for a UK-German partnership in developing a joint programme on ocean 

acidification (see letter by NERC Theme leader Prof. Tim Jickells, p. 37). The planned 

cooperation with UK NERC is consistent with BMBF policy as stated at the German-British 

conference on “Climate Change: Meeting the Challenge together” held in November 2004 in 

Berlin ending with a commitment to strengthening German-British cooperation in building up 

joint activities in climate research. 

9.2. National Cooperation 

BIOACID will be closely coordinated with National research programmes focussing on related 

aspect, in particular the DFG Priority Programme AQUASHIFT (The Response of Aquatic 

Ecosystems to Climate Change) and the BMBF Verbundprojekt SOPRAN (Surface Ocean 

processes in the Anthropocene). 

AQUASHIFT (coordinator: U. Sommer, IFM-GEOMAR, Kiel) assesses the effect of global 

warming on aquatic ecosystems. The anthropogenically enhanced greenhouse effect and the CO2induced 

ocean acidification are two sides of the same coin. Their impacts on marine ecosystems 

will go hand in hand, with a high probability of synergistic effects. Collaboration between 

AQUASHIFT and BIOACID opens the opportunity to address such synergistic effects, with the 

aim to allow a more realistic representation of future ocean conditions and the corresponding 

ecosystem responses.


SOPRAN (coordinator: D. Wallace, IFM-GEOMAR, Kiel) focuses on biogeochemical processes 

operating within and close to the surface ocean as drivers of ocean-atmosphere material 

exchanges. Particular attention is given to the impacts of climate induced changes in surface 

ocean processes (upwelling, mixing, light, nutrient supply) and of changes in atmospheric 

composition (e.g. increased CO2, dust) on surface ocean biogeochemistry and air-sea exchange of 

climatically relevant gases. Due to its focus on the organismal and community level, BIOACID 

will provide valuable input to SOPRAN. With its focus on a wide spectrum of global change 

forcings, SOPRAN will allow BIOACID to upscale its results to the ocean-wide level. 

Specifically, Theme 1 Primary production, microbial processes and biogeochemical feedbacks 

and Theme 5 Integrated assessment: Sensitivities and uncertainties of BIOACID will serve as 

cross-over with SOPRAN, facilitating cross fertilization between the two programmes and 

opening opportunities for joint investigations, particularly at the level of field observations and 

model simulations. 

BIOACID includes project leaders of the two marine Excellence Clusters “The Future Ocean” 

(Kiel) and “The Ocean in the Earth System” (Bremen/Bremerhaven) which host a variety of 

projects with relevance to ocean change, and will contribute to the cooperation between these and 

other national marine institutions. 

35

36 

BIOACID: Biological Impacts of Ocean Acidification


37

10. Summary Budget 

38 



11. Detailed descriptions of Themes and Projects 

0.1 Projektkoordination (Ulf Riebesell) 

0.2 Daten-Management (Michael Diepenbroek) 

0.3 Infrastruktur-Entwicklung (Hans Pörtner) 

0 

0.4 Training und Wissenstransfer (Michael Meyerhöfer) 

Koordination 

2 3 4 5 

1 

Integrierte 

Abschätzung: 

Sensitivitäten und 

Unsicherheiten 

Interaktionen zwischen 

Arten und die 

Zusammensetzung der 

Gemeinschaften in 

einem sich ändernden 

Ozean 

(Maarten Boersma) 

Kalzifizierung: 

Empfindlichkeiten 

von Phyla bis zu 

Ökosystemen 

Leistungsmerkmale bei 

Tieren: Reproduktion, 

Wachstum und 

Verhaltensweisen 

(Dirk de Beer) 

(Hans Pörtner) 

Primärproduktion, 

mikrobielle Umsätze 

und biogeochemische 

Rückkopplungsmechanismen 

(Maren Voß) 

Themen 

(Andreas Oschlies) 

Einfluss der 

Alkalinitätsflüsse vom 

Wattenmeer auf den 

Kohlenstoffkreislauf und 

die Primärproduktion der 

Nordsee 

(Johannes Pätsch) 

Zelluläre Mechanismen 

der Kalzifizierung 

3.1 

Auswirkungen auf 

Weidegänger und Filtrierer 

2.1 

Akklimatisierung v ersus 

Anpassung in 

autotrophen Organismen 

(Thorsten Reusch) 

1.1 

5.1 

Effekte der Ozeanversauerungauf 

artspezifische Performance 

sowie Konkurrenz und 

Struktur in mikrobiellen und 

makrobiellen 

Gemeinschaften 

(Martin Wahl) 

4.1 

(Frank Melzner) 

(Thomas Brey ) 

Kalzifizierung unter pH- 

Stress: Wirkungen auf 

organismischer und 

ökosystemarer Ebene 

(Ralph Tollrian) 

Physiologische 

Langzeiteffekte auf 

Lebensstadien 

benthischer Krebse 

Fig. 0.1: Consortium structure: overarching activities, themes and projects. Responsible PIs in 

parentheses. 

3.2 

Abbau und Umsetzung v on 

organischen Substanzen 

Ev aluierung und 

Parameteroptimierung der 

pelagischen 

Kalziumkarbonat 

Produktion in globalen 

biogeochemischen 

Modellen 

(Andreas Oschlies) 

2.2 

1.2 

(Felix Mark) 

(Anja Engel) 

5.2 

Effekte der 

Ozeanv ersauerung auf 

Nahrungsnetze und 

kompetitive Interaktionen in 

pelagischen Ökosystemen 

(Maarten Boersma) 

4.2 

Ultrastruktur und 

elementarer Aufbau von 

Kalkskeletten 

(Foraminiferen, Korallen) 

3.3 

Auswirkungen auf Top- 

Prädatoren: Fische und 

Cephalopoden 

2.3 

Modellierung biogeochemischer 

Rückkopplungsmechanismen 

der organischen 

Kohlenstoffpumpe 

(Birgit Schneider) 

1.3 

(Catriona Clemmesen) 

(Jelle Bijma) 

Bew ertung unsicherer 

ökologisch-ökonomischer 

Projekte 

Folgen der Ozeanv 

ersauerung mit der 

Viabilitäts-Methode 

(Martin Quaas) 

5.3 

Durch Mikroumgebungen 

kontrollierte (De)- 

Kalzifizierungsmechanismen 

(Michael Böttcher) 

3.4 

Der Einfluss rezenter und 

fossiler Ozean Versauerung 

auf den Stoffwechsel, die 

Biomineralisation und die 

Biodiv ersität pelagischer und 

neritischer Kalkschaler 

(Adrian Immenhauser) 

3.5 

39

40 


Table 0.1: Links between projects. Coloured boxes indicate projects with direct exchange (e.g. joint experiments, joint 

use of equipment and measurement capacity, exchange of samples, etc.) 

0.3.1 

0.3.2 

0.4 

1.1.1 

1.1.2 

1.1.3 

1.1.4 

1.1.5 

1.2.1 

1.2.2 

1.2.3 

1.2.4 

1.2.5 

1.3 

2.1.1 

2.1.2 

2.1.3 

2.2.1 

2.2.2 

2.3.1 

2.3.2 

3.1.1 

3.1.2 

3.1.3 

3.1.4 

3.2.1 

3.2.2 

3.2.3 

3.2.4 

3.3.1 

3.3.2 

3.4.1 

3.4.2 

3.4.3 

3.5.1 

3.5.2 

3.5.3 

4.1.1 

4.1.2 

4.1.3 

4.1.4 

4.2.1 

4.2.2 

5.1 

5.2 

5.3 

0.3.1 

0.3.2 

0.4 

1.1.1 

1.1.2 

1.1.3 

1.1.4 

1.1.5 

1.2.1 

1.2.2 

1.2.3 

1.2.4 

1.2.5 

1.3 

2.1.1 

2.1.2 

2.1.3 

2.2.1 

2.2.2 

2.3.1 

2.3.2 

3.1.1 

3.1.2 

3.1.3 

3.1.4 

3.2.1 

3.2.2 

3.2.3 

3.2.4 

3.3.1 

3.3.2 

3.4.1 

3.4.2 

3.4.3 

3.5.1 

3.5.2 

3.5.3 

4.1.1 

4.1.2 

4.1.3 

4.1.4 

4.2.1 

4.2.2 

5.1 

5.2 

5.3


Theme O: Overarching activities 

Project 0.1: Project coordination 

Coordinator: Ulf Riebesell 

Project Coordination 

BIOACID will be coordinated at the Leibniz-Institute of Marine Sciences (IFM-GEOMAR) in 

Kiel by Prof. Ulf Riebesell. The coordinator will be responsible for the day-to-day management 

of BIOACID. He will ensure that the project runs in compliance with the requirements of the 

contract (administrative, operational and scientific aspects). The Coordinator Project Office will 

be organized at IFM-GEOMAR. Prof. Hans-Otto Pörtner at the Alfred-Wegener-Institute for 

Polar und Marine Sciences in Bremerhaven will act as Deputy Coordinator. He will fill in for the 

Coordinator position under circumstances when the Coordinator is unavailable. 

To uphold an efficient project management, the project coordinator will employ a full time 

project manager (PM) and a half time secretary. The assistance of financial officers and 

personnel at central administrative departments will be provided at no cost. 

The PM will regularly report to the Coordinator. His scientific and administrative tasks will be to: 

• assist the Coordinator as the overall contact person on scientific matters and reporting, 

and serve as the intermediary for communication between the partners on administrative 

and financial matters 

• organise meetings and workshops, as well as any action for training and dissemination 

purposes 

• prepare agendas for all meetings 

• communicate all decisions regarding the implementation of the scientific work to the 

theme leaders and project PIs 

• work in close collaboration with the theme leaders and project PIs, collecting all formal 

research and training plans and progress reports 

• coordinate and run the day-to-day administrative and financial tasks 

• control that financial contributions are transferred to contractors on time 

• assist the project communication. 

Budget and Budget Justification 

Personnel costs 

Project manager E13-5 

Secretary E6 1/2 

Student helpers (8 x 75hs) 

First Year Second Year Third Year Total 

41

Overtime technicians 

Dissemination: Camera 

personnel for TV – 

production 

Subtotal 

Consumables 

Printing costs management 

Meetings, workshops, 

catering 

Consumables office 

Dissemination: printing 

costs (poster, flyer etc.) 

Dissemination: material 

TV production 

Subtotal 

Travel 

Coordinator 

Project manager 

Dissemination: camera 

personnel 

Subtotal 

Investments 

Computer project manager 

Computer secretary 

Subtotal 

Other costs 

Dissemination: cutting TV 

production 

Subtotal 

TOTAL 

Budget justification 

42 



Personnel: The scientist position is for a full time project manager (PM). This position will be 

offered to Dr. Michael Meyerhöfer, who has assisted during the development of the BIOACID 

programme and has contributed immensely to the preparation of the BIOACID pre- and full 

proposals. The PM will handle logistical organisation and workshop coordination as well as 

reporting tasks for the project and dissemination activities. Due to the administrative complexity 

of BIOACID the PM will be supported by a half time secretary and student helpers. 

One important dissemination activity of BIOACID will be the production of a TV-quality video 

for distribution to different media, decision makers and other stakeholders interested in ocean 

acidification. This includes the engagement of professional film and production personnel.


Consumables: A yearly budget is requested to cover costs of printing, copying and publication 

charges for BIOACID relevant material for administrative (e.g. reports) and dissemination 

purposes. Further costs arise for the organisation of the BIOACID annual meetings, preparatory 

meetings for joint research and dissemination activities, meetings of BIOACID’s Executive 

Board individually and jointly with the coordination team of the UK-NERC programme on ocean 

acidification, as well as meetings with governmental and NGO stakeholders. All meetings of the 

project will be held at locations where meeting rooms are available at low or no charge so that 

costs can be kept at a minimum level. 

Travel: To maintain close contact to partners within the project and with international 

collaborators and to guarantee an effective exchange of information, travel funds are requested 

for the coordinator and the PM. 

Investment: A desktop computer including licenses for standard software is requested for the 

project manager and the secretary. 

43

44 


Project 0.2 Data management 

PI: Michael Diepenbroek, WDC-MARE / PANGAEA, University of Bremen 

i. Objectives: 

• To coordinate data capture within BIOACID themes 1-5 and promote dataflow between 

these themes and their working groups. 

• To store, manage and distribute output from the modelling exercises. 

• Perform technical quality control on all data and check the validity and completeness of 

metadata. 

• Provide open access to all BIOACID data and information in a timely and efficient 

manner via interfaces that are scalable for different partners and other stakeholders. 

• To ensure the long-term archiving, publication, and distribution of data according to 

international standards and protocols. 

• To harmonize data management and facilitate data exchange between BIOACID and 

related projects, such as EPOCA and SOPRAN, which both use PANGAEA for data 

management 

The sharing of data and information among partners of large coordinated projects, such as 

BIOACID, strengthens the collaboration between different disciplines and research groups and 

ultimately leads to an increased scientific output through synergistic effects. Experience from 

previous projects has shown that both individual scientists as well as the project as a whole 

benefit from data sharing and collaboration. The benefit of BIOACID to the wider scientific 

community also requires data generated within the project to be freely available and easily 

accessible. To encourage and promote data and information exchange in a timely and efficient 

manner, BIOACID will establish a data policy with binding rules for all project partners. While 

calling for openness and free flow of data between partners, these rules must also protect the 

intellectual property rights of the data producers. 

A major requirement of the data policy is that all data must be lodged in the BIOACID database 

within 3 months after the time of measurement (data provided at zero cost to the project may by 

lodged in other databases as long as they are publicly-accessible). Access restrictions may be set 

up during the proprietary rights period (2 years after measurement) on request from the data 

originator. During the proprietary rights period, data will only be passed to project partners with 

the data originators agreement. BIOACID will encourage all PIs to share data during the 

proprietary rights period and should ensure that the data originators contribution is respected 

appropriately through co-authorship or acknowledgement. 

ii. Background and Previous Work 

The proponents of this subproject have a broad and substantiated background in the fields of data 

management and related IT subjects (databases, data infrastructures, protocols, system design, 

and automatization). During the past 15 years the group has built up an information system 

(PANGAEA – Publishing Network for Geoscientific & Environmental Data – www.pangaea.de),


which is a system for acquisition, processing, long-term storage, and publication of georeferenced 

data related to all earth science fields. In 2001 the PANGAEA group founded the 

ICSU World Data Center for Marine Environmental Sciences (WDC-MARE – www.wdcmare.org), 

which uses PANGAEA as operating platform. The organization of data management 

includes quality check and publication of data and the dissemination of metadata according to 

international standards. 

The challenge of managing the heterogeneous and dynamic data of environmental and 

geosciences was met in the PANGAEA system through a flexible data model which reflects the 

information processing steps in the earth science fields and can handle any related analytical data. 

The basic technical structure corresponds to a three-tiered client/server architecture with a 

number of clients and middleware components controlling the information flow and quality. On 

the server side a relational database management system (RDBMS) is used for information 

storage. To ensure fast data access the data are mirrored in a data warehouse which is also used 

as interface to the German GRID community. RDBMS and warehouse currently hold about 

450.000 fully documented data sets, comprising more than 2 billion analytical values 

(observational data). Each data set can be referenced by a DOI (Digital Object Identifier). All 

interfaces to the information system are based on web services including a simple map supported 

(WFS, UMN) search engine (PangaVista). 

For a number of international projects, PANGAEA is providing data portals for community 

specific data access to a heterogenic data center infrastructure [Schindler U et al., 2006]. 

Currently portals for CARBOOCEAN and EUR-OCEANS are running. 

The PANGAEA and the WDC-MARE are operating on a long-term basis. The institutional frame 

is supplied by RCOM in cooperation with the Alfred Wegener Institute (AWI), Bremerhaven. 

PANGAEA is a partner in over 50 national and international projects covering all fields of 

environmental sciences (http://www.pangaea.de/Projects), in particular the EU Integrated Project 

EPOCA, EU Coordinated Project EPOCA and the German BMBF Coordinated Project 

SOPRAN, which all have close links with BIOACID. 

References 

Diepenbroek, M; Grobe, H; Reinke, M; Schindler, U; Schlitzer, R; Sieger, R & Wefer, G (2002): PANGAEA - an Information 

System for Environmental Sciences. Computer & Geosciences, 28, 1201-1210, doi:10.1016/S0098-3004(02)00039-0 

International DOI Foundation (2003): DOI Handbook. doi:10.1000/182 

Schindler, U; Brase, J; Diepenbroek, M (2005): Webservices Infrastructure for the Registration of Scientific Primary Data. In: 

Rauber, A et al. (eds.): ECDL 2005, LNCS 3652, pp. 128-138, 2005, doi:10.1007/11551362_12 

Schindler, U; Diepenbroek, M (2006): Generic Toolbox for Metadata Portals. In prep. 

iii. Detailed Description of the Work Plan 

The BIOACID DIS will be developed and implemented as early as possible so that data and 

information flow is initiated in parallel with the first research activities. The purpose of the 

BIOACID data management plan is to create a semi-distributed, scalable, and flexible 

international data system into which research, observation, and modelling activities can submit 

data and which will provide services that will increase the value of the data. Data capture and 

data flow will be managed by an experienced data manager employed half-time and located at 

WDC-MARE at the University of Bremen. The data manager will be responsible for: 

• Maintenance of the BIOACID web-page 

45

46 


• Planning of data reporting procedures with the PIs prior to data collection 

• Keeping track of project data and the associated metadata 

• Web-based data tracking, including provision of common access to data by project PIs 

• Archival of data at the end of the project 

According to the rules of good scientific practice (ESF 2001) all data sets will be archived longterm, 

and published using the partners’ data centres, in accordance with the wishes of the 

responsible PIs. The data manager will watch that intellectual property rights are obeyed. 

To provide access to the project-wide information BIOACID will be embedded in a common data 

and information infrastructure implemented and maintained by the data manager and technical 

staff from WDC-MARE. Contents and services of the website include: 

• General project information (documents, workshops, cruises, news etc.) 

• Data portal 

• Inventory of site and sampling locations (dynamic map) 

• Bibliography of BIOACID specific publications 

• “Yellow pages” for BIOACID related scientists 

The data retrieval and access system will be based on distributed metadata catalogues located at 

the cooperating data centres. Catalogue contents and access protocols are based on international 

standards and protocols for geospatial data and metadata (e.g. ISO19115, DIF, DC, OGC-WFS, 

OAI-PMH). Use and maintenance of these standards will ensure a high level of data consistency 

and quality and will also facilitate later incorporation or union with information systems from 

other communities. User interfaces will range from a map-based full text search engine to 

hierarchical access through dynamic web pages. The implementation of the data infrastructure 

will be carried out with regard to international SDI initiatives, in particular EU initiative 

INSPIRE and GSDI. 

Dissemination will be a continuous process throughout the project. All dissemination actions 

and products will be negotiated with the BIOACID Management Team. Dissemination of project 

results and deliverables will be carried out to the scientific community, political decision makers, 

and the general public. Data management activities will be evaluated and critically reviewed 

during workshops and the annual BIOACID meetings. 

iv. Specific Tasks and Deliverables 

1. Coordination of data capture, integration and quality control activities for the five BIOACID 

Themes. 

As outlined in the Overview paper, original data sets have to be compiled from a number of 

different sources, including data from BIOACID related cruises, experiments and from existing 

data centres and working databases. Furthermore, the quality and state of these data sets might


vary enormously, as data are stored in a variety of formats with varying levels of metadata. 

Therefore, in the initial phase an implementation plan for data and information management will 

be set up. This plan will outline a structure and schedule for overall data flow between working 

groups, quality management schemes, and data processing schemes. In addition it will regulate 

the property rights of the data originators and the appropriate time intervals for public release 

from creation of the data, as well as confidentiality issues. 

An initial metadata catalogue of available data sets will be drawn up and used to define a priority 

list for data acquisition and compilation. The catalogue will be continuously updated to enable all 

BIOACID participants to locate and download target data sets. The metadata catalogue will be 

complemented by a BIOACID specific bibliography. 

2. Archiving and publishing data sets and data collections online and as offline products (DVD) 

using persistent identifiers as DOIs. 

Data sets will be described consistently according to the common metadata schema used. Data 

still subject to access restrictions will be password secured and for internal use only. Published 

data sets will be registered and made citable through usage of Digital Objects Identifiers (DOI) as 

persistent identifiers (http://www.std-doi.de/). By way of the DOI registry data sets can also be 

retrieved from library catalogues. 

3. Implementation of the BIOACID data infrastructure 

The data infrastructure (SDI) developed in this task has two objectives: 

• Enabling a distributed storage of observational and model simulation data within a 

common networked structure, and 

• Establishing a robust and long lasting data network which can be extended by or 

integrated into ongoing projects and programs. 

The first is directly necessary for BIOACID to handle the potentially large mass of data from 

modelling as well as the highly heterogeneous observational data. The second objective 

corresponds to the worldwide efforts (as with INSPIRE) in developing a global spatial data 

infrastructure (GSDI). A first setup will be based on the Open Archives Initiative protocol (OAI- 

PMH) with Dublin Core as content standard. OAI-PMH is a widely used “de facto” standard, 

supported by ad hoc usable open source software packages. One potential difficulty arises from 

the currently rapid pace of SDI developments. Therefore, from the start of the project, all 

implementations will follow a generic and flexible design allowing for later modifications and 

extensions. 

4. Maintain the website and data portal for BIOACID 

The existing BIOACID website will be converted to WDC-MARE standards. The website will 

include the data portal which enables efficient access to the individual and integrated data sets 

(‘one stop shopping’). Key components will be a BIOACID specific search engine allowing for 

retrieving and downloading of data sets relevant to the project. The search engine will form the 

47

48 


front-end of the common metadata catalogue. For best performance a harvesting approach will be 

chosen for assembly and maintenance of the catalogue. The index for the search engine will be 

complemented by a thesaurus. In order to minimise any friction losses during requests and 

delivery of data sets a first version of a general SOPRAN data portal will be set-up in the first 

phase of the project. 

v. Budget and Budget Justification 


Data manager E13 1/2 

Subtotal 

Consumables 

Office consumables 

Subtotal 

Travel 

Domestic travel 

Subtotal 

Investments 

Computer 

Supplies 

Subtotal 

Other costs 

Subtotal 

TOTAL 


0.1 


Personnel costs: A half-time data manager position will be established by WDC-MARE at the 

University of Bremen. The data manager will be integrated in the staff of WDC-MARE while at 

the same time keeping close contact with the coordinator and the project manager at all times 

during the project. 

Consumables: This will cover costs related to day-to-day data managing, including copying 

charges for management relevant material (e.g. reports) and dissemination purposes. 

Travel: To maintain close contact to the different partners within the project, to participate in the 

BIOACID annual meetings and meetings relevant to data management and to guarantee an


effective exchange of information, travel funds are requested for the data domestic travel oif the 

data manager. 

Investment: Funding is requested for a workstation for the data manager and related supplies. 

Project 0.3 Infrastructure development 

PI: Hans O. Pörtner 

i. Objectives 

This project intends to support and develop the aquarium facilities and research infrastructure 

needed for long term incubations (0.3.1.) and analyses of CO2 partial pressures in body fluids of 

marine animals and ambient waters (0.3.2.). Technique development described under this project 

is relevant for many projects of the network relying on animal maintenance and experimentation. 

Firstly, the use of maintenance and culture facilities in recirculated aquaria like the one at AWI 

Bremerhaven brings with it the inherent shortcoming that water physicochemistry is fluctuating 

over time (largely due to net proton equivalent ion exchange of the organisms) and needs to be 

monitored and stabilized on long time scales. Such shortcomings are not experienced by marine 

institutes at rocky shores where flow through systems can be used for maintenance of water 

physicochemistry (However, such flow through systems will strongly be influenced by 

environmental variability). Such aspects also have to be considered for experimental mesocosms 

and larval cultures under recirculated versus flow through conditions. The techniques proposed in 

subproject 0.3.1. are intended to close this gap and should support establishment of stable 

baseline conditions in recirculated systems. The solutions to be developed are also suitable to 

establish baseline conditions in larger experimental systems at pre-industrial levels and various 

levels of CO2 induced ocean acidification and thus appear as a relevant contribution which 

widens the scope of experimental research in the field of ocean acidification and may also 

become relevant in aquaculture systems. 

Secondly, the online monitoring of carbon dioxide as outlined in subproject 0.3.2. can support the 

elaboration of new knowledge of the effects of increased carbon dioxide content on marine 

organisms and environment. The aim of this subproject is the development of new chemical 

optical sensors and instrumentation for the determination of carbon dioxide partial pressure 

(pCO2) in fluids of marine organisms and in marine environments based on the dual lifetime 

referencing (DLR) method patented by PreSens. Emphasis will be put on the development of 

pCO2 sensors with appropriate dynamic range in order to meet the conditions present in marine 

systems and on the development of suitable instrumentation. The performance of the new 

equipment will be optimised according to the experimental applications provided by project 

partners with the prototype pCO2 sensors and instruments. 

ii. State of the Art 

Aquarium maintenance and mesocosms 

Stability of sea water physicochemical parameters needs to be secured in animal maintenance 

systems. This is a matter of significant concern in all laboratories which do not have access to 

49

50 


open ocean flow through seawater and rely on recirculated aquaria with a water treatment 

section. This includes the large recirculated aquarium system at AWI Bremerhaven, funded by 

BMBF upon the unification of AWI and BAH. 

Oxidative catabolism of heterotrophs is characterized by net proton production (Pörtner 1995) 

which is compensated for by ion exchange mechanisms and leads to a net release of protons into 

the water with the result of disturbed physicochemical parameters at constant or falling DIC (the 

latter occurs once excess CO2 generated by titration of (bi)carbonate is released into the 

atmosphere). Furthermore, any delay in the equilibration of air or gas mixtures with the aquarium 

water will prevent precise analysis of relevant water physicochemistry and thus sabotage any 

efforts to maintain constant settings of relevant parameters. The goal of this development must 

therefore be to monitor the effects of non-respiratory proton loads to the system water under 

equilibrium conditions and to compensate for any pH disturbance resulting from net proton 

equivalent ion exchange by aquarium maintained organisms. 

Rearing of larvae of crustaceans, cephalopods and fishes (esp. cod) also needs to be carried out in 

recirculated aquarium systems under largely undisturbed conditions. These activities will make 

the putative most sensitive life stages available for studies of effects of ocean acidification. 

Rearing would ideally occur under control vs. elevated CO2 conditions. The proposed system is a 

miniaturized AquaInno Pond-in-Pond system for hatchery developed by AWI and IMARE (R. 

Fisch, B.H. Buck). It offers a “system within a system” approach by providing process water with 

additional purification and water exchange on demand, without disturbing sensitive life stages. 

Modification and change of modules is possible and support a wide range of experiments. The 

system can be isolated from air and ambient water for equilibration with various CO2 levels. 

During long term incubations repeated water exchange is feasible in the pH stabilized 

recirculated large scale aquarium systems (see above). 

Fig. 0.2 (1): Draft of the floating AquaInno nearshore technology illustrating its components for water treatment. The 

waste water flows from the fish sector (A) to a first waste collector (B), where particles will sediment in the tubes 

system. A following bioreactor (C) eliminates dissolved nitrogen and phosphate. The re-use of water is controlled by 

sensors (G) and sediment disposal (F) takes place. A pneumatic system simplifies handling and harvesting (E) of 

cultured organisms. The platform (D) around the culture module is improving the overall handling of the system. 

Surrounding algal cultures (I) eliminate the final product nitrate. D and I are not included for indoor use. 

Fig. 0.3 (2): First freshwater prototype of a floating in pond raceway used for cultivation of ornamental fish in the year 

2004. Currently, the system is developed for hatchery and grow-out in seawater and has been modified as a 

recirculation system with different modules. 

For CO2 equilibration according to IPCC emission scenarios large temperature controlled 

experimental chambers have to be maintained at constant CO2 partial pressures according to gas 

mixtures provided by commercial suppliers. A prototype developed by AWI (B. Klein) needs to


be further modified to reduce gas quantities used and improve stability of physicochemical 

parameters. 

Fig. 0.4: Prototype of larger scale CO2 incubation system developed at AWI (B.Klein) (total volume 2.5 m 3 , aerated 

and filtered, temperature controlled). 

CO2 sensor development 

Standard methods for the determination of gaseous carbon dioxide include the direct detection 

via infrared spectroscopy (Werle et al. 1998). Most sensors for the determination of dissolved 

CO2 are modified pH sensors exploiting the acidic nature of carbon dioxide in contact with 

aqueous media. In 1958 Severinghaus and Bradley introduced an electrochemical pCO2 sensor 

comprising a pH glass electrode immersed into a small volume of a weak bicarbonate buffer 

solution which is separated from the sample by a gas-permeable but ion-impermeable membrane, 

such as Teflon. This type of sensors has found widespread application in industrial (e.g. 

biotechnology or beverage) analyses (www.mt.com). 

Various efforts were made to develop optical pCO2 sensors since Lübbers and Opitz (1975) 

reported an optical pCO2 sensor based on the pCO2-dependent fluorescence intensity change of 4methylumbelliferone 

in a PTFE-covered bicarbonate buffer. These efforts resulted in optical 

pCO2 sensors for biotechnology applications (www.ysilifesciences.com, www.fluorometrix.com) 

based on fluorescence measurements. DeGrandpre et al. (1995) developed a submersible 

autonomous moored instrument for CO2 which is based on a colorimetric pH measurement and 

commercialised by Sunburst Sensors, LLC (www.sunburstsensors.com). Yet, this system is bulky 

and not suitable for laboratory measurements where only small amounts of sample volume (e.g. 

body fluids of marine organisms) are available. 

51

52 

CO 2 

I - 

I - 

CO 2 

CO 2 


Carbon dioxide dissolves in aqueous solutions both physically and chemically. Carbonic acid 

thereby produced causes a pH drop which can be visualised with pH-sensitive indicators. Thus 

principally all chemical optical carbon dioxide sensors are based on the detection of the pH 

change in a CO2-sensitive matrix - whether they are based on colorimetric or luminescence 

detection methods. Luminescence detection-based methods are superior to colorimetric 

approaches due to their higher sensitivity. Here, the luminescence lifetime detection technique 

offers an advantage compared to intensity-based methods because intensity fluctuations - caused 

by e.g., fluctuations of excitation light source or background light - can be easily referenced. The 

decay time is insensitive to such fluctuation whereas the luminescence intensity is affected. The 

principle of luminescence quenching by carbon dioxide is illustrated in Fig. 0.5. 

Excitation Emission 

HI 

I - 

HI 

Protonation No Emission 

Fig. 0.5: Principle of luminescence quenching by 

carbon dioxide. The deprotonated form of the pHsensitive 

indicator I - displays fluorescence when excited 

at a distinct wavelength. In the presence of carbon 

dioxide a portion of the pH-sensitive dye in the sensor is 

protonated. The emission intensity and thus the average 

lifetime of the indicator system decrease at increasing 

carbon dioxide concentration since the protonated form 

cannot be excited by the excitation light used. 

The measurement principle of chemical optical pCO2 sensors to be developed in this project is 

based on the determination of the luminescence decay time of pH-sensitive dyes incorporated in a 

gas-permeable – but ion-impermeable – membrane. With the DLR method patented by PreSens it 

is possible to internally reference the luminescence decay time detected and to transfer it into the 

µs time regime. This enables the use of easier and economically affordable instrumentation based 

on the phase modulation technique. 

Together with the luminescent dye sensitive to carbon dioxide an inert luminescent dye is 

incorporated into the sensing membrane. Both dyes are excited by sinus-modulated light. Thus, 

the luminescent response – a superposition of the CO2-sensitive dye and the inert dye – is also 

modulated, but with a phase shift proportional to the mean decay time of the overall system, 

which is again a function of the carbon dioxide concentration. The principle of the luminescence 

decay time detection by phase modulation technique is visualised in Fig. 0.6. This technique is 

used for all devices from PreSens. It proved to be a very reliable and robust method.

Signal 


Φ 1 

Φ 2 

time / µs 

iii. Previous Work of the Proponents 

0.3.1 

Excitation 

high CO2 

low CO2 

Fig. 0.6: Principle of luminescence 

decay time detection. At low pCO2 the 

contribution of the CO2-sensitive dye 

(short decay time) to the total signal is 

high resulting in a low phase shift (Φ1) 

compared to excitation. With increasing 

pCO2 the weight of the reference dye 

(long decay time) increases resulting in 

an increase of phase shift (Φ2). 

Franz Josef Sartoris has a strong background in the physiology of marine animals, including 

acid-base regulation, ion- and osmoregulation, energy metabolism and respiration (Sartoris and 

Pörtner, 1997; Pörtner and Sartoris, 1999; Sartoris et al., 2003; Metzger et al., 2007). He has a 

broad experience with the implementation of physiological and biochemical methods in smallsized 

animals like amphipods, copepods and decapod larvae. He is the Scientific Manager of the 

state of the art recirculating seawater aquarium at the AWI with a capacity of 150 m 3 for the 

rearing of temperate and polar animals. 

Bela Hieronymus Buck is a marine biologist experienced in various aspects concerning the 

cultivation of marine organisms (e.g. Buck et al., 2002, 2005 a,b). Research topics addressed 

were the physiology and symbiotic interaction of giant clams, parasite infestation of bivalve, 

cultivation strategies of various species including fish, and the development of new system 

design featuring its realization by accepting socio-economic aspects and co-management. He also 

works on the modification of modern aquaculture recirculation systems (RAS) for fish 

cultivation. Bela H. Buck is the head of the unit “Marine Aquaculture, Maritime Technologies 

and ICZM” in AWI, and of the unit “Marine Aquaculture for responsible fisheries” of IMARE 

(Institute for Marine Resources).. 

Ralf Fisch is a biologist experienced in bionics and aquaculture (Fisch and Buck 2006, 2008). 

During his early work he was studying the biological surfaces of arthropods. To explore the 

multifunctional nano-structures he used oSEM, AFM and TEM technologies. Today he is 

working in aquaculture research topics including the modelling of nutrient budgets and ecological 

aspects. Also educated as an engineer, he is experienced in the construction and design of 

aquaculture systems and co-inventor of a floating recirculation aquaculture system (FRAS). His 

prime interest is the development and implementation of a sustainable aquaculture. 

Guido Krieten is the technical manager of the seawater aquarium system at AWI. He is a 

qualified mechanic and builder of central heating and ventilation systems. He has a degree in 

maritime engineering. He is responsible for the technical aspects of aquarium maintenance, the 

53

54 


maintenance of seawater quality and the operation in compliance with safety regulations. 

Hans O. Pörtner has a long standing history in studying acid-base regulation of marine 

ectotherms and its metabolic impacts in relation to ambient water conditions (e.g. Pörtner et al. 

1991, 1998). His experience in analyses of proton equivalent ion exchange will be an asset in the 

monitoring and establishment of stable water physicochemistry. Current interest covers the use of 

these systems in studies of interaction between CO2 levels and other climatic factors, the 

mechanisms shaping cellular and whole-animal energy budgets under various thermal and 

carbon-dioxide regimes, and the molecular mechanisms of environmental adaptation and 

limitation. He has published more than 190 publications in peer-reviewed journals. 

0.3.2. 

PreSens has been developing and selling optical sensors since 1997 (Klimant, 1997, Apostolidis 

et al, 2004, Schröder and Klimant, 2005). The product portfolio includes the production and 

marketing of fiber-optical chemical sensors and scientific instrumentation for the determination 

of oxygen, pH and temperature. Oxygen and pH sensors were successfully applied by AWI and 

IFM-GEOMAR in various experimental setups, among many other scientific and industrial 

customers. pCO2 sensors and instrumentation based on the dual lifetime referencing method 

patented by PreSens are currently under development for biotechnology and medical 

applications. It is reasonable that this concept can be transferred also to the development of pCO2 

sensors with dynamic range in the trace concentration range (380 ppm – 1900, in body fluids 380 

– 3000 ppm). The scientific team has gained expertise on the development of optical sensors for 

more than 10 years. 

Athanas Apostolidis is a research engineer at PreSens GmbH, Regensburg. He received his 

diploma in chemistry on development of a multi-channel optical protein detector for continuous 

annular chromatography. He received his Ph.D. funded by the BMBF in 2004 on a combinatorial 

approach for development of optical gas sensors at the Institute of Analytical Chemistry, Chemo- 

and Biosensors (Prof. Wolfbeis) at the University of Regensburg. During his Ph.D. he was 

working on the development of optical carbon dioxide sensors. In 2005 he was working at 

Mühlbauer AG, Roding, Germany, as process engineer and materials specialist in the 

development of production processes for Smart Card and passport solutions. His present interest 

is in the development of optical chemical sensors for carbon dioxide determination in biological, 

biotechnology and medical applications 

Christian Huber is a research engineer at PreSens GmbH, Regensburg. He received his Ph.D. in 

2000 on design and characterization of novel anion-selective optical sensors at the Institute of 

Analytical Chemistry, Chemo- and Biosensors (Prof. Wolfbeis) at the University of Regensburg. 

During his Ph.D. he was working on the development on anion-selective optical chemical sensors 

based on the DLR method. At PreSens he is a product specialist for application of chemical 

optical oxygen sensors in food and beverage, pharmaceutical, biotechnology and medical 

industry.


iv. Work Programme, Schedules, and Milestones 

Subproject 0.3.1. 

Recirculated mesocosms and larval cultures (full strength sea water) 

PI: Franz J. Sartoris, Bela H. Buck, Ralf Fisch, Guido Krieten, Hans O. Pörtner 

Work Programme 

a. Development of a large recirculated aquarium system into a stable system maintaining baseline 

physicochemical parameters. 

The adaptation of proton equivalent ion exchange monitoring systems according to Heisler 

(1989, cf. Pörtner et al. 1991, 1998) will support monitoring of physicochemical parameters in 

equilibrium. Furthermore, carbonate buffers need to be used for compensation of non-respiratory 

proton loads to the water. Fine-tuning of pH should be possible through pH-stat controlled 

addition of bicarbonate solutions. Developments thus include: 

• Instrumentation for the monitoring of pH and DIC content under well controlled 

conditions of PCO2 and temperature. 

• Establishment of adequate calcite buffering in large scale animal maintenance systems 

(simulating the CO2 dependent buffering action of ocean sediments). 

• Complementary pH-stat controlled addition of bicarbonate solutions. 

b. Development of a rearing and incubation system for larvae of various groups 

Available prototypes will firstly be tested for use with established cephalopod cultures (Sepia 

officinalis, project 3.1, 2.3) and crustacean larvae (project 2.2.). It will be investigated whether 

the rearing of fish larvae can be carried out under those conditions, thereby complementing or 

deviating to some extent from established aquaculture procedures. One of us (B. Klein) has 

carried out the rearing of cod larvae in aquaculture facilities in Tromso, Norway. 

c. CO2 incubation system for mesocosms (pH stat) 

Combining the methods established under a. with the large experimental CO2 incubation system 

will support long term stability and provide the basis for pH stat control of the water through 

controlled addition of high concentration bicarbonate solutions (available inhouse funding 

20.000,- €). 

Internal cooperation: Projects 2.1.1. / 2.1.2. / 2.2.1. / 2.2.2. / 2.3.1. / 2.3.2. etc. 

55

Schedule 

56 


0.3.1 First Year Second Year Third Year 

Establishment of pH bicarbonate 

monitoring system 

Establishment of stabilized aquarium 

physicochemistry 

Establishment of prototype for larval 

culture 

Successful rearing of cephalopod 

larvae under various water 

physicochemistry settings 

Successful rearing of crustacean larvae 

under various water physicochemistry 

settings 

Successful rearing of fish larvae under 

various water physicochemistry 

settings 

Analysis of system quality, success 

evaluation, data interpretation 

Manuscript preparation, presentation of 

results at conferences 

Milestones 

I II III IV I II III IV I II III IV 

- Implementation of pH bicarbonate analysis system month 6 

- Establishment of stabilized aquarium physicochemistry month 12 

- Complementation of aquarium equipment month 20 

- Establishment of prototype for larval culture month 12 

- Successful rearing of cephalopod larvae under various water 

- physicochemistry settings month 18 

- Successful rearing of crustacean larvae under various water 


- Successful rearing of fish larvae under various water 


- Evaluation of combined data sets, system properties and uncertainties month 33


Subproject 0.3.2. 

Development of Chemical Optical Sensor Technology for the Determination of pCO2 

Partial Pressure in Fluids of Marine Organisms and Marine Environment 

PI: Athanas Apostolidis, Christian Huber 

PreSens Precision Sensing GmbH, Josef-Engert-Str. 11, 93053 Regensburg 

phone: +49 941 942 72 150 / 114 fax: +49 941 942 72 111 

e-mail: athanas.apostolidis@presens.de , christian.huber@presens.de 


WP 1: Sensor Chemistry Development 

This work package is divided into two main tasks. First, materials feasible for the production of 

pCO2 sensors for seawater application will be selected according to the environmental conditions 

present for the application of a pCO2 sensor. This selection includes decision for feasible pH 

indicator dyes, buffer systems and matrix polymers. Second, from these components sensor 

membranes will be produced and tested with respect to their dynamic range and operational 

performance in a calibration setup to verify best performing membrane compositions. 

Duration: 12 months 

Milestone M1: Sensor chemistry feasible for the determination of pCO2 in the concentration 

range present in marine environment. 

WP 2: Sensor Housing Development 

The sensor membranes developed in WP 1 will be integrated into mini sensor housings (e.g. flow 

through cell, dipping probe) that need to be designed based on the 2mm polymer optical fibers 

(POF) used by PreSens. For applications that cannot be served with the mini sensor setup the 

sensor chemistry will be adapted to micro sensor housings based on 140µm optical glass fibers. 

The different sensor / housing will be subject to storage stability tests to evaluate storage 

conditions required for retaining sensor performance during transport and storage of sensors at 

the end user. 


Milestone M2.1: Housing for mini pCO2 sensors and packaging of sensors for storage. 

Milestone M2.2: Housing for micro pCO2 sensors and packaging of sensors for storage. 

WP 3: Instrument Development 

Optical and electronic setup of the instrumentation will be developed to fit to sensor chemistry 

and to provide reliable measurements based on sensing technology from PreSens. Instrument 

housing will be developed for applications under laboratory and field conditions. Optical setups 

will be developed to fit a) mini sensor setups and b) micro sensor setups. 


Milestone M3.1: Laboratory demonstrator for pCO2 measurement in marine application 

for use with mini pCO2 sensors. 

Milestone M3.2: Demonstrator for measurement of pCO2 with micro pCO2 sensors. 

WP 4: Optimisation of Sensor Performance 

57

58 


According to the performance evaluated for the sensor chemistry developed in WP 1 and the 

feedback of field testers the sensors will be improved with respect to their accuracy, sensitivity 

and stability. The stability improvement will include both storage and operational stability. 


Milestone M4: pCO2 sensor with improved performance according to sensitivity and, both, 

operational and storage stability. 

WP 5: Optimisation of Device Performance 

The laboratory demonstrators for pCO2 measurement with mini or micro sensors will be 

improved with respect to environmental conditions of pCO2 determination in field. This can 

include battery operated systems, PC-independent setup, etc. 


Milestone M5: Instrument for field application pCO2 measurement with optimised performance. 

WP 6: Provision of Instruments, Sensors and Support to Project Partners 

PreSens will provide some instruments and sensors developed to field testers to verify 

performance of the calibration results in our labs to real experimental conditions. Presens will 

implement the feedback of the testing partners to the sensor chemistry and the instrument 

development and improvement. 


Result: Valuable feedback information for the validation of setup performance and the 

improvement of the pCO2 measurement system. 

Schedule 


WP 1 Sensor Chemistry Development 

WP 2 Sensor Housing Development 

WP 3 Instrument Development 

WP 4 Optimisation of Sensor 

Performance 

WP 5 Optimisation of Instrument 

Performance 

WP 6 Provision of Instruments, Sensors 

and Support to Project Partners 

I II III IV I II III IV I II III IV


Milestones (see work packages above) 

- Sensor chemistry feasible for determination of pCO2 in the concentration 

range present in marine environment (M1) 

- Housing for pCO2 mini sensors and packaging of sensors for storage 

(M2.1) 

- Housing for pCO2 micro sensors and packaging of sensors for storage 

(M2.2) 

- Laboratory demonstrator for pCO2 measurement in marine application for 

use with pCO2 mini sensors (M3.1) 

- Laboratory demonstrator for pCO2 measurement in marine application for 

use with pCO2 micro sensors (M3.2) 

- Instrument for field application pCO2 measurement with optimised 

performance (M5) 

- Instrument for field application pCO2 measurement with optimised 

performance (M5) 

Internal cooperation: 

month 12 

month 12 

month 21 

month 15 

month 24 

month 33 

month 36 

The sensors and devices that will be developed in this project will provide instrumentation 

infrastructure for online monitoring of carbon dioxide partial pressure in organism body fluids 

and marine environment for laboratory experiments of the project partners [link to 2.1.2, 2.1.3, 

2.3.1, 2.3.2, 3.1.3, 3.1.4, 3.2.2, 3.4.2]. Their experience with prototype instrumentation and 

sensors will be implemented in optimisation of performance. 

Cooperation with other projects outside the Verbundproject 

Collaborations beyond the project consortium that are relevant to the subproject project are not 

planned. A request for funding for this subproject has not been submitted to any other addressee. 

In case a request will be submitted, we will inform the Federal ministry of Education and 

Research (BMBF) immediately. 

59

60 





0.3.1 Technician 

0.3.2 Chemical Engineer, PhD * 7PM 

0.3.2 Electronic Engineer, PhD * 3PM 

0.3.2 Electronic Engineer, M.Sc * 3PM 

0.3.2Technician * 6PM 

Subtotal 

Consumables 

0.3.1. 

0.3.2 Chemicals and Gases * 

0.3.2 Polymer Optical Fibers * 

0.3.2 Consumables Sensor Production * 

0.3.2 Electronic and Optical 

Components * 

Subtotal 

Travel 

0.3.1 

0.3.2 National * 

0.3.2 International * 

Subtotal 

Investments 

0.3.1 

Subtotal 

Other costs 

0.3.1. 

0.3.2 * 

Subtotal 

TOTAL 

*:For subproject 0.3.2. numbers represent 50% of the total budget. The other 50% will be contributed by the company 

(Presens) itself. 

Budget justification 0.3.1 

Personnel costs: 

The success and feasibility of this project is dependent on the approval of the technical assistant, 

who will be in permanent charge of the very time-consuming process of larval rearing (e.g.


crustaceans, cephalopods, fishes) and associated assessments of water quality. The success of the 

project is dependent on a uniform manner of larval rearing to avoid errors and to minimize 

variability in conditions or ways of handling larvae. This excludes the repeated hiring of 

fluctuating personal, like students before or during their Masters periods. It is thus inevitable to 

have technical support available for continuous larval rearing, to provide sufficient good quality 

material for experimental work and to not distract scientific personal from the objectives of the 

science projects. 

After a period of vocational training and gaining daily routine, a full time position is needed to 

establish the rearing of larvae of the various groups under both control and experimental 

conditions. 

Consumables: 

Chemicals, Artemia eggs, algal and other (e.g. mysid) cultures for feeding of larvae 

Travel: 

The collection of animal specimens (ovigerous females) and eggs from various locations in 

Northern Europe demands travel funds for the technician and other personnel to support 

collection and safe arrival of fresh material at AWI. Exchange visits may also be needed with 

laboratories experienced in the rearing of e.g. cod larvae. 

Investments: 

Control and maintenance of water physicochemistry 

In accordance with the reasoning the aquarium systems mentioned above need complementation 

by flow through tanks containing buffer material of various compositions. They are also 

equipped by components of the pH bicarbonate system for online monitoring of pH under 

equilibrium conditions as well as a computer controlled pumping system for the addition of 

concentrated base. 

Larval rearing system 

In accordance with the reasoning above finalized development and construction of the prototype 

will demand a lump sum of per 3 years. 

CO2-Incubation mesocosm 

In accordance with the reasoning above long term incubation at various CO2 tensions in larger 

volume systems needs reproduction of the existing prototype, one for each CO2 tension. Together 

with available inhouse funding an amount of is requested to install 4 further CO2 systems to 

cover preindustrial, 2 x preindustrial, year 2100 and higher CO2 levels. 

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62 


Budget and Budget Justification 0.3.2 

Personnel: 

Chemical Engineer, PhD: Responsible for planning, execution and the development of pCO2 

sensors and measurement instruments; 

Electronic Engineer, PhD: Development and production of measurement instruments for pCO2 

sensors, especially circuit design and optical design; 

Electronic Engineer, M.Sc.: Development and production of measurement instruments for pCO2 

sensors, especially software development; 

Technician: Execution of sensor characterisation with laboratory calibration setups 

Consumables: Chemicals and gases are required for the production and the characterisation of 

the pCO2 sensors. Polymer Optical Fibers will be coated with the pCO2 sensor materials to be 

developed and will be supplied to project partners for evaluation. 

Consumables Sensor Production: For the production of pCO2 sensors supplies are required that 

are prone to deterioration and need regular replacement (e.g. membrane support materials, 

pipetting tips or needles); 

Electronic and Optical Components: Presens will build up to 5 demonstrators of pCO2 

measurement instruments for marine applications and supply to project partners. 

Travel: National: Travels to project partners and annual meetings; International: 1 international 

congress p.a. 

vi. References 

Apostolidis A, Klimant I, Andrzejewski D, Wolfbeis OS (2004). Combinatorial Approach for Development of Materials for Optical Sensing of 

Gases. J Comb Chem 6: 325 – 331 

Buck BH, Buchholz CM (2005b) Response of offshore cultivated Laminaria saccharina to hydrodynamic forcing in the North Sea, 

Aquaculture, 250: 674-691. 

Buck BH, Rosenthal H, Saint-Paul U (2002) Effect of increased irradiance and thermal stress on the symbiosis of Symbiodinium 

microadriaticum and Tridacna gigas, Aquatic Living Resources 15: 107-117. 

Buck BH., Thieltges DW, Walter U, Nehls G, Rosenthal H (2005a) Inshore-offshore comparison of parasite infestation in Mytilus edulis: 

Implications for open ocean aquaculture. J Appl Ichthyol 21: 107-113. 

DeGrandpre, M.D., Hammar, T.R., Smith, S.P., and F.L. Sayles. (1995)In situ measurements of seawater pCO2, Limnol Oceanogr, 40: 969 – 

975 

Fisch R, Buck BH (2006) Neues Aquakultursystem für das Meer made in Germany, Fischerblatt 12: 13-16. 

Fisch R, Buck BH (2008) Waste reduction in finfish aquaculture within suspended semi closed and closed systems. J Appl Ichtyol, in press. 

Heisler N (1989) Parameters and methods in acid-base physiology. In Techniques in Comparative Respiratory Physiology: an experimental 

approach (ed. C.R. Bridges and P.J. Butler). pp. 305-332, Society for experimental Biology Seminar Series. Cambridge, Cambridge 

University Press. 

Klimant I (1997) Verfahren und Vorrichtung zur Referenzierung von Fluoreszenzintensitätssignalen, Ger. Pat. Appl. DE 198.29.657 

Lübbers DW, Opitz N (1975) The pCO2/pO2 optrode: A new probe for measuring pCO2 and pO2 of gases and liquids. Z Naturforschung 30C: 

532-533. 

Metzger R, Sartoris FJ, Langenbuch M, Pörtner HO (2007) Influence of elevated CO2 concentrations on thermal tolerance of the edible crab 

Cancer pagurus. J Thermal Biol 32: 144-151 

Pörtner HO, Sartoris FJ (1999) Invasive studies of intracellular acid-base parameters: quantitative analyses during environmental and 

functional stress. In: regulation of Tissue pH in Plants and Animals, edited by EW Taylor, S Egington & JA Raven. S.E.B. Seminar 

Series, Cambridge University Press. 68-98 

Pörtner, HO (1995) pH homeostasis in terrestrial vertebrates: a comparison of traditional and new concepts. Adv Comp Env Physiol 22: 51-62. 

Pörtner, HO, Reipschläger A, Heisler N(1998) Metabolism and acid-base regulation in Sipunculus nudus as a function of ambient carbon 

dioxide. J exp Biol 201: 43-55. 

Pörtner, HO, Andersen NA, Heisler N (1991) Proton equivalent ion transfer in Sipunculus nudus as a function of ambient oxygen tension: 

relationships with energy metabolism. J exp Biol 156: 21-39. 

Sartoris FJ, Bock C, Serendero I, Lannig G, Pörtner HO (2003) Temperature dependent changes in energy metabolism, intracellular pH and 

blood oxygen tension in the Atlantic cod, Gadus morhua. J fish biol 62: 1239-1253 

Sartoris FJ, Pörtner HO (1997) Temperature dependence of ionic and acid-base regulation in boreal and arctic Crangon crangon and Pandalus 

borealis. J Exp Mar Biol Ecol 211: 69-83


Schroeder CR, Klimant I. (2005) The influence of the lipophilic base in solid state optical pCO2 sensors: a comparative study. Sens. Actuators 

B 107: 572 – 579. 

Severinghaus JW, Bradley AF (1998) Electrodes for blood pO2 and pCO2 determination. J Appl Physiol 13: 515 – 520. 

Werle P, Mücke R, Amato F D, Lancia T (1998). Near-Infrared Trace-Gas Sensors based on Room-Temperature Diode Lasers, Appl. Phys. B 

67: 307 – 315. 

Project 0.4: Training and transfer of know-how 

PI: Michael Meyerhöfer 

Training and transfer of know how between the BIOACID participants will be carried out by the 

following workshops, offered by the different institutions. 

1.: Workshop on microsensors (Dirk de Beer, MPI Bremen) 

Each of the two workshops should last ten days. The first half will be sensor making, the second 

sensor use. Participants can bring there own samples for measurements. 

2.: Workshop on isotope geochemistry and laser-ablationtechniques (Anton Eisenhauer, 

IFM-GEOMAR, Kiel) 

We intend to perform during four days a training workshop on the use of isotope geochemistry 

and on its use for biomineralization studies. This workshop will also give an introduction to the 

use of laser-ablation techniques as a new micro-analytical tool. 

3.: Physiological approaches to body fluid physicochemistry and acid-base regulation 

(Franz Sartoris, Christian Bock, Hans Pörtner and colleagues, AWI, Bremerhaven) 

We intend to host a three day workshop on the concepts of quantitative acid-base physiology and 

the use of various techniques associated with the quantification of changes in tissue and body 

fluid acid-base and associated ion and metabolic status in animals. The workshop will enable 

PhD students to interpret their findings in the light of changes in water physicochemistry and 

their effect on acid-base, ionic and metabolic regulation. 

4.: Workshop on seawater carbonate chemistry (Kai Schulz and Ulf Riebesell, IFM- 

GEOMAR, Kiel) 

All experimental projects proposed in BIOACID share a common aspect, i.e. the manipulation of 

the seawater carbonate chemistry for different carbon dioxide (CO2) scenarios. Adjusting, 

maintaining, and controlling the seawater carbonate system will be crucial for data interpretation 

and quality control. 

A joint two-day workshop on seawater carbonate chemistry will be held at IFM-GEOMAR at the 

beginning of BIOACID to ensure high data quality and inter-comparability. The workshop will 

cover the following topics:1) Principles of the seawater carbonate system: concepts, parameters 

and speciation2) Calculations: techniques to calculate the carbonate system from measured 

parameters 3) Measurements: dissolved inorganic carbon (DIC), total alkalinity (TA), pCO2 and 

pH) Experimental manipulations: acid / base additions, CO2 aeration, DIC additions at constant 

TA The workshop will enable all participants to choose the most appropriate CO2 manipulation 

63

64 


for their specific experimental approaches, determine the parameters necessary to calculate 

carbonate chemistry speciation and evaluate the data obtained for quality. 

5.: Workshop "Experimental design" (Martin Wahl, IFM-GEOMAR, Kiel) 

We plan a five day workshop on setting up experiments in consideration of: 

Schedule 

• Exact questions asked 

• Carefully chosen response variables 

• Temporal, spatial, methodological and biological scope of the investigation 

• Sources of variance 

• Desired data quality 

• Available resources (time, funds, infrastructure) 

• Synergetic combination with other investigation 

• Suitable statistical approach 

Workshop on microsensors 

Workshop on isotope geochemistry 

and laser-ablationtechniques 

Physiological approaches to body 

fluid physicochemistry and acid-base 

regulation 

Workshop on seawater carbonate 

chemistry 

Workshop "Experimental design" 

First Year Second Year Third Year 





Subtotal 

Consumables 

Workshop on microsensors 

Workshop on isotope 

geochemistry and laserablation- 

techniques 

Physiological approaches 

to body fluid 

physicochemistry and acidbase 

regulation 

Workshop on seawater 

carbonate chemistry 

Workshop "Experimental 

design" 

Subtotal 

Travel 

Subtotal 

Investments 

Subtotal 

Other costs 

Subtotal 

TOTAL 


The costs for the course will largely be carried by BIOACID. The participants need to take care of accommodation, food, and travel. 


The requested funds will only cover costs directly related to organizing and carrying out the 

training workshops at the various partner institutions. Costs related to travel and accommodation 

of the participants will be covered from travel funds of the individual sub-projects. 

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66 



11.2: Theme 1: Primary Production, microbial processes and 

biogeochemical feedbacks 

i. Common Background 

The partial pressure of CO2 (pCO2) in the atmosphere and the oceans has increased by 30% over 

the past century and model scenarios predict a further increase to twice the current concentrations 

by the end of the century, which is app. 700ppm by the year 2100 (Houghton et al. 2001). This 

increase is far more rapid than any change over the last 400,000 years and expected to 

significantly impact marine life (Fabry et al. 2008). Understanding the response of marine 

autotrophs and of heterotrophic bacteria to rising CO2 concentrations and decreasing pH is of 

special importance since these organisms provide the basis of marine food webs (link to theme 

4). All aquatic plants/primary producers are adapted to pCO2 concentrations that varied only 

between 200 and 300 ppm over the past 420 million years as ice core data suggested (Falkowski 

et al. 2000), while they are facing novel pCO2 conditions that are several times higher on a very 

rapid time scale within the next decades. To what extent species may currently be in the process 

of adaptation to higher pCO2 levels by evolutionary adaptation is totally unresolved. 

Fig. 1.1: Representatives of phytoplankton guilds that will be studied in theme 1 A Coccolithophore: Calcidiscus 

leptoporus B diatom:Skeletonema costatum C. diatom:Thallsiosiara angulata D Cyanobacteria:Nodularia spumigena. 

While increased pCO 2 may enhance photosynthesis by alleviating pCO 2 limitation, the extent and circumstances under 

which CO2 is limiting for photosynthesis and growth of primary producers in the field are still 

unclear. Preliminary data reveal that species with effective carbon concentration mechanisms 

(CCMs) are less sensitive to increased CO2 levels than those lacking efficient CCMs, analogous 

to findings in terrestrial vegetation (Collins et al. 2006). Currently, our ignorance of the 

metabolic diversity of oceanic autotrophy decreases the possibility of any realistic projection of 

marine primary production in response to increased carbonation. Calcifying organisms like 

coccolithophores and corals may suffer under decreasing calcification potentially leading to a 

negative feedback to atmospheric CO2 (Gattuso et al. 1998, Wolf-Gladrow et al. 1999, Riebesell 

et al., 2000, Orr et al 2005). 

Species specific differences in inorganic carbon acquisition moreover indicate changing 

competitive relationships between phytoplankton guilds (Fig. 1.1) at future CO2 concentration 

(Rost et al., 2003). Experimental work on diazotrophic cyanobacteria suggests an increase in 

productivity with increasing pCO2 (Hutchins et al., 2007; Ramos et al., 2007, Levitan et al. 

2007). Recent data from seagrasses and several groups of micro- and macroalgae suggest an 

enhancement of photosynthetic rates (Collins and Bell, 2004). Positively correlated CO2 

concentrations and uptake rates potentially constitute an important negative feedback to 

anthropogenic CO2 emissions, if enhancing the so-called biological carbon pump (Riebesell et al. 

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68 


2007). A systematic assessment of autotrophic responses considering possible co-limiting factors 

for photosynthesis such as nutrients, light and mixing depth, however, is still missing. 

Evolutionary adaptation to enhanced pCO2 levels as response of species and populations 

to environmental change is neglected in studies until now. In marine unicellular algae and 

bacteria, in particular, rapid evolution to increasing pCO2 levels is expected due to short 

generation times of


export of particulate matter from the upper water column is affected by high CO2 is essential to 

improve the predictive capacity of global biogeochemical models. 

An effect on the elemental stoichiometry of particles was found in large mesocosms where 

increasing DIC was translated to carbon rich organic matter (Engel et al. 2005, Riebesell et al., 

2007). The combined influence of light, nutrients, and CO2 concentration, on elemental 

stoichiometry and the relative importance of these factors individually is largely unknown 

(Burkhardt and Riebesell, 1997, Burkhardt et al., 1999). 

In the face of a not acceptable lack of information, the major objectives of theme 1 are to gain a 

better understanding of the response of autotrophic communities and heterotrophic bacteria to 

ocean acidification, and the related consequences for organic matter cycling, and the turnover of 

key elements. 

ii. Collaborative research 

In theme 1 various aspects of responses to ocean acidification and carbonation are studied 

including the export of primary production into the food-web and the formation of organic matter 

by aggregation of particles. Currently, data on the responses of the diverse plankton groups to 

Fig. 1.3: Set up with 5 CO 2 controlled 

chemostats in the lab of A. Engel at 

the Alfred Wegener Institute in 

Bremerhaven where joint 

collaborations are planned within 

project 1.2. (Photo: A. Engel) 

ocean acidification are limited to a few species, a research gap that this collaborative effort is 

intended to close through studies of key species and whole plankton communities in the field 

(projects 1.2). 

Broadening the taxonomic repertoire would also allow geographic predictions of sensitivities and 

compositional shifts in major plankton groups to a global scale pCO2 increases. Several 

subprojects under 1.1 address longer-lasting adaptations of phytoplankton and bacteria. Projects 

under 1.2 focus on the interaction between autotrophs and heterotrophs and especially the role of 

extracellular organic substances and aggregation. Finally project 1.3 models the impact of ocean 

acidification on the soft tissue pump (POC + DOC) as a potential removal process of carbon from 

the surface waters. 

Additionally to the pCO2 induced changes the combined effect of the temperature increase will 

be simulated under theme 1. Although most studies will be carried out in controlled labexperiments 

(e.g. chemostats from the AWI, Fig. 1.3) some studies will include open sea 

plankton to test the response of communities on OA. Within the BIOACID Project we have 

agreed upon several levels of CO2 which are: 280ppm (preindustrial) 380ppm (today), 560ppm 

(twice preindustrial), 700ppm (2.5 preindustrial and the level of the IPCC "business as usual" 

prediction for 2100), 980 – 1000ppm (3.5 preindustrial). Our experiments under Theme 1 will 

69

70 


follow these agreements and we plan joint experiments in several subgroups of Theme 1. 

Furthermore it is planned to perform experiments by carbonation and not acidification only for 

the pH reduction. This approach avoids conflicts from diverging culture setups. The effects of 

changes in the temperature which are predicted for the same time span as the OA are tested for in 

all the subprojects of theme 1. 

Overall the results of theme 1 will supply a basis for modelling and future prognoses on the 

ecological consequences of ocean acidification in biogeochemical model under theme 1.3. All 

subprojects will feed their results and findings into a better assessment of acidification and global 

warming on the so called “soft tissue pump”, especially the production and particle formation by 

means of TEP. A more reliable estimate of the biogeochemical feedbacks of changing 

productivity and export will be reached through a more realistic formulation of carbon production 

and export in global marine biogeochemical models. 

Project 1.1: Acclimation versus adaptation in autotrophs 

T. Reusch (University Münster/ IFM-GEOMAR Kiel), M. Hippler/J. LaRoche (University 

Münster/ IFM-GEOMAR Kiel), G. Jost/K. Jürgens (Leibniz Institute Warnemünde), M. Müller 

(IFM-GEOMAR Kiel), U. Karsten/T. Hübener (University Rostock) 

i. Objectives 

The five subprojects within Project 1.1 address the effects of ocean acidification (OA) and 

carbonation (rising seawater CO2 concentrations) on marine primary producers of diverse 

taxonomic affiliation, including chemoautotrophic bacteria, benthic and planktonic microalgae. 

Project 1.1 mainly studies effects of rising pCO2 on primary productivity and is mutually 

complementary to project 3.1 (and here particularly 3.1.1) that focuses on calcifying primary 

producers. The overall objectives are to: 

• Identify guilds and taxonomic groups of autotrophic organisms that are sensitive to 

acidification and carbon enrichment, compare treatments based on direct pCO2 

enhancement versus changes in seawater carbon balance 

• identify processes carried out by autotrophs that are affected by ocean acidification or 

increased pCO2 

• analyse short- and long-term physiological acclimation mechanisms that modulate 

carbon acquisition in response to carbonation 

• assess interactions between OA, nutrient status (in particular iron limitation) and ocean 

warming for photosynthesis and photosystem proteomics of important phytoplankton 

groups 

• establish selection lines in order to assess evolutionary changes in physiology and carbon 

acquisition under increased pCO2 

• analyze genetic and proteomic changes as a response to long-term exposure under 

increased pCO2 

• quantify shifts in taxonomic composition among important autotrophic groups as a result 

of acidification and carbonation, and possible second order effects on ecosystem 

performance 


Whereas effects of increased pCO2 in terrestrial systems have been intensely studied in the past 

15 years (summarized in Ainsworth and Long 2005), work on marine primary producers is in its


infancy. Because planktonic and benthic microalgae form the basis of marine food chains (with 

the exception of deep sea vent ecosystems), changes at the level of primary production may result 

in cascading effects throughout the food web (link to Theme 4). 

Increasing atmospheric pCO2 not only leads to drastic increases in seawater CO2 but also causes a 

reduction in pH (i.e. acidification), and comparatively minor increases in HCO3 - concentrations 

(i.e. carbonation). Recent data suggest that the primary short-term response of micro- and 

macroalgae is an enhancement of photosynthetic rates with rising pCO2. This may constitute an 

important negative feedback to anthropogenic CO2 emissions, potentially enhancing the so-called 

biological carbon pump (Riebesell et al. 2007). However, it is presently unclear to what extent 

and under what conditions carbon availability limits photosynthesis and growth of primary 

producers in the field. On the other hand, growth rates of calcifying phytoplankton species such 

as coccolithophores may be impaired by rising pCO2 due to lower carbonate availability 

(Riebesell et al. 2000) (link to theme 3), but recent data challenge this notion (Iglesias-Rodriguez 

et al. 2008). On objective of this project will be to perform critical experiments that compare 

different isolates with respect to their different (heritable) physiological performance. Moreover, 

we will compare different pCO2 enrichment protocols to evaluate whether conflicting results may 

arise from experimental procedures. 

Because the sensitivity to carbon enrichment differs widely among taxa, rising CO2 levels will 

alter competitive relationships and result in shifts of plankton species composition (Rost et al. 

2003). Currently, data on the responses of the diverse plankton groups to ocean acidification is 

limited to a few species, a research gap that this collaborative effort is intended to close. 

Broadening the taxonomic repertoire is also a prerequisite for geographic predictions of 

sensitivities and compositional shifts in major plankton groups to ocean acidification and 

increasing pCO2. The project will study the response of several important photoautotrophic 

groups such as non-calcifying planktonic diatoms and calcifying coccolithophores, benthic 

diatoms to increases in pCO2 within a common analysis framework and largely congruent 

experimental conditions. 

In contrast to photoautotrophs, consequences of increased CO2 concentration on 

chemoautotrophic organisms are probably quite different than on the surface phototrophic 

primary production. Pelagic oxic-anoxic transition zones (redoxclines) found in several coastal 

environments and marginal seas (e.g., Baltic Sea, Black Sea) are sites of significant 

chemoautotrophic production (dark CO2 fixation) (Taylor et al., 2001; Jost et al. 2008). By 

modifying the overall nutrient cycles, these redox-related transformations have an impact far 

beyond the spatial scale of the redoxcline (best known for N losses). It is not known how changes 

in CO2 concentration and pH affect the biogeochemistry of these areas. Because dark CO2 

fixation is based on chemical energy from reactions including mainly pH-sensitive substances 

(e.g. H2S, Mn, Fe), subproject 1.1.1 (Jost /Jürgens) hypothesizes significant changes in 

biogeochemical process rates related to chemolithotrophy, microbial key organisms and 

microbially based food webs. The thermodynamics and kinetics of some of the respiratory 

processes, for instance Fe(III) and Mn(IV) reduction by microbes, are highly pH dependent 

(Canfield et al. 2005). Changes in pH can induce changes in energy gain for microorganisms, in 

the delicate balance of biogeochemical redox sensitive processes, and even the element fluxes of 

the benthic-pelagic coupling in shallow euxinic systems. Subproject 1.1.1 will examine model 

microorganisms that have been shown to constitute key players for distinct biogeochemical 

transformations in pelagic redoxclines, and which had been successfully cultivated. 

Ocean acidification accompanied with several concomitant changes in abiotic parameters that can 

affect photosynthetic rates, one of which is the availability of dissolved iron. Despite its 

71

72 


abundance on earth, iron-deficiency is the most common nutritional deficiency in the world. Iron 

deficiency affects at least 10% of oceanic photosynthetic productivity and is most prevalent in 

vast high nutrient low chlorophyll (HNLC) regions of today’s oceans (Martin and Fitzwater 

1988, Behrenfeld and Kolber 1999). As consequence, photosynthesis and carbon assimilation are 

restricted by the low iron-bioavailability in these areas. 

The decrease in pH will reduce the precipitation of dissolved iron as Fe oxides while also 

reducing the binding of Fe to organic ligands, both factors being important in controlling Fe 

bioavailability. The effects of ocean acidification on the bioavailability of iron and hence, on 

phytoplankton productivity is currently not well understood. One hypothesis addressed by 

subproject 1.1.2 (Hippler/LaRoche) is that OA minimizes the pressure of iron limitation on 

primary productivity. Iron is essential for virtually all forms of life because it participates in 

electron transfer reactions as a crucial cofactor in enzymes that catalyze redox reactions. Since 

iron can also react with oxygen to generate cytotoxic agents, its accessibility within the cell has 

to be under tight homeostatic control, which requires complex regulatory mechanisms (Finney 

and O'Halloran 2003). 

At the molecular level, photosystem I (PSI) is a prime target of iron-deficiency, probably because 

of its high iron content. In the short-term, iron-deficiency leads not only to a pronounced 

degradation of PSI, but also to a remodeling of the PSI associated light-harvesting antenna 

(LHCI) (Moseley et al. 2002). Long-term adaptation to iron-deficiency even causes constitutive 

differences in the photosynthetic architecture as demonstrated for coastal and oceanic diatoms 

(Strzepek and Harrison 2004). The oceanic diatom strain contained up to fivefold lower PSI and 

up to sevenfold lower cytochrome b6f complex concentrations as compared to a coastal diatom. 

Surprisingly, the changes to the photosynthetic apparatus while decreasing the cellular iron 

requirements of the oceanic diatom maintain its photosynthetic performance relative to the 

coastal diatom. However, the lower iron-requirements of the oceanic diatom result in a reduce 

ability to acclimate to changing light conditions. Additionally, iron-limitation appears to directly 

interfere with carbon uptake and assimilation in phytoplankton (Schulz et al. 2007, Allen et al. 

2008) although the mechanisms by which this occurs are poorly studied. 

Insights into the interplay between carbon-and iron-availability and its impact on photosynthesis 

of primary producers in the ocean subjected to increasing dissolved CO2 are thus highly 

warranted. Subproject 1.1.2 will also provide a basis for better understanding the impact of open 

ocean iron-fertilization, a controversial practice which is pursued by some companies (Buesseler 

et al. 2008). In a high CO2 ocean the proposed study will bring additional knowledge essential in 

assessing the effectiveness of open ocean Fe fertilization as a strategy to mitigate global 

atmospheric CO2 increase. A recent transcriptomic study identified genes involved in silicon 

bioprocesses in T. pseudonana (Mock et al. 2008) proving the suitability of this technique in the 

analysis of the biology of diatoms. In Phaedactylum tricornutum, a species that shows a high 

tolerance to iron limitation, several genes involved in Fe acquisition and in the remodeling of the 

photosynthetic apparatus have been identified through a whole-cell system’s approach (Allen et 

al. in press). Several of these genes are absent in the T. pseudonana genome, pointing to a genetic 

basis for the phenotypic difference in their acclimation to iron limitation. An additional genome 

project for the oceanic diatom T. oceanica, has been initiated jointly at the IFM-GEOMAR and 

University of Kiel CAU. The outcome of this genome project together with the genome sequence 

of T. pseudonana and P. tricornutum will provide a solid basis for comparative transcriptomic 

and proteomics analyses (see also subproject 1.1.4). 

Most experiments done so far examining phytoplankton are restricted in time and are observing 

only a few generations of these single cell organisms. Regarding the fast generation time of most


phytoplankton groups (~one division per day) a possible acclimatisation and/or adaptation has to 

be considered for prediction of future phytoplankton response (Collins and Bell 2004). Current 

primary producers have co-evolved with a partial pressure of CO2 that never exceeded 300 ppm 

in the past 20 Mio years. Conversely, evolutionary changes are to be expected as a response to 

ongoing increases in pCO2. Such evolution in action is expected to be particularly rapid in 

microalgal species having short generation times, such as unicellular plankton and bacteria. 

Strain-to-strain differences of Emiliania huxleyi in their sensitivity to CO2 enrichment indicate 

standing genetic variation for traits related to carbon acquisition under altered CO2 chemistry 

(Delille et al. 2005). Interestingly, the only long-term experiment addressing evolutionary 

changes in a microalgal species over approximately 1000 generations revealed that the green 

unicellular algae Chlamydomonas rheinhardtii lost its carbon concentrating mechanism (CCM) 

in response to increased pCO2. These findings are in line with evolutionary theory, predicting that 

costly carbon concentration is no longer selected for in a CO2 rich environment. If long-term 

exposure to increases in pCO2 leads to degeneration of CCMs, then short-term responses of 

phytoplankton, for example as higher export production, may only be transient. On the other 

hand, under sexual recombination, positive selection of novel physiological function may also 

take place that produce physiological trait values outside the boundaries of original trait 

distributions (Reusch and Wood 2007) 

Because long-term effects are crucial but understudied in the marine plankton and 

chemoautotrophic bacteria, three subprojects will address long-term physiological responses 

(subproject 1.1.1 Jost/Jürgens; 1.1.3 Müller; 1.1.4 Reusch/Riebesell) and compare the 

reversible plastic acclimation response with longer-lasting adaptive changes. Changing responses 

of important target species to carbon enrichment have already been identified in pilot 

experiments (cf. Fig. 1.1.1a,b). Cultures of coccolithophores showed a higher sensitivity for high 

pCO2 regarding their division rates in comparison to short-term experiments (Riebesell et al. 

2000, Langer et al. 2006). Responses of phytoplankton to single parameters (e.g. temperature, 

pCO2 and calcite saturation state) are still in the progress of investigation. However, in nature all 

physical parameters are predicted to change more or less in the next centuries. Therefore, 

subproject 1.1.3 will also investigate any interaction effects of the two most rapid changing 

parameters (temperature and pCO2) on different phytoplankton key groups (diatoms, 

coccolithophores and calcareous dinoflagellates), which are the main primary producers 

(diatoms), the main pelagic producers of biogenic calcite (coccolithophores and calcareous 

dinoflagellates) and jointly contribute to the basis of the oceanic food web. A strong link will be 

forged to subproject 3.1.1 that focuses on changes in calcification rates of coccolithophores. 

In order to verify whether or not physiological changes are due to genetic adaptation, we aim at 

identifying the genetic basis of observed changes using state-of-the art genomic and 

transcriptomic techniques in two species where genomic and transcriptomic resources are readily 

available, Thalassiosira pseudonana and Emiliania huxleyi (subproject 1.1.4 Reusch 

/Riebesell). In these fast generation species, rapid evolutionary change may take place through 

the combination of any of the following three mechanisms (i) clonal selection, whereby preadapted 

multi-locus genotypes outcompete others by mitotic division and population increases 

(ii) de novo mutation in particular genotypic lines (iii) genetic recombination which may bring 

trait values beyond the range of values seen in the original population. Recombination is 

expected to bring together favourable mutations, while breaking up negative gene associations. 

For E. huxleyi, culture conditions are established that allow us to introduce intermittent rounds of 

sexual recombination during the course of the experiment (Laguna et al. 2001), permitting a test 

for the role of sexual reproduction in promoting evolutionary adaptation. Genomic approaches 

will be coordinated with subproject 1.1.2 for the diatom transcriptomic and proteomic analysis. 

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As additional response variables, information on the elementary composition of primary 

producers will provide important information for subproject 4.2.1. that addresses the effects of 

food quality on higher trophic levels. 

Fig. 1.1.1a: Division rates of three coccolithophore species 

under high pCO 2 (filled red) and ambient pCO 2 (green) 

over time. Open red circles indicate the transition from 

ambient to high pCO 2. The percental decrease in division 

rate is shown by the red numbers. Müller et al unpublished 

Fig. 1.1.1.b: Change of the optimal division curve of 

Emiliania huxleyi from ambient pCO 2 (green) to high 

pCO2 (red) in relation to different temperatures. Müller 

et al unpublished 

Benthic diatoms are another key primary producer guild with important ecological role on 

intertidal and shallow subtidal marine sediments along the German coastline of the North and 

Baltic Sea, being responsible for particularly high rates of elemental cycling and high primary 

productivity. These communities provide a major food source (e.g. fatty acids) for benthic 

suspension- or deposit-feeders, and act as control barrier for oxygen fluxes at the sediment/water 

interface, and as stabilizer of sediment surfaces against erosion by the excretion of extracellular 

polymeric substances (EPS) (Cahoon 1999). EPS are mainly composed of polysaccharides and 

proteins, and besides their role in biostabilisation of sediments they are involved in gliding 

motility and adhesion. 

Despite their importance for production and elemental cycling almost nothing is known on the 

ecophysiology and production biology per se, and even less on the interactive effects of rising 

CO2 concentrations and temperatures on microphytobenthic primary production (Forster et al. 

2006), a research gap that subproject 1.1.5. (Karsten/Hübener) intends to close. Building upon 

such ecophysiological data, the relationship between biodiversity of benthic diatom communities 

and their productivity will be assessed. Subproject 1.1.5. will also explore whether or not diatom 

taxa are valuable indicator species for environmental changes associated with OA, as has been 

shown for eutrophication before (EDDI: Battarbee et al. 2000, MOLTEN: Clarke et al. 2002, 

2003, Juggins 2004). Notably, these organisms possess small tolerance values for important 

control factors of water-quality (pH, nutrients, salinity). As a second step, it will explore how the 

diatom community changes as a response to predicted ocean acidification, in combination with 

temperature increases. 


Subproject 1.1.1: G. Jost has experience in the work with autotrophic prokaryotes and the 

analysis of their role in the biogeochemistry of pelagic redoxclines. K. Jürgens has extensive


experience in the analysis of prokaryotic and eukaryotic microorganisms, microbial food webs 

and trophic interactions, including model systems and field studies in diverse aquatic systems. In 

the last years a newly established group at the IOW in molecular and microbial ecology has made 

significant progress in the understanding of redoxcline microbial communities (Labrenz et al. 

2007, Jost et al. 2008), the identification of bacterial key players (Grote et al. 2007) and major 

transformation processes (Hannig et al. 2007). 

Subproject 1.1.2: Prof. J. LaRoche is an expert in marine phytoplankton ecology and 

ecophysiology. She has performed and participated in seminal experiments on the role of iron on 

phytoplankton productivity and physiology (LaRoche et al. 1996, McKay et al. 1997, Boyd et al. 

2000, Jickells et al. 2005, Allen et al. 2008). She is also an expert in diatom transcriptomics. M. 

Hippler is an expert in algal proteomics and physiology and particularly in adaptive remodelling 

of the photosynthetic apparatus to iron-deprivation (Moseley et al. 2002, Naumann et al. 2005, 

Allmer et al. 2006, Naumann et al. 2007). This combined expertise in physiology, transcriptomics 

and proteomics will ensure the successful complementation of the outlined working program. 

Subproject 1.1.3: M. Müller is currently working in the ESF project ’Casiopeia’ as a PhD, 

finishing his doctoral degree in autumn 2008. He previously worked on coccolithophores (Müller 

et al. 2008b) and gained over the last years a solid expertise in culturing phytoplankton in dilute 

batch cultures and semi-continuous cultures under a manipulated carbonate system (Müller et al. 

2008a). At the moment a variety of coccolithophorid species are cultured since several years and 

will be available for experimental work in this project. 

Subproject 1.1.4: The expertise and research interest of both applicants are mutually overlapping. 

U. Riebesell is an expert in marine phytoplankton ecology and ecophysiology (Riebesell et al. 

2000, Riebesell 2004, Riebesell et al. 2007). T. Reusch has broad experience in design and 

execution of selection experiments (Reusch and Wood 2007, Wegner et al. 2007), and in 

characterizing the genetic basis of phenotypic change (Wegner et al. 2003), including 

transcriptomic work (Reusch et al. 2008) 

Subproject 1.1.5: U. Karsten has worked with benthic diatoms from brackish and marine waters 

and successfully established a culture collection of polar diatoms at the University of Rostock. 

The research interest of Prof. Karsten is mainly related to the ecological and physiological 

performance of these microalgae under different environmental conditions, as well as to the 

development of new methodological approaches (e.g. Karsten et al. 2006; Wölfel et al. 2007; 

Gustavs et al. 2008). T. Hübener has a long record in studies on diatom ecology and taxonomy 

(Dreßler and Hübener 2006, as well as on diatoms as indicators for environmental change 

(palaeolimnology) (e.g. Adler and Hübener 2007, Hübener et al. 2007). The expertise of U. 

Karsten and T. Hübener is well reflected in numerous grants funded and refereed publications. 

iv. Work Programme, Schedules and Deliverables 

Work Programme – general 

The experimental levels and the rate of increase in pCO2 as well as other important experimental 

manipulations will be coordinated as much as possible especially with the experiments of 

Jost/Jürgens, LaRoche/Hippler, Müller, and Reusch/Riebesell. We also plan to share some of the 

experimental set-ups, in particular the chemostats, among the subprojects. All projects have 

agreed upon the specific levels of pCO2 for their experimental manipulation, namely the five core 

treatment levels 280ppm (pre-industrial), 380 (present day), 560ppm (2x pre-industrial), 700ppm 

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(IPCC business as usual for year 2100), 1000ppm (3x pre-industrial). These treatment levels 

should be established by a slow change from the present day level. 

Subproject 1.1.1 Impact of changing pCO2 and temperature on a chemolithoautotrophic 

Epsilonproteobacterium from a pelagic redoxcline 

G. Jost & K. Jürgens 


The response of microbial populations at pelagic redoxclines to changes in single parameters 

(pCO2, pH, and temperature) will be studied in controlled experimental laboratory systems 

(diluted batch and chemostat cultures, including a comparison between both systems). The effects 

of ambient pCO2 and slowly increasing pCO2 on the performance of a representative 

chemoautotrophic prokaryote will be compared. Additionally, it will be investigated whether 

increasing pCO2 in combination with decreasing pH or only changes in the pH are the important 

factors. The selected model organism epsilonproteobacterium strain GD1, which is abundant in 

the redoxclines of the central Baltic Sea (Grote et al. 2007) will be used for selected process 

studies (e.g. chemolithotrophic denitrification). The changing performance of this strain in 

response to the investigated factors will be compared to relative changes in the performance of 

other single organisms (1.1.3, 1.1.4) as well as whole (bacterial) communities (1.1.2, 1.1.5, 

1.2.4). The performance of the microorganisms will be assessed as specific growth rate, 

elemental composition (CNP) and related chemical transformations (e.g., sulphur oxidation, 

denitrification). To differentiate between phenotypic and genotypic long-term adaptations 

molecular biological methods will be applied (e.g. differential gene expression of functional 

genes) in cooperation with other projects. 

In order to assess the acclimation potential, different rates of increase pCO2 attaining the final 

treatment levels will be compared. Finally, in line with sub-projects 1.1.3 and 1.1.5, the 

combined versus single effects of temperature and pCO2 will be assessed in factorial experiments. 

The results will be used for biogeochemical modelling, including the evaluation of the influence 

of proton activity on biogeochemical processes. Additional, a comparison of the bacterial 

population at the end of the long-term experiments grown under different conditions will be done 

not only by investigating phenotypic performances but also genotypic differences (e.g. 

differential gene expression of selected functional genes). A comparison of the results of the 

reaction a single bacterium will show on changing conditions related to climate change with 

observations of carbon dioxide fixation in CO2-enriched sediments and the appearances of 

different phylogenetic groups of bacteria including epsilonproteobacteria (subproject 4.1.4.1) 

may give a hint on possible extrapolations of the results even to sediments. 

Work Schedule 


Set-up of culturing facility, instrument 

calibration 




Diluted batch experiments 

Long-term chemostat experiment 

Sample processing and measurements 

Combined CO2/tempertaure perturbation experiments 

Molecular investigations 

Data analysis, statistical evaluation, 

data interpretation 


results at conferences, PhD paper 


Milestones (1.1.1) 

- Implementation of experimental facility for controlled continuous cultures month 9 

- Experimental data set on CO2/pH sensitivity of epsilonproteobacterium GD1 month 18 

- Data set on synergistic effects of CO2 and temperature on strain GD1 month 24 

- Evaluation of combined data sets, sensitivities and uncertainties month 33 

Subproject 1.1.2 The interplay between carbon-and iron-availability and its impact on 

photosynthesis of primary producers in the ocean 

J. LaRoche & M. Hippler 

Work programme 

We will study the effects of ocean acidification on photosynthesis and carbon assimilation in 

diatoms, exposing them to iron-limited and replete conditions. Physiological approaches will be 

combined with systems biology approaches where transcriptomics and proteomics will be 

applied. (i) Physiological measurements of growth and photosynthetic performance for a oceanic 

and costal diatom strain will be conducted at distinct CO2 partial pressures of 380, 700 and 1000 

ppm combined with iron-sufficient and –deficient conditions. (ii) Comparative transcriptomic 

profiling of the two diatom strains T. pseudonana and T. oceanica will be performed under the 

outlined physiological conditions taking advantage of 454 DNA sequencing (Eveland et al. 2008) 

to elucidate dynamics in gene expression in regard to different carbon and iron resources. (iii) 

Comparative and quantitative proteomics will be performed to unravel alterations in the proteome 

with a focus on plasma and thylakoid membranes in response to iron and carbon availability. In 

particularly we will concentrate on changes of the photosynthetic apparatus. Here we will take 

advantage of high precision and resolution mass spectrometry using an LTQ-Orbitap mass 

spectrometer (ThermoFisher) and proteotypic peptide profiling. 

The project is complementary to 1.1.1 and 3.3 which also study the effect of OA and pCO2 

increase on the speciation of trace metals and their effect on chemoautotrophs and calcifyiers. 

The result of our study on short term effects of high CO2 on the photosynthetic apparatus of 

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diatoms can be used to help predict the long term genetic effects of elevated CO2 studied in 

1.1.4. The results will also be used in biogeochemical models (1.3). 

Work Schedule 


Establish trace-metal buffered cultures 

Experiments with T. oceanica 

Experiments with T. pseudonana 

Establishment of proteomic techniques 

with T. oceanica 

Transcriptomic analysis T. oceanica 

Data analysis & synthesis 

Establishment of proteomic techniques 

with T. pseudonana 

Transcriptomics T. pseudonana 

Data analysis & synthesis 

PhD. Thesis preparation 



- Establishment of semi-continuous trace metal-buffered culture system and CO2 levels month 6 

- Experimental data set with T. oceanica grown at varying CO2 and iron levels month 18 

- Experimental data set with T. pseudonana grown at varying CO2 and iron levels month 27 

- Evaluation of combined data sets month 36 

Subproject 1.1.3 Long-term response 

multidimensional approach 

M. Müller 

of phytoplankton on climate change: a 


For each phytoplankton species (Thalassiosira pseudonana, Emiliania huxleyi and 

Thoracosphaera heimii) incubations will be run in duplicate under lab-controlled conditions 

(light, temperature, nutrients and carbonate system) over 1.5 year what corresponds to 500 to 

1000 generations depending on the species. For this purpose 24 chemostat units will be combined 

in a computer-controlled incubation system where the pCO2 and the temperature can be slowly 

increased to a certain level. The newly constructed CO2 aeration system at the IFM-GEOMAR is 

a ideal tool to set up a chemostat system where the carbonate system can be controlled in a


appropriate manner. The system will be monitored by dissolved inorganic carbon, alkalinity and 

nutrient analysis carried out in the lab of U. Riebesell. Primary response variables that are 

coordinated throughout projects 1.1.2 – 1.1.5 are division rates, photosynthesis, calcification rates 

and chlorophyll production. Photosynthesis and calcification rates will be measured at the isotope 

lab using radioactive 14 C accompanied by carbon acquisition determination with the Inlet-Mass- 

Spectrometry (MIMS) in cooperation with K. Schulz (3.1.1). Division rates and chlorophyll 

autofluorescence will be determined by flow cytometry. Genetic analysis at the end of the 

experimental exposures will be realized in cooperation with J. LaRoche, T. Reusch and M. Bleich 

(1.1.2; 1.1.4 and 3.1.4), additionally we will identify stress proteins in the named species in 

regard to pCO2 and temperature (LaRoche, 1999). Calcite analytics (δ 13 C, δ 18 O, δ 11 B) can be 

performed in the geochemistry lab at the Ruhr-University Bochum (3.5.2) and in collaboration 

with subproject 3.5.3 (S. Meier). During the acclimation phase we plan to analyze the expression 

of stress proteins such as heat shock proteins (hsps) in order to identify surrogates of the 

acclimation response. 

Work Schedule 


Establishing chemostats 

Long-term CO2 enrichment exps 

Physiological measurements 

Identify stress proteins in dilute batch 

cultures 

Quantification of stress proteins in 

chemostats 

Synthesis of results & analysis 





- Completion of chemostat system and data set on facility-accuracy month 9 

- Identification of stress proteins month 15 

- Data set on synergistic effects of CO2 and temperature under long-term conditions month 30 

- Evaluation of combined data sets (with all collaborators), sensitivities and month 32 

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Subproject 1.1.4 Rapid evolution of key phytoplankton species to a high pCO2 ocean 

T. Reusch & U. Riebesell 


We propose to establish replicated selection lines with large population sizes (>10 6 cells) in two 

important members of the marine phytoplankton, a diatom (Thalassiosira spp.) and a 

coccolithophorid (Emiliania huxleyi) under ambient (= present day; 380 ppm) and increased 

pCO2 (~700 ppm/1000 ppm). Ideally, pCO2 will be increased by bubbling enriched atmosphere 

into culture water. Experiments will start from novel isolates obtained from natural 

phytoplankton assemblages (Rynearson and Armbrust 2000). Control populations /lines in all 

diversity levels will also be established and subjected only to laboratory selection under ambient 

pCO2. Cultures will be run for at least 500 generations (~6-12 months), with some replicates 

being shared among the subprojects (i.e. 1.1.3 Müller). 

Using diagnostic microsatellite markers in combination with quantitative real-time PCR, we will 

closely follow the genotypic composition of all multi-clonal populations to detect genotypic 

selection, i.e. the dominance of certain pre-adapted genotypes over others. Results from this part 

of the project will be compared to results from project 4.1.2 (M. Wahl) that addresses the role of 

genetic diversity for juvenile survival in invertebrates. Because recombination may enhance 

selection responses, in E. huxleyi, half of the experimental lines will undergo sexual reproduction 

every 10 – 50 mitotic cells cycles (Laguna et al. 2001). The published methods may have to be 

modified in order to allow several (~10-20) intermittent sexual generations. Relative fitness of 

selected and ambient replicates will be assessed singly, and as direct competition experiments 

under a reciprocal combination of selection regimes. Response variables to be monitored every 

50 mitotic generations will be coordinated throughout 1.1.2 – 1.1.4 and include photosynthetic 

rates, pigment content/composition, division rates, chemical stoichiometry, calcification rates and 

stable isotope composition. The latter data will be interconnected with project 4.2.1 (M. 

Boersma) that studies the effects of changing chemical composition of phytoplankton for 

zooplankton. As in subproject 1.1.3, calcification rates will be measured at the isotope lab using 

radioactive 14 C accompanied by carbon acquisition determination with the Inlet-Mass- 

Spectrometry (MIMS) in cooperation with K. Schulz (3.1.1). In order to identify the genetic basis 

of adaptive evolution in a high pCO2 ocean, we will assess the transcriptomic response of both 

selection lines (ambient / high pCO2) in full combination with the actual test regime (ambient / 

high pCO2). In E. huxleyi, transcription profiling will be done using tagged 3’-expressed 

sequence tags in combination with direct 454 DNA sequencing (Eveland et al. 2008). In 

Thalassiosira, we use the whole genome tiling array developed by (Mock et al. 2008). Changes 

in transcriptomic response in Thalassiosira will be compared to responses obtained in project 

1.1.2 (J. LaRoche) where the response to iron deficiency is studied. Interpretation of data on 

evolutionary responses will be closely discussed with the long-term data on the fossil record 

obtained through project 3.5.2 (J. Mutterlose).

Work Schedule 



Collect field isolates, establish 

Selection lines 

Induction of sexual reproduction 

Establish Q-PCR to discriminate 

genotypes 


Run evolution experiment 

Test for adaptation in reciprocal 

experiment 

Perform transcriptomic analyses 




results at conferences, PhD thesis 

preparation 


Milestones (1.1.4.) 

- Selection lines are running month 9 

- Sexual Induction possible month 12 

- Experimental evolution lines finished (~1000 generations) month 24 

- Transcriptomic data obtained month 26 

- Bioinformatic analysis of evolutionary changes month 33 

Subproject 1.1.5 Interactive effects of CO2 concentration and temperature on 

microphytobenthic biodiversity and ecosystem function 

U. Karsten, T. Hübener 


The main goal is to evaluate the interactive effects of rising pCO2 and temperature on 

microphytobenthic ecophysiology, biodiversity and primary production under controlled 

laboratory conditions and at representative shallow water stations in the Baltic Sea. In order to 

test species specific responses of benthic diatoms to a range of CO2 concentrations and 

temperatures we expose unialgal cultures of dominant species from experimental habitats in 

laboratory cultures and measure photosynthesis, division rates and primary production under 

different light, CO2 and temperature conditions using fluorimetric techniques and O2 optodes. For 

the improvement of the methodological approach strong collaboration with project 0.3.2. 

(Apostolidis/Huber, Presens) is intended. The response parameters will be compared to relative 

changes in the performance of other aquatic organisms and communities within theme 1.1., 

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particularly project 1.1.1 (Jost/Jürgens) as well as to other macrobenthic primary producers in 

project 4.1.1 (Asmus). As a next step, in order to study the community level response, we will 

artificially mix communities with up to 10 diatom taxa. Species selection will be guided by 

abundance estimates from natural sediment communities of the shallow Baltic Sea. Community 

experiments will allow us to test the hypothesis that there is a predictable shift in taxonomic 

distribution upon OA, possibly interacting with temperature, and driven by different 

ecophysiological requirements obtained from previous experiments. Microphytobenthic primary 

production will be assessed via area-based biomass determination (standing stock, chlorophyll a 

content in sediment cores). We also apply in situ primary production measurements using new 

benthic chambers with planar O2 optodes and puls amplitude modulation (PAM) fluorometry. 

There are many scientific overlaps to projects 3.4.1 and 3.4.2 concerning the metabolic activity of 

benthic microorganisms and their effects on the water column, sediment and porewater 

chemistry. 

Work Schedule 


Isolation, identification,cultivation of 

diatom species 

Assessment of natural diversity 

Ecophysiological measurements 

Mesocosm experiments on primary 

productivity versus diversity 

Data analysis & completion of PhD 

thesis 


Milestones (1.1.5.) 

- Field data on natural biodiversity month 6 

- Unialgal cultures for experimental studies month 9 

- Ecophysiological response profiles for CO2 x temperature conditions month 18 

- Experiments on primary production as function of pCO2 and temperature month 24 

- Mesocosm data on diversity versus primary productivity month 30 

- Statistical evaluation of data, Interpretation, conclusion month 34 

vi. Budget and Budget Justification 


1.1.1 (PhD) 

1.1.1 (HiWi) 

1.1.2 (PhD) 

1.1.3 (Post Doc) 

1.1.3 (HiWi) 

First Year Second Year Third Year Total

1.1.4 PhD (Reusch) 

1.1.4 Tech (Riebesell) 

1.1.5 (PhD, tech supp.) 

Subtotal 

Consumables 

1.1.1 

1.1.2 Kiel 

1.1.2 Münster 

1.1.3 

1.1.4 

1.1.5 

Subtotal 

Travel 

1.1.1 

1.1.2 

1.1.3 

1.1.4 

1.1.5 

Subtotal 

Investment 

1.1.1 

1.1.2 

1.1.3 

1.1.4 

1.1.5 

Subtotal 

Other costs 

1.1.1 

1.1.2 

1.1.3 pub charges 

1.1.4 (Münster) 

1.1.5 

Subtotal 

Total 

Budget justification 1.1.1. 



Personnel costs: One PhD-student, will establish an anaerobically run chemostat system, 

allowing the application of changing pCO2 for long term cultivation of the isolated 

epsilonproteobacterium strain GD1. During the period of establishing the chemostat system 

already diluted batch experiments will be done to investigate the effect of changing pCO2 versus 

only pH changes on the growth of the selected strain. The proposed student assistant will help in 

the preparation of media for cultures and during sampling of the chemostat experiments as well 

as in preparation of samples for elemental and flow cytometric measurements. 

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Consumables: Will cover the preparation of media for the chemostat and batch cultures, 

measurements of bacterial concentrations by flow cytometry, comparisons with epifluorescence 

microscopy, estimation of elemental composition of the bacterial biomass, estimations of the 

concentrations of the chemical reactants, radioactive substrates and chemicals for rate 

measurements, chemicals for molecular biological methods to estimate genotypic differences. 

Travel: Project collaborators will take part in workshops ensuring comparative measurements of 

the pCO2 and the establishing the proposed concentrations in the different experimental systems 

as well as in workshops concerning the construction and running of chemostats with constant and 

slowly increasing pCO2. Especially, regular exchanges will take part during the establishing of 

the chemostat system with the WP (M. Müller). Participation within regular annual project 

meetings as well as in international conferences, exchanging ideas and presenting the results of 

the project, is also necessary. 

Budget justification 1.1.2 

Personnel costs: The Ph.D. student position will be split between the two institutions as follows: 

2 years for J. LaRoche in Kiel and 1 year for M. Hippler in Münster. The Ph.D. student from the 

LaRoche lab will be responsible for setting up the trace metal buffered cultures at the various 

CO2 and Fe levels and for making the basic physiological measurements (photosynthesis, growth 

rate, iron uptake, flavodoxin accumulation, PAM fluorometry). The student will also collect the 

samples for transcriptomics and proteomics. He will be responsible for the RNA extraction and 

transcriptomics analysis. 

The Ph.D. student (one year) in the Hippler lab will be responsible for the analysis of the 

proteomics samples grown at the various CO2 and Fe levels. 

Consumables: The consumables will be used to pay for the cost of sequencing with the 454 

tagged pyrosequencing (estimated at for 6 conditions run in duplicates) and for the 

proteomics analysis. 


Personnel costs: Post Doc is necessary to construct and maintain the required chemostat system. 

Additionally, to culture the organisms over long time scales reliable skills are essential what is 

not fulfilled by a PhD student and even a Post Doc will need additive help (student assistance). 

Consumables: To sample and monitor the chemostat system a bunch of consumables are needed 

and analysis is the base for publications. 

Travel: Visiting international conferences is essential to present the latest results and to keep in 

contact with the scientific community. 

Investment: The construction of the chemostat system will provide a unique platform to 

investigate the combined effect of CO2 and temperature on phytoplankton under completely 

controlled lab conditions over a long time scale. Accessorily, it provides the possibility to 

identify acclimatization and/or adaptation in marine organisms over time. This system has to be 

controlled by a set of intertwined computer framework. Other costs: Flow cytometric 

measurements are needed to monitor the cell abundance in the chemostat system.



Personnel costs: One technician ( ) for maintaining the selection lines, 

will be shared with project 1.1.3. Selection lines need to be replicated under different pCO2 

conditions. For E.huxleyi, they will be combined with inducing intermittent sexual phases. For 

both species we plan to establish 48 batch cultures of 1.5 L in total which will be very time 

consuming. One PhD student will focus on the induction of sexual phases, on developing the Q- 

PCR assays, and on trasncriptomic analyses at the end of long-term experiments. 

Consumables: for general maintenance of cultures, continual physiological 

measurements, genetic analyses for determining genotypic composition (clonal selection) using 

Q-PCR and clone specific microsatellites 

Travel: paid by institute. Investment: none required 

Other costs: replicated transcription profiling of evolved and non-evolved strains under ambient 

and novel conditions (complete reciprocal design; for transcriptomic analyses using 

454 tagged pyrosequencing (Emiliania), for whole genome transcript profiling 

(Thalassiosira) using whole genome tiling array designed by (Mock et al. 2008). 


Personnel costs: One PhD-student ( ) for evaluation of morphometric and molecular 

diversity shifts in different microphytobentic habitats (field work) and artificial lab-communities 

(mesocosms, incl. isolation, cultivation) under different pCO2 and temperature conditions. 

Students technical support ( ) for maintaining stock cultures, growth experiments under 

different light, pCO2 and temperature conditions(growth fluorimeter for benthic microalgae) and 

primary production measurements with O2 optodes with unialgal cultures and defined mixed 

benthic diatom communities. 

Consumables: for chemicals, glass and plastic vessels for culturing, microscopy, 

chemical analysis, molecular biology. 

Travel: for field work, participation in workshops ensuring comparative measurements 

of pCO2, in annual project meetings and in international conferences presenting results of the 

project. 

Investments: once; CO2 aeration system for the mesocosm, growth and primary 

production measurements, multichannel O2 optode system. 

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Project 1.2 Turnover of Organic Matter 

Anja Engel (AWI, Bremerhaven), Maren Voss (Leibniz Institute Warnemünde), M. Nausch/ G. 

Nausch (Leibniz Institute Warnemünde), Hans-Peter Grossart (IGB, Berlin), Laurenz Thomsen/ 

Giselher Gust (Jacobs University Bremen) 

i. Objectives 

This projects aims to elucidate the effects of ocean acidification on the turnover of organic matter 

in pelagic ecosystems. During field and laboratory studies, we will quantify and characterize how 

the production, exudation and microbial processing of organic matter (OM) respond to changes in 

seawater pCO2 and pH. Specifically, the project will investigate: 

• physiological and community structure responses of marine microbes. This will enable 

us to differentiate between functional and structural changes of planktonic communities. 

• the turnover of biochemical key components, such as polysaccharides, proteins and 

organic phosphorus compounds, and the resulting changes in the C:N:P stoichiometry of 

organic matter. 

• how the remineralisation and final deposition of sinking OM, i.e. aggregates, will 

become affected by potential changes in mineral ballast. 

Our ultimate goal is to contribute to a better understanding of the direction and strength of 

biogeochemical feedback processes in the future ocean. We aim to generate reliable data on 

highly variable chemical and microbial processes in order to provide a better basis for modelling 

and future prognoses on the ecological consequences of ocean acidification. 


There is now increasing awareness that ocean acidification will affect marine algae and biogenic 

production in the ocean (Orr et al. 2005, Arrigo 2007, Riebesell et al. 2007, Fabry et al. 2008). 

Direct responses of the microbial food-web to changes in seawater pCO2 and pH, or in the supply 

with organic resources are yet little exploited. In order to estimate the sensitivity of 

biogeochemical cycles to ocean acidification, an improved understanding of potential changes in 

the turnover of organic matter during production and decomposition processes is urgently 

needed. 

Recent studies indicated that the production of acidic polysaccharide particles, also known as 

transparent exopolymer particles (TEP), is sensitive to changes in seawater CO2 concentration 

(Engel, 2002; Engel et al., 2004a, Mari 2008). Acidic polysaccharides are exuded by 

phytoplankton cells as a carbon-rich ‘overflow’ product of photosynthesis under nutrient 

depletion (e.g. Obernosterer and Herndl, 1995; Biddanda and Benner, 1997; Søndergaard et al., 

2000), or derive from bacterial oxidation of dissolved carbohydrates (Giroldo et al., 2003). 

Acidic polysaccharide can facilitate the bonding between organic components and therewith 

affect particle stickiness, and the partitioning between the pools of dissolved and particulate 

organic matter, and organic matter export (Logan et al., 1995; Engel 2000, Passow 2002). On the 

other hand, varying concentrations and chemical signatures of released organic matter potentially 

can affect the activities and community composition of heterotrophic bacteria (e.g. Biersmith and 

Benner, 1998; Grossart et al., 2005, 2006a). Thus, besides exudation and aggregation, the


concentration of polysaccharides in seawater is determined by bacterial decomposition processes, 

which may respond differently to ocean acidification (Piontek et al., 2007a, b). 

In general, elemental cycles do not exist independently from each other in marine ecosystems. 

The cross-linking of C, N and P-cycles is pronounced in heterotrophic bacteria, as the availability 

of DOC has consequences for P utilization and can decide if DIP or DOP is the preferred 

compound (Hoppe and Ullrich 1999, Nausch and Nausch 2004, 2007). Changes in carbon 

exudation due to ocean acidification may therefore affect the phosphor demand of bacteria 

(Tanaka et al. 2007). On the other hand, N2-fixation rates can also be enhanced under high CO2 

(Hutchins et al. 2007, Ramos et al., 2007). DOM release can reach up to 40% of assimilated N 

(Bronk et al., 1994), indicating that DON- and DOC- release are tightly coupled (Bronk. and 

Ward, 1999). The regulation of release and uptake processes in the environment is poorly 

understood (Bronk et al., 2007). Potentially, competitive interactions for resources may define the 

community of autotrophs and small heterotrophs in the field. To reliably predict organic matter 

turnover in the future, a better understanding on factors regulating DOM release and uptake and 

their sensitivity towards changing pCO2/pH are necessary. 

Export of organic carbon into the benthic boundary layer (BBL) is enhanced by acidic 

polysaccharides and by mineral ballast (Passow 2002, Armstrong et al., 2002; Klaas and Archer 

2002). Recent studies indicate that ballast through calcification can either be decreased or 

increased due to CO2-induced changes in seawater chemistry (Fabry, 2008). Until final burial 

and once on the sea floor, organic aggregates are more easily remobilised into the benthic 

boundary layer than the bulk sediments beneath, and are frequently resuspended and hence 

modified in the water column (Thomsen et al., 2002, Keil et al., 2004). After an extended 

residence time of weeks to month, oxygen consumption within most organic aggregates is often 

similar to that of the background sediment and produces a minimal impact on sediment 

mineralization rates (Beaulieu and Smith 1998.). The part of this organic matter that seems too 

refractory to be recycled is buried in ocean sediments, sequestering carbon for long time periods 

and consequently influencing atmospheric carbon dioxide concentrations (Hedges et al., 2002). 

Bioavailability of aggregates in the BBL may be more related to their organo-mineral content 

than to the molecular composition of organic matter and consequently changes in mineral ballast 

will affect future carbon sequestration. 


Subproject 1.2.1: A. Engel (AWI): A. Engel has comprehensive experience on studying CO2 

effects on marine ecosystems, mineral ballasting, and in particular on organic matter exudation 

(i.e. Engel 2002, Engel et al. 2004a,b, Engel et al. 2005, Engel et al. 2008, Engel et al. in press). 

Since 2005, she leads a Helmholtz Young Investigators Group to investigate global change 

effects on marine biogeochemistry. The group has been involved in several national and 

international programs dealing with potential effects of global change (PeECE, PEACE, 

AQUASHIFT, SOPRAN, and EPOCA). A Ph.D. thesis has been performed dealing with the 

effects of ocean acidification on organic matter enzymatic hydrolysis and bacterial 

decomposition in calcifying algae. 

Subproject 1.2.2: M. Voß (IOW): M.Voß co-leads the subproject in the SOPRAN project 

“Ecosystem response to CO2 enrichment” where the role of nitrogen fixers under high CO2 is 

studied in free drifting mesocosms. Within the WGL network, TRACES she supervises a PhD 

thesis on the DON release of cyanobacteria and the transfer of fixed nitrogen into higher trophic 

levels. She worked on the regulation of N-fixation in various regions (Wasmund et al., 2005; 

Wasmund et al., 2001, Voss et al., 2006, Capone et al., 1998, Voss et al., 2004). Voss leads the 

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88 


“N-cycle group” at the IOW consisting of 5 externally funded people with a special focus on 

marine nitrogen cycling, nitrogen fixation and stable isotope ecology. 

Subproject 1.2.3: M. and G. Nausch (IOW): Phosphorus dynamics in surface water was a major 

topic of research during the last years and resulted in the formation of an interdisciplinary 

“phosphorus” group (2 senior scientists, 1PhD, 3 diploma students, 1 technician). Substantial 

investigations on alkaline phosphatase activity as an indicator for P-limitation were done (Nausch 

1998; Nausch 2000; Nausch and Nausch 2000). A new method for the determination of 

bioavailable DOP (BAP) was established (Nausch and Nausch 2004) allowing investigations of 

BAP and its importance for phytoplankton and bacterial nutrition (Nausch and Nausch 2006, 

2007). Currently, they participate in the SOPRAN-project “Ecosystem response to CO2 

enrichment”. 

Subproject 1.2.4: H.-P. Grossart (IGB): H.-P. Grossart has intensively worked on 

phytoplankton-bacteria interactions, especially on effects of phytoplankton release, its subsequent 

aggregation and on mineralization by specific bacterial communities (Grossart et al., 2005, 

Grossart et al., 2006a+b, Grossart and Simon 2007, Grossart et al. 2007). He has participated in 

the PeECE II and III studies, collaborating with national and international partners (Løvdal et 

al.,2008, Tanaka et al. 2008, Allgaier et al. 2008, Riebesell et al. 2008). Grossart et al. (2006c) 

showed effects of pCO2 on bacterial abundance and activities, which was mainly linked to algal 

and particle dynamics. H.-P. Grossart is currently collaborating with M. Lunau, A. Engel, and M. 

Voss in the SOPRAN mesocosm studies. 

Subproject 1.2.5: L. Thomsen/ G. Gust (Jacobs University): L. Thomsen has long experience in 

BBL studies at continental margins, (OMEX, HERMES, ESONET, CORAMM). G. Gust has 

developed and utilized high pressure laboratories with decompression free access and benthic 

chambers (LOTUS, OMEGA, GRAL, SEDYMO). Joint studies Thomsen/Gust indicate that with 

increasing residence time within the benthic boundary layer, a carbon protection mechanism is 

built up through further aggregation processes, which reduces their bioavailability; including a 

pressure effect (Thomsen and McCave, 2000; Thomsen and Gust, 2000; Thomsen et al., 2002; 

Thomsen, 2004; Tengberg et al., 2004; Mendes et al., 2007, Kleeberg et al., 2008; Bigalke et al., 

in press). 

iv. Detailed Description of the Project Work Plan 

Description of common experiments and field studies 

To establish a close collaboration between partners within this project, IOW, IGB, JU, and AWI 

will perform joint laboratory experiments, using natural and cultured key microorganisms 

(diatoms, cyanobacteria, heterotrophic bacteria) under CO2 perturbation. The group of A. Engel 

(AWI) provides a system of five CO2, T, nutrient and light controlled chemostats (10L, Fig. 1.3). 

Chemostat experiments will be set up for five different core CO2 concentration with two groups 

of primary producers (diatom and nitrogen fixer) in two separate experiments in the 1 st and 3 rd 

year. Those experiments will run over 3 month periods with sampling of subprojects 1-4 for their 

respective programs (see below). We will study changes in chemical composition as well as 

microbial colonization and decomposition of sinking aggregates in pressure chambers established 

by Thomsen/ Gust at JU. In the 2 nd year two cruises into the central Baltic Sea are planned for the 

investigation of the responses of natural plankton population to CO2 and pH perturbations during 

onboard experiments. The cruises will be organized by IOW. Additional laboratory experiments 

will be performed to investigate combined effects of CO2/ pH and temperature, pressure, light 

and nutrient availability.


Subproject 1.2.1 Production and decomposition of exudates 

A. Engel (AWI) 

We propose to test the following hypotheses: 

• An increase in seawater CO2 concentration stimulates the production and 

exudation of exopolymer substances, in particular polysaccharides, by autotrophic 

organisms 

• CO2 induced ocean acidification affects the enzymatic hydrolysis of organic matter 

and processes to follow, resulting in changes of the microbial decomposition of 

exopolymer substances 

• The impact of ocean acidification (OA) on production and decomposition 

processes is different for different organic matter components (carbohydrates, 

proteins). 

Ocean acidification may therefore affect the turnover organic matter in the water-column 

differently, with adverse biogeochemical consequences and feedbacks. During the joint activities, 

1.2.1 will determine the production and decomposition of exopolymers, i.e. transparent 

exopolymer particles (TEP), coomassie stained particles (CSP), and bulk organic matter (POC, 

PON, DOC, DON). To better describe processes relevant for biogeochemical feedbacks we will 

measure the chemical composition of exudates, i.e. size fractionated neutral and acidic sugars, 

and amino acids, together with rate measurements of processes governing their production, i.e. 

14 C-exudation, DOM aggregation. Information on the chemical composition of algal exudates at 

changing pCO2 will further be used to evaluate the food quality of OM for heterotrophic 

organisms in collaboration with 4.2.1. 

For comparison between different phytoplankton functional groups exudates production and 

composition will be determined in selected samples provided by 4.2.2. To study pH effects on 

organic matter decomposition, we will investigate the potential activities of major bacterial 

exoenzymes (glucosidases, peptidases, phosphatases, lipases), as well as bacterial production 

(thymidine/ leucin), and respiration. A comparison between pH effects on the activities of 

bacterial exoenzymes and of those produced by grazers will be made in collaboration with project 

4.1.1. Substrate uptake (carbohydrates) by bacteria will be investigated in collaboration with M. 

Voss, M. Nausch and H.-P. Grossart. 

Information on algal performance during chemostat experiment will be used by modelling 

projects in theme 5, in particular in project 5.1, where results on the production of DOM and its 

dependence on pH, temperature, and nutrient availability will be used for model refinement, and 

by 5.2., where information on the production and decomposition of exudates will be included in a 

Bayesian meta-analysis of experimental data 

This subproject will also participate in the Svalbard mesocosm experiment planned within the 

frame of the European project EPOCA to investigate effects on arctic pelagic communities in 

2009. 

89

Work Schedule 1.2.1 

90 



1.2.1 I II III IV I II III IV I II III IV 

lab work 

field work 

Chemostat-Exp. 

Synthesis and Publication 


- Set-up of improved PCHO/AAS analysis (microwave hydrolysis) month 6 

- Dataset from Mesocosm-Experiment Svalbard month 9-10 

- Chemostat data set on CO2/pH sensitivity of diatoms month 18 

- Field data set from spring/ summer cruise to the Baltic Proper month 24 

- Chemostat data set on CO2/pH sensitivity of cyanobacteria month 33 

Subproject 1.2.2 Dissolved organic nitrogen release and uptake under stress 

M. Voß (IOW) 

To better understand the release and uptake of DON and uptake of DOC of primary producers 

under different pCO2 concentrations and under nutrient stress situations several approaches will 

be used. 1. In the IOW batch cultures of diatoms and cyanobacteria under variable pCO2 will be 

established together with M. Nausch. The different nutrient stress situations will be limitation of 

PO4 3- for N2-fixers and PO4 3- and NO3 - limitation for diatoms. Additional to this stress 3-5 pCO2 

levels will be established. A suite of variables will be then be analysed; nitrogen fixation and 

primary production rates, POC/PON, DON/DOC concentrations, DON release and DON/DOC 

uptake with 15 N / 13 C labelled substrates. These experiments aim to separate between stresses from 

nutrient limitations from the one by acidification 2. We will participate in two 3-4 month long 

chemostat experiments at the AWI (see workplan 1.1.1) with again the same group of 

phytoplankton organisms and make the same analysis as before. The chemostats will be 

investigated by Engel, Nausch, and Grossart at the same time to generate a broad overview over 

production and degradation processes. 3. In the field a spring bloom of diatoms and summer 

bloom of cyanobacteria will be studied. Here we are faced with a whole plankton community 

which will affect rates and processes. All groups will cooperate again during field sampling and 

during large volume (0.1m³) mesocosm experiments on board where the plankton community 

will be studied under enriched pCO2 conditions. 

Relations to theme 4 are given through the study of C:N ratios in phytoplankton species which 

are studied in 4.2.2. and in 4.2.1 in different phytoplankton species. We will produce similar 

results on relationships between C:N ratios of primary producers and CO2 concentrations in 

culture experiments as planned by Rost and Boersma and will compare the results. Furthermore 

our data will be used by project 5.2 to improve the model parameterisation of biological 

responses to OA.





lab work 

field work 




- Establishing the δ 15 N -DON measurements at the IOW month 6 

- Data from batch culture exp. with cyanobacteria or diatoms month 12 

- Field data set from spring and summer cruise to the Baltic Proper month 24 

- Data sets from chemostat experiments month 30 

- Evaluation of the regulation of DOM release under high pCO2 

month 36 

Subproject 1.2.3 DOM availability and phosphorus utilization 

M. Nausch and G. Nausch (IOW) 

To study the DOM (DOP) release by phytoplankton and the DOM effects on phosphor utilization 

by heterotrophic bacteria different approaches will be used. Continuous culture experiments and 

batch culture experiments under controlled pCO2 conditions with representative organisms of the 

spring and summer bloom (diatoms and cyanobacteria) will be conducted (cooperation with 

subprojects 1.1.1., 1.1.2, and 1.1.4). In our responsibility are measurements of DOP and its 

constituents (ATP, DNA, RNA, Phospholipids) during the course of the experiments. Two joint 

field experiments in spring and summer will show how the whole plankton community will affect 

rates and processes compared to the individual organisms in batch and chemostat cultures. 

Experiments for DOP utilization by heterotrophic bacteria will be done in batch cultures. Similar 

experiments in chemostats are envisaged in cooperation with A. Engel. Supernatants of algal 

cultures will be inoculated with bacteria (strain or mixed population) and changes in DOP and 

bacterial P will be followed and compared with the turnover of radioactive compounds. The 

response of the natural bacterial community to additionally supplied DOC compounds under 

variable pCO2 will be investigated. Changes in DOP and bacterial P will be followed. The results 

will be incorporated in the carbon cycle model of subproject 1.3. The taxonomic composition of 

bacteria in the experiments and the contribution of the different groups to the P uptake will be 

studied with molecular biological methods (cooperation with H.-P. Grossart, see subproject 

1.2.4). 

Our investigations on the effects of ocean acidification on the turnover of organic matter and the 

expected changed C:N:P of DOM are urgently needed to explain the hypothesis in subprojects 

4.1.1, 4.1.4, 4.2.1 and 4.2.2. 

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92 




lab work 

Field work 




- Establishing batch cultures with different CO2 levels month 3 

- Data from batch and chemostat cultures for DOM production month 12 

- Data for DOM effects on phosphorus utilization by bacteria month 24 

- Field data from spring cruise to the central Baltic Sea month 18 

- Field data from the summer cruise to the central Baltic Sea month 24 

- Evaluation of both chemostat experiments month 33 

Subproject 1.2.4 Microbial response to DOM release and aggregation 

H.-P. Grossart (IGB) 

We aim to address the following major questions to achieve a better understanding of the 

coupling between ocean acidification and biogeochemical processes mainly controlled by 

activities of heterotrophic bacteria: 

• Do changes in pCO2/pH and subsequent changes in particle quality affect bacterial 

colonization rate and do they select for specific phylogenetic groups? 

• Do increasing fractions of C-rich phytoplankton-derived matter affect microbial organic 

matter remineralisation? 

• Does nutrient availability control microbial degradation of the presumably C-rich 

phytoplankton-derived matter and, thus, control the efficiency of the biological pump? 

• Are changes in pH and/or substrate quality reflected by the microbial transcriptome, i.e. 

expression of key genes? 

For this purpose, batch and continuous cultures of diatoms and cyanobacteria in the lab as well as 

mesocosms in the Baltic Sea will be used at different pCO2 concentrations. The experiments will 

be jointly conducted with A. Engel, M. Voss, and M. and G. Nausch. We will perform 

experiments both at atmospheric pressure (see above) as well as in pressurized incubation 

chambers together with L. Thomsen and G. Gust (mimicking particle sinking) to get more 

realistic data on pCO2-induced changes of microbial community composition and related


processes as well as their potential impact on carbon cycling. In all of our studies, we will 

consistently distinguish between free-living and attached bacterial communities. Field 

investigations with natural communities will be done in spring during the bloom of diatoms and 

in summer during the bloom of cyanobacteria in the Baltic Sea. By simulating various pCO2 and 

nutrient concentrations subsequent changes in microbial community structure and function of 

specific key players will be studied using molecular (DGGE, RFLP, CARD- and MAR-FISH), 

traditional microbial and biogeochemical methods. By combining molecular and biogeochemical 

methods, such as Stable Isotope Probing (SIP) we will be able to better link physiological and 

community structure response of heterotrophic bacteria to ocean acidification. 

Besides the tight linkage between sub-projects of the present cluster this project is linked to the 

following projects: 3.4.2 knowledge on the microsensor technique will be exchanged, 4.1.4 direct 

effects of ocean acidification on bacteria will be simultaneously measured and knowledge on 

physiological effects will be exchanged, 4.2.1 changes in plankton C:N:P ratios and effects on 

bacterial populations will be studied together, for project 5.3 our data provide valuable 

information for ocean acidification impact assessment. 



lab work 

field work 





- Establishing SIP method month 6 

- Experimental data set on direct CO2/pH sensitivity (microbial processes) month 18 

- Field data for verification of lab data month 28 

- Data set on combined effects of CO2 and pressure month 33 

- Exchange and compilation of data (for modelling) month 33 

Subproject 1.2.5 Effect of changing calcareous/lithogenic ballast on aggregates in the 

benthic boundary layer 

L. Thomsen, G. Gust (Jacobs University) 

In collaboration with AWI, Phytoplankton (diatoms, coccoliths) and cyanobacteria will be 

cultivated in bioreactors (100 – 5000 l) under different pCO2, temperature, radiation and nutrient 

inputs and then transferred into BBL laboratories. Detrital aggregates will be mixed with fine 

sediments typical for continental margins and further formed on roller tables (differential settling) 

and in shear tanks (turbulent shear). Then the aggregates will be transferred into in benthic flow 

simulation chambers and flumes and exposed to different pCO2, temperature, flow and 

hydrostatic pressures (0.1 – 20 MPa). The bioavailabity (via HPLC) and mineral composition 

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94 


will be analysed and aggregates classified. One set of experiments will evaluate the importance 

of increased CO2 levels and temperature on calcification and ballast formation on carbon 

degradation within the BBL with special focus on sorptive preservation. Data will be discussed 

with other partners to evaluate the pathway/transportation of aggregates from pelagic to benthic 

environments. Phytodetrital aggregates formed under current CO2 levels are compared with those 

formed in a future ocean. Another set of experiments will evaluate the importance of increased 

pressure on carbon degradation to determine sensitive biogeochemical parameters at atmospheric 

and in-situ pressures to understand and model the fate of organic matter in a future ocean. A third 

test of experiments will be carried out during spring/summer 2010/2011 to take in situ samples 

and expose them to varying pCO2, temperature and hydrodynamic conditions. Background of this 

research, the results and discussion will be made available to outreach and training. 

Our investigations are linked to the Themes 3 and 5. We will support subproject 3.4.3 to further 

understand the importance of the BBL for “carbonate sediment dissolution”, and improve and 

better understand the model results of 5.2 "Evaluating and optimising parameterisations of 

pelagic calcium carbonate production in global biogeochemical ocean models". 




calibration 

Production of organisms 

CO 2 BBL and ballast 

CO 2 BBL and pressure 

Field campaign 




- Implementation of experimental facility month 6 

- Exp. data set on CO2 & temp. on BBL ballast formation month 18 

- Exp. data set on CO2 & pressure on BBL carbon degradation month 24 

- In-situ data set on CO2 & pressure on BBL ballast & carbon degradation month 30 


vi. Budget and Budget Justification 


1.2.1 

1.2.2 

1.2.3 

1.2.4 


1.2.5 

Subtotal 

Consumables 

1.2.1 

1.2.2 

1.2.3 

1.2.4 

1.2.5 

Subtotal 

Travel 

1.2.1 

1.2.2 

1.2.3 

1.2.4 

1.2.5 

Subtotal 

Investment 

1.2.1 

1.2.2 

1.2.3 

1.2.4 

1.2.5 

Subtotal 

Other costs 

1.2.5 

Total 




Personnel costs: 1 Scientist (PhD-candidate; diploma or master required, ): Set-up and 

sampling of chemostat experiments at the AWI, and participation in field studies. The Ph.D. 

students will examine effects of changes in pCO2 and pH on the release and decomposition of 

exopolymer substances, i.e. polysaccharides and amino acids, TEP and CSP, incl. measurements 

of bacterial decomposition activities using enzyme assays, bacterial biomass production and 

respiration. Analyses of the elemental composition of organic matter will be supported by a 

technician at the AWI. Funds for student helpers are to support the set-up and sampling of 

chemostat experiments, and to help with cruises preparations, routine lab work and maintenance 

of cultures. 

Consumables: Consumables are primarily requested for Nuclepore-, membrane- and GF/F 

filters, Cytoclear slides for the microscopy of TEP and CSP, chemical reagents, glasware, 

nutrient media, ion exchange columns, HPLC columns, isotopes ( 14 C-bicarbonate, 14 C Leucine, 

3 H Thymidine), fluorogenic substrate analogues, and for Macrosept tubes (10, 100, 1000 kDa) for 

ultrafiltration. 

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96 


Travel: Travel funds are for exchange visits between partners, and for national meetings and 1 

international conference per year. In 2009, funds include travel costs to Svalbard (mesocosm 

experiment). 

Investments: Investments are needed for a START-1500 MLS microwave, which enables high 

throughput sample pre-processing during the hydrolysis and drying steps of CHO and AAS 

analyses under defined and reproducible conditions. Investments are also required for O2-optodes 

to determine bacterial respiration during the CO2 perturbation experiments, and for a, Shimadzu 

TNM-1 expansion of TOC analyzer. 


Personnel costs: A PostDoc will work half time on the project to establish the stable isotope 

measurements in DON in the IOW where all technical prerequisites are given. This work can be 

accomplished by an experienced person within the first 6 months. Batch culture experiments 

under 3-5 constant pCO2 levels will be carried out on cooperation with subproject 1.2.3. Cruises 

to the Baltic Sea need to be prepared and equipped; furthermore the person will participate in the 

common chemostat experiments at the AWI and carry out the analysis as described above. We 

aim to find one PostDoc to work the other half time for the subproject 1.2.4. A student is 

supposed to help with the preparation of the cruises and routine lab analysis (nutrients, CO2) for 

app. 60 hours per week throughout the whole project time. 

Consumables: Consumables include isotope tracer substances ( 15 N2 gas, and Na-H 13 CO3 - ), 

bottles, septa and syringes for the nitrogen fixation and carbon uptake experiments. The isotope 

mass spectrometers need high purity compressed gases and for the combustion of filters a number 

of reagents. 

Travel: The travel funds are for exchange visits to the partner institutes in Bremerhaven, Berlin, 

and Kiel and at least one international conference. 


Personnel costs: A PhD student will be employed to perform the extensive and ambitious 

experiments in the lab at IOW and AWI as well as during planned cruises, to interpret the data 

and to summarizes them in PhD thesis. It is aimed to engage a students help throughout the whole 

project to conduct routine pCO2, DOC, DON and DOP measurement. These basic parameters like 

the stable CO2 conditions in the experiments have to be ensured for this subproject but also for 

the cooperating subproject 1.2.2. 

Consumables: Consumables include experimental equipment (microwave digestion tubes, 

incubation bottles, CO2-gas etc.) and chemicals for the determination of DOC, DOP and their 

components, radioactive phosphorus compounds, and CO2 standards and filters. 

Travel: Travel funds are for stays in the partner institutes (Bremerhaven, Berlin) for joint 

experiments, to meetings and workshops (Kiel), and for participation in international 

conferences. 


Personnel costs: A PostDoc will work half time on the project to establish the stable isotope 

probing method at IGB where all molecular prerequisites are given and to measure stable


isotopes at IOW (in collaboration with M. Voss). This work can only be accomplished by an 

experienced person within the first 6 months. Batch culture and pressure chamber experiments (in 

cooperation with subproject 1.2.5) under 3-5 constant pCO2 levels will be carried out. Cruises to 

the Baltic Sea need to be prepared and equipped and the person will be responsible for all 

molecular work in the common chemostat experiments at the AWI. We aim to find one PostDoc 

to work the other half time for the subproject 1.2.2. A student is supposed to help with the 

preparation of the cruises and routine lab analysis (PCR, DGGE) mainly throughout the 

chemostat experiment and the cruises. 

Consumables: Consumables include molecular supply (such as Primers, Taq-polymerase, DNA 

extraction and purification kits, acrylamide gels etc.) but also substrates labelled with either 

stable (SIP method) or radioactive isotopes (bacterial production), fluorogenic substrates 

(enzymes), fluorochroms (SYBR Gold for DNA labelling and cell counts) and optodes for 

respiration measurements. Further it will include consumables for stable isotope analyses at IOW 

(see 1.2.2), reaction tubes, pipette tips etc. 

Travel: The travel funds are for exchange visits to the partner institutes in Bremerhaven, Berlin, 

and Kiel and at least one international conference. 


Personnel costs: One PhD student will create organic rich aggregates of different organic and 

inorganic compositions and carry out the flume experiments and pressure tank studies under the 

guidance of the supervisors and support of the OceanLab technician teams. The student will also 

participate in the field campaigns. Student helpers will support these activities and prepare 

specific web-modules (animations, class material) for outreach. 

Consumables: Consumables are primarily requested for Nuclepore-, membrane- and GF/F 

filters, chemical reagents, glassware, nutrients, HPLC columns, and chemicals for mineralogical 

analyses. 

Travel: Travel funds are for exchange visits with partners, annual meetings and one international 

conference. 

Investment: One investment during year 1 is needed for three identical benthic chambers with 

control electronics for the specific experiments under varying pCO2 levels, fully integratable into 

the pressure laboratory electronics system. 

Other costs: Other costs for 1.2.5 include the purchase of different, highly concentrated 

microalgae species from a commercial vendor (each year) and access (rent) to the mobile DL2 

pressure lab of TUHH (0.1 - 50 MPa) with full thermodynamic/hydrodynamic controls in year 2 

and year 3. The rental costs include transport, support by technician and access to specific 

sampling gear for in-situ studies during the field campaign. As Giselher Gust is retired from TU 

HH, these costs are not covered by the TU HH. 

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Project 1.3: Modelling biogeochemical feedbacks of the organic carbon pump 

B. Schneider (Universität Kiel) 

i. Objectives 

The aim of the project is to assess the impact of ocean acidification and global warming on the 

marine soft-tissue pump. In particular the cycling of dissolved organic carbon (DOC) will be 

investigated, as its production and decay is sensitive to changes in pH and/or temperature and it 

strongly interacts with particle flux dynamics. The processes involved are presently not very well 

understood, consequently implementations of DOC in global marine carbon cycle models can be 

considered as tentative. In collaboration with the experimental projects in Themes 1 and 3 this 

modeling sub-project will review current knowledge about DOC and transparent exopolymer 

particles (TEP) to be implemented in a global marine biogeochemical model (PISCES). 


The vertical transport of particulate and dissolved organic carbon (POC, DOC) in the ocean, the 

organic carbon (soft-tissue) pump, is largely responsible for the vertical gradient in the 

concentrations of dissolved inorganic carbon (DIC) in the water column (Volk and Hoffert, 

1985). Consequently, ocean carbon storage is more effective than it would be without this 

mechanism. A change in the organic carbon pump might result in alterations of the partitioning of 

carbon between the oceanic and atmospheric carbon reservoir. In current marine biogeochemical 

models the organic carbon pump is generally insensitive to changes in atmospheric pCO2. 

However, recent observations from mesocosm studies have shown a CO2-stimulated enhanced 

carbon drawdown by marine phytoplankton (Riebesell et al., 2007), which is partly transferred 

into DOC. Enhanced production of transparent exopolymer particles (TEP), a successor of DOC, 

has been documented from earlier mesocosm and laboratory studies (Engel et al., 2004; Engel et 

al., 2002). DOC plays a key role for the CO2-exchange between atmosphere and ocean as (1) its 

oceanic reservoir is much larger than that of POC, (2) it is sensitive to ocean circulation changes 

as its production depends on nutrient supply and export takes place via subduction, and (3) due to 

its sticky nature when transformed into TEP it is favoring aggregate formation and may thus 

affect particle flux dynamics to either enhanced (Engel et al., 2004) or reduced particle flux 

(Mari, 2008). A model-sensitivity study has already shown that a change by one unit in the C:N 

ratio of sinking particulate organic matter would have a considerable and persistent effect on the 

atmospheric pCO2 (Schneider et al., 2004), whereas global warming is generally expected to 

increase DOC degradation by microbial processes (Pomeroy and Wiebe, 2001). The combined 

effect of CO2 and temperature on DOC and the resulting alterations in DOC subduction, particle 

flux dynamics and air-sea CO2-exchange have not yet been studied. 

iii. Previous Work of the Proponent 

Birgit Schneider is the head of the Junior Research Group ‘Ocean Circulation and Paleo- 

Modeling’ in the framework of the Cluster of Excellence ‘The Future Ocean’ at the University of 

Kiel, Germany. The aim of this group is to address the interactions of climate and marine 

biogeochemical cycles, in particular with relevance to the global carbon cycle on time scales of 

present (Schneider et al., 2008; Schneider et al., 2003), past (Schneider and Schmittner, 2006) 

and future (Steinacher et al., in prep.). In collaboration with M. Gehlen and L. Bopp (LSCE, 

France) she has already performed model simulations estimating the effect of ocean acidification


on calcite forming phytoplankton (Gehlen et al., 2007) and aragonite forming zooplankton 

(Gangstø et al., 2008). Furthermore, she has worked on sensitivity studies estimating the 

relevance of changes in the vertical flux of particulate organic and inorganic carbon (POC, PIC) 

on the uptake and storage of CO2 by the ocean, which have demonstrated the potential for 

considerable and persistent effects on the atmospheric CO2 concentration (Schneider et al. 2004; 

Schneider et al., 2008b). 

iv. Detailed Description of the Project Work Plan 

In a first step (six months) an extensive review of what is currently known about the processes 

contributing to the production and decomposition of DOC and TEP will be summarized. 

Therefore, next to current literature the experience of the collaborators within Themes1 and 3 and 

their hypotheses for the new experimental setups will be explored, in particular the dependence 

of POC/DOC/TEP production and decay on pH and/or pCO2 (Schulz, 3.1.1; Engel, 1.2.1; 

Grossart, 1.2.4; Müller, 1.1.3; Nausch, 1.2.3, and Voß, 1.2.2, temperature (Müller, 1.1.3, Voß, 

1.2.2), light and nutrients (Hippler, 1.1.2; Nausch, 1.2.3, Voß, 1.2.2) as well as the role of DOC 

and TEP in the formation and fate of marine aggregates (Engel, 1.2.1, Thomsen, 1.2.5; Riebesell, 

3.2.1. Examples from paleo analogues will be considered in collaboration with two sub projects 

from Theme 3 (Mutterlose, 3.5.2; Meier, 3.5.3). In a second step (one year) the current 

implementation of DOC cycling in the marine biogeochemical model PISCES (Aumont and 

Bopp, 2006) will be revised. Therefore, together with participants from Themes 1 (see above), 3 

(Hoppema, 3.4.3) and 5 (Pätsch, 5.1; Oschlies, 5.2) and with colleagues at LSCE (France) new 

parameterizations for DOC cycling will be developed and implemented into PISCES. If 

available, these parameterizations should already include new experimental results obtained in 

Themes 1 and 3. The major points to be addressed here will be the effects of changes in the 

environmental conditions (pCO2, pH, temperature, light, nutrients) on (1) species specific DOC 

exudation rates of the (four) plankton functional types represented by the model, (2) DOC 

mineralization, and (3) the influence of DOC (TEP) on particle aggregation. For the latter a 

higher (lower) abundance of DOC and thus TEP will automatically increase (decrease) the 

frequency of collisions between DOC and particles and thereby favor (inhibit) aggregate 

formation, however, an increase in the stickiness of organic material with lower pH as observed 

by (Engel et al., 2004) is so far not considered by the model. In the following phase (1 year) the 

complex interplay of physical and biological responses to anthropogenic CO2 perturbations on 

the marine carbon cycle and potential climate feedbacks will be addressed by the application of a 

state-of-the-art coupled climate carbon cycle model (KCM - Kiel Climate Model, PISCES). By 

driving the biogeochemical model with pre-determined ocean circulation fields for either 

constant or changing climate conditions the effects of ocean acidification and global warming 

will be assessed separately and in combination. The final step (six months) will be the synthesis 

of new DOC and TEP related findings from Theme 1 that will also be relevant for the 

synthesizing projects in Theme 5 (Oschlies, 5.2; Quaas, 5.3). 

Work Schedule 

1.3 First Year Second Year Third Year 

review sensitivity of DOC and TEP to 

changes in environmental conditions 


99

100 



analyse implementation of DOC 

cycling in the model 

develop new parameterizations for 

DOC cycling also including TEP 

run CO2 scenarios 

syntesize DOC and TEP related results 

from Theme 1 

Milestones (1.3) 

- Review of the sensitivity DOC and TEP production and decay to changes in 

environmental conditions 

- Review of the sensitivity of POC, DOC and TEP turnover processes to hydrostatic 

pressure effects 

- Modelled climate feedbacks of the organic carbon pump under ocean acidification 

and climate change 

- Synthesis of new results on the sensitivity of DOC and TEP cycling to ocean 

acidification and climate change 



1.3 

Subtotal 

Consumables 

1.3 

Subtotal 

Travel 

1.3 

Subtotal 

Investments 

Subtotal 

Other costs 

1.3 

Total 

Budget justification 1.3 


month 6 

month 18 

month 30 

month 36 

Personnel costs: A PhD-student will work in Project 1.3 over the entire 3-year period of Bioacid 

(first phase), whereby the initial synthesis work provides optimal conditions to get familiar with 

the related science and to establish solid cooperations with the other project partners. 

Consumables: As consumables mainly data storage media like DVDs and external hard disks 

will be needed, e.g. one year of biogeochemical model (PISCES) output in monthly resolution on 

the ORCA2-grid has about 1.5 GB. Running several climate change scenarios over 250 years 

(preindustrial-2100) will afford disk space on the order of several terabytes.


Travel: Travel funds will be needed to visit national (Bioacid) and international collaborators (in 

particular L. Bopp, LSCE, France) and to attend at least one international conference per year. 

Other costs: Further costs will appear for a desktop PC including screen as well as for 

publication fees. All necessary model simulations can be performed on the NEC-SX8 supercomputer 

at the University of Kiel without additional costs. 

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11.3: Theme 2: Performance characters: reproduction, growth and 

behaviours in animal species 


Theme 2 will address the contribution of ocean acidification (OA) and hypercapnia (i.e. higher 

than normal CO2 concentrations in water, animal blood or tissue) to climate change effects on 

marine ecosystems through experimental studies in marine water breathing animals. Initial 

findings suggest decreased growth and enhanced mortality of sensitive species, e.g. among 

molluscs or echinoderms, in response to a doubling of CO2 levels from pre-industrial to 560 ppm 

(Shirayama and Thornton 2005). This value is likely exceeded during this century. Marine 

invertebrates are hypothesized to be among the organisms most sensitive to anthropogenic CO2 

accumulation, especially those heavily calcified and with a hypometabolic mode of life. 

Unravelling the physiological mechanisms responding to OA is required for the development of 

cause and effect relationships (Pörtner et al., 2004, 2005). However, ecosystem effects of OA 

will not be understood by solely analysing the direct influence of ocean physicochemistry on 

individual organisms and species. The CO2 dependent modulation of responses to other changing 

abiotic factors, in particular temperature, also have to be included. 

Previous analyses of CO2 effects have frequently addressed short-term stress responses or lethal 

thresholds. According to recent insight, however, performance decrements occur prior to the 

onset of stress and are responsible for climate-induced reductions in population density (Pörtner 

and Knust, 2007). Long-term performance and thus fitness is key to survival and success of a 

species. CO2 induced limitation may occur firstly through a decrease in the capacity for growth, 

reproductive output, or diverse behaviours including the ventilation of burrows or bioturbation 

activities or exercise capacities in general. Secondly, considering the known range of 

mechanisms affected by CO2, it appears that a shift of acid-base status, including a shift of 

extracellular pH, likely reduces functional capacity of affected mechanisms (including 

calcification) and the whole organism in due course (see overview in Fig. 2.1). Preliminary data 

indicate that a narrowing of thermal windows results and the effect observed suggests a large 

sensitivity of the width of thermal windows (Pörtner et al., 2005, Metzger et al., 2007) and 

associated temperature dependent zoogeographical ranges to CO2. Physiological key processes 

setting sensitivity to CO2 include the regulation of organismal and cellular acid-base and ion 

status and their feedback on other processes associated with organismal performance (Pörtner et 

al. 2004, 2005). The physiological principles shaping performance are likely also involved in 

multi-step processes affecting marine food webs. Here, species-specific responses and 

sensitivities cause various species of an ecosystem to be affected differently, resulting in changes 

in species interactions, food web structure and associated carbon fluxes. 

Physiological mechanisms at molecular to systemic levels are subject to long term modification 

during acclimation (within individuals) and adaptation (between generations). This will, at the 

whole organism level, cause shifting tolerance ranges and adjustments in performance optima, 

capacities and limits. Conversely, limits become effective where species and firstly, their critical 

life stages reach their limits of physiological plasticity and also, of acclimation capacity. 

Acclimation or evolutionary adaptation implies genetic changes that can be detected using 

genomic and transcriptomic tools. Preliminary experiments indicate in fact that long term 

acclimation occurs through enhanced gene expression processes of mechanisms involved in ion 

and acid-base regulation. However, it is presently unclear how and to what extent this type of 

acclimation improves performance. Similarly, evolutionary adaptation may improve resilience 

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over the time course of generations. As a proximate indicator of potential evolutionary 

adaptation, acclimation capacity to CO2 may reveal heritable variation between populations of the 

same species along a latitudinal cline. Differential acclimation may relate to temperature 

dependent differences in CO2 exposure due to different CO2 solubilities and associated water 

physicochemistry. 

metabolic equilibria 

protein synthesis rate 

Calcification site 

calcification - H + Ω 

- 

H + HCO - 

3 

i 

intracellular space 

3 Na + 

extracellular space 

Fig. 2.1: Hypothetical overview of processes and mechanisms potentially affected by CO2 in a generalized water 

breathing animal, at different stages in the life cycle, from egg to adult, emphasizing a key role for extracellular pH 

(top right) in defining sensitivity to ocean hypercapnia and acidification (after Pörtner et al. 2004, 2008). Some 

regulatory factors and processes relevant for growth and behavioural performance are indicated by shaded areas. As in 

thermal sensitivity, the first line of hypercapnia tolerance is likely set at the level of functional capacity of the intact 

organism and defined e.g. by its capacity for performance. e.g. growth (bottom left). Hypercapnia is expected to cause 

a reduction in performance and enhanced sensitivity to thermal extremes (conceptual model for impact of extracellular 

pH was adopted from Pörtner 2008; picture sources: www.valdosta.edu, www.imagequest3d.com, B. Niehoff, J. 

Pickering, www.fische.kanaren.org, O. Heilmayer, C. Bock & G. Lannig). 

H + 

gene 

expression ( + or - ) 

Low water pH and high CO 2 

and reduced HCO 3 - 

CO 2 

Cl - 

H + 

H 2O 

ATPase 

membrane 

ion equilibria - 

2 K + 

Na + 

HCO 3 - 

Na + 

- 

- 

H 2O 

H + e 

- 

blood 

pigment 

Na + /H + -exchange etc. 

Epithelia (gill, gut, kidney) 

Brain 

Chemosensory 

Neurons pHi ↓ 

Adenosine 

accumulation 

and release 

Heart 

functional 

capacity 

Tissues 

Muscle 

- 

ventilation rate (some groups) 

+ 

Operculum


Theme 2 will investigate various measures of performance as early indicators of sensitivity and 

also addresses their ecosystem consequences in various climate scenarios. Our goal is to 

elaborate the physiological mechanisms setting performance levels in various species and animal 

phyla as well as their capacity to shift those mechanisms and associated tolerance thresholds 

through acclimation or evolutionary adaptation at transcriptional and translational levels. 

Comparative work focuses on those species that are key in the functioning of the respective 

ecosystems or of commercial interest. Marine fish and invertebrates with likely different levels of 

sensitivity to CO2 and pH will be exposed to conditions simulating climate scenarios as expected 

from anthropogenic emission strategies (Caldeira and Wickett, 2003, 2005). Application of more 

extreme conditions may be required for an identification and characterization of key 

physiological mechanisms responding to CO2, from molecular to organismal levels of biological 

organisation. 

These efforts explore whether unifying physiological principles are operative across life stages 

and are shaping various degrees of sensitivity in both calcifiers and non-calcifiers. Potential 

methods and approaches include invasive (optodes and chromatography techniques) and noninvasive 

(NMR, optical techniques) studies of systemic and cellular acid-base regulation. They 

also include analyses of metabolic and calcification rates, of energetics and of correlates or 

functional rates of cellular processes (e.g. protein synthesis) associated with energy budget and 

growth. Investigations of gene expression capacities using microarrays include those genes 

shaping performance levels, in line with the concept of oxygen and capacity limited thermal 

tolerance and the assumed key role for acid-base regulation capacity in shaping sensitivity to 

CO2. CO2 effects on motor activity (e.g. grazing), oxygen consumption, circulatory and 

ventilatory activity, fertilization, fecundity or egg production and hatching, development and 

somatic growth will be studied. Temperature dependent acid-base and ion regulation will also be 

explored in model species, e.g. through studies of the levels of haemolymph or blood ion and gas 

variables (pH, PO2, PCO2, Mg 2+ , Ca 2+ , K + levels, total CO2, succinate). 

These efforts will include three projects, according to the functional role of marine fauna. Project 

1 will study grazers and filtrators, project 2 will focus on benthic reptant decapod crustaceans and 

project 3 will address aspects of ontogeny, allometry and mechanisms shaping performance and 

sensitivities in top predators. The latter project will include the development of mechanism-based 

modelling. 

Objectives 

• Identify critical stages in the life cycle (e.g. eggs, larvae) of functionally important marine 

organisms based on performance measures as indicators of sensitivity to OA 

• Analyse physiological mechanisms defining performance levels and sensitivity as well as 

potential consequences for behaviours 

• Estimate acclimation capacity (gene expression capacity) for that mechanism as the 

background of physiological plasticity 

• Compare species from various phyla and habitats with respect to evolutionary constraints 

in acclimation or adaptation as well as differential sensitivities 

• Quantify impact and tolerance thresholds (tipping points) 

• Identify potential consequences for shifts in species composition of ecosystems and 

associated changes in ecosystem functioning and food web structure 

• Assess interaction between effects of OA and ocean warming 

• Compare responses and mechanisms in different populations of a species (e.g. in a climate 

gradient) reflecting potential for evolutionary adaptation (genetic differences) 

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Building on previous experience (e.g. Pörtner et al. 2001, Bailey et al. 2003, Clemmesen et al. 

2003, Michaelidis et al. 2005, Caldarone et al. 2006, Metzger et al. 2007) collaborative research 

between projects (2.1., 2.2., and 2.3. as well as 3.1.3) will be carried out in populations of fish 

species like cod (Gadus morhua) or molluscs like pectinids (Aequipecten opercularis, Chlamys 

islandica) or mussels (Mytilus edulis) or crustaceans (Hyas aranaeus, Cancer pagurus) across a 

northern hemisphere latitudinal cline, between the North Sea and the Barents Sea. Species 

populations from various latitudes may display variable sensitivity to environmental hypercapnia 

in relation to CO2 solubility and carbonate saturation, to temperature dependent shifts in steady 

state acid-base status and associated decrements in e.g. growth performance (2.1.2, 2.1.3, 2.2.2). 

After adequate testing of mechanistic hypotheses in the adults, the respective knowledge will be 

transferred to juveniles or larvae (2.2.1; 2.3.1. and 2.3.2) for an investigation of the specific 

sensitivity of developmental processes and of the allometry of CO2 effects. 

Efforts will build on jointly used infrastructure developments (0.3.1. and 0.3.2.). Joint laboratory 

studies will use common CO2 incubations, with levels covering preindustrial values as well as 

those expected from IPCC emission scenario values. These values will be those commonly used 

throughout the network. They will be combined with temperature scenarios adopting realistic 

CO2 temperature combinations according to climate scenarios and local climates, adopting 

similar steps of e.g. 3°C including e.g. either 0,3,6,9,12,15 °C etc. 

Data from these joint experiments will provide a basis to understand the differences in 

sensitivities to OA among various marine organisms of major Metazoan phyla (Mollusca, 

Arthropoda, Echinodermata, Chordata (Pisces)). This allows testing of the hypothesis that the 

capacity of ion and acid-base regulation in relation to metabolic capacities defines the level of 

sensitivity in all life stages. Data will contribute to an understanding of tolerance thresholds 

(tipping points), of the relationships of changes in performance and in biocalcification (3.1.3., 

3.1.4.), and to the development of an integrative molecular to ecosystem picture of CO2 effects. 

Data will also improve the parameterization of mechanism based physiological, ecological and 

biogeochemical models. 

Project 2.1: Effects on grazers and filtrators 

PI: Thomas Brey 

i. Objectives 

Project 2.1 tackles two questions: 

- How will ocean acidification and temperature rise affect the invertebrate egg fertilisation 

process as well as organism fitness and performance in calcifiers and non-calcifiers, and 

- How will these effects translate into changes at the ecosystem level? 

Eggs that are directly released into seawater – as in our model organism sea urchin - are exposed 

unprotected to an acidified environment. Sea urchin eggs are dormant cells until fertilisation and 

are activated within seconds via an external messenger, the invading sperm cell. As a 

consequence, signal transduction pathways are induced in between seconds for the activation of 

metabolic processes needed for rapid synthesis and hardening of the fertilisation membrane to


avoid lethal polyspermy. Most processes of signal transduction involve ordered sequences of 

biochemical reactions inside the cell, which are carried out by enzymes, activated by second 

messengers. The second messenger calcium as a common regulator of many of these signal 

transduction pathways has been shown to have a key role in the regulation of enzyme activation 

and cell growth. Potential consequences of intracellular pH changes are depletion of intracellular 

calcium stores and destruction of the acid-base balance. Enzymes working in a narrow pH 

window are suspected to respond on intracellular pH changes by deactivation or inhibition. 

In isoosmotic animals an increase of seawater CO2 concentration will translate quickly into a 

corresponding CO2 concentration in the body fluids. This causes physiological responses at the 

cell and the organism level, as the organism intends to maintain pH-homeostasis. Within certain 

limits, higher costs of acid-base regulation may be compensated for by metabolic adjustments, 

but organism fitness and performance may be reduced seriously beyond those limits, particularly 

at elevated temperatures. Basically, OA represents the same problem for both non-calcifying and 

calcifying taxa. In the latter, however, the energetics of shell/skeleton formation may be 

particularly sensitive to OA, as may be the quality of the product. Therefore we target two 

ecologically important taxa here, calanoid copepods and bivalve molluscs, albeit, owing to 

differences in size and morphology, with different methodical approaches. In calanoid copepods 

the focus is pH-homeostasis, i.e. the organism level, with the aim to create a multi-species 

response matrix to OA and temperature rise that may facilitate our understanding of changes at 

the food web level. In molluscs, we explore responses at the cellular and the organism level 

simultaneously, with the intention to establish a physiological response model that may assist in 

explaining OA induced modifications of physical and morphological properties of carbonate 

shells/skeletons. 


Across species, the rapid formation of a fertilisation membrane after the acrosom reaction with 

one sperm is essential to avoid lethal polyspermy. The construction of this membrane occurs 

within milliseconds and is induced by the release of calcium (Ca 2+ ) from intracellular stores. The 

resulting Ca 2+ pools function as second messengers and induce a signal transduction cascade 

leading to the initial cell divisions for forming the embryo (Santella et al. 2006). Ocean 

acidification and high CO2 concentration are suspected to affect the function of membrane ion 

channels, particularly in sodium, calcium and potassium (compare Jonz and Barnes 2007, 

Doering and Mcrory 2007) and thus intracellular ion levels. In mammalian cells, Ca 2+ levels and 

signalling are strongly pH sensitive (Kostyuk et al. 2003, our unpublished results). Hence we 

presume drastic effects of OA on the Ca 2+ related signal transduction pathways during the 

sensitive phase of fertilisation and initiation of embryogenesis. To our knowledge, however, the 

impact of acidification on fertilisation success and embryonic development has not yet been 

studied. Sea urchins are particularly vulnerable to acidification (Miles et al. 2007), and sea urchin 

eggs have served as models for the analysis of fertilisation and development processes. However, 

how calcium signalling in oocytes of echinoderms is affected remains to be elucidated. 

Calanoid copepods contribute up to 80% to zooplankton biomass (e.g. Longhurst 1985) and are a 

key component in pelagic marine ecosystems, linking primary production to higher trophic levels 

(e.g. Runge 1988). Herbivorous species can control phytoplankton development (e.g. Bathmann 

et al. 1990); as consumers of phyto- and microzooplankton, they have a strong impact on the 

microbial loop in the upper water layers (e.g. Turner et al. 1998), and their faecal pellets 

contribute to the carbon flow from the surface to deeper water layers (e.g. review by Schnack- 

Schiel & Isla 2005). The role of CO2 for copepod ecophysiology will, however, not only be 

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understood through its direct influence, but even more so, through the modulation of responses to 

thermal extremes. A recent study shows that synergistic effects of carbon dioxide and 

temperature strongly affect the edible crab, Cancer pagurus, by narrowing its thermal tolerance 

window (Metzger et al. 2007). In copepods, to our knowledge, only two studies focussed on the 

effect of elevated CO2, indicating that hatching success and/or egg production of calanoid 

copepod decrease at concentrations >8,000 ppm CO2 (Acartia spp. Kurihara et al. 2004; Calanus 

finmarchicus, Mayor et al. 2007). Knowledge on the direct effects of CO2 on the organism 

physiology and on possible synergistic effects with increasing temperature is lacking completely. 

Elevated CO2 levels modify biogenic calcification. Most experiments demonstrated a reduction in 

calcification and size at elevated pCO2 (Bijma et al. 1998; Leclercq et al. 2000; Riebesell et al. 

2000; Michaelidis et al. 2005; Berge et al. 2006; Gazeau et al. 2007), but the opposite trend has 

been observed, too (Iglesias-Rodriguez et al. 2008). Shell formation consumes energy through 

active ion regulation but its integration into whole animal functioning and energy metabolism is 

little understood. OA and associated hypercapnia requires acid-base regulation as extra- and 

intracellular pH levels decrease (for review see Pörtner et al. 2005). Three main mechanisms are 

involved in this regulatory process: (i) metabolic production and consumption of protons, (ii) 

buffering of extra- and intracellular compartments e.g. by dissolution of CaCO3 exoskeletons of 

bivalves and other calcifiers, and (iii) active transport of equivalent ion across cell membranes. 

Chronic CO2 exposure has been shown to reduce shell growth rate and soft body growth in the 

bivalve, Mytilus galloprovincialis combined with suppressed aerobic energy metabolism 

(„metabolic depression“, Michaelidis et al. 2005), likely correlating with depressed protein 

anabolism as observed in isolated cells (Langenbuch & Pörtner 2003). The shell stabilises body 

form and function and protects the animal against predators and environmental forces and agents. 

Thus, understanding the temperature-dependent physiological mechanisms that restrict aerobic 

performance and thus fitness during hypercapnic conditions as well as its effect on shell 

properties are crucial to assess the degree of sensitivity of calcifiers to the future global scenario. 


2.1.1: 

Angela Köhler is cell biologist and pathologist with emphasis on toxicology and is involved in 

the development and implementation of biomarkers (OSPAR, AMAP, HELCOM, MEDPOL, 

Viarengo et al. 2007). She is experienced in enzyme and immunocytochemistry, light-, 

epifluorescence and confocal microscopy. Her group has 5 years experience in in vitro 

fertilisation experiments with sea urchins and in metabolic mapping by enzyme kinetic studies 

and immunolocalisation of proteins during fertilisation. 

Ulf Bickmeyer is a cell biologist experienced in optical (CCD-imaging, confocal laser scanning 

microscopy) and electrophysiological techniques (patch clamp and others) combined with cell 

and tissue culture (vertebrate and invertebrate cells). Expertise: Properties and modulation of ion 

channels, toxicity and cellular effects of secondary metabolites from marine organisms, 

intracellular signal pathways (cAMP and calcium as second messenger, Bickmeyer et al 2007), 

pH sensitive dyes. 

2.1.2 

Barbara Niehoff is an expert on zooplankton ecology and population dynamics. Since 1992, she 

combines laboratory experiments with field observations and, to obtain the large picture of the 

response of an organism to its environment, she conducts biochemical, histological and stable


isotope analyses (Kreibich et al. 2008). Currently, she is the head of the Helmholtz Young 

Investigator Group “Trophic interactions in pelagic ecosystems: the role of zooplankton”. 

Franz Josef Sartoris is an expert on the physiology of marine invertebrates, including acid-base 

regulation, ion- and osmoregulation, energy metabolism and respiration (Metzger et al 2007). He 

has broad experience with the implementation of physiological and biochemical methods in 

small-sized animals like amphipods, copepods and decapod larvae. 

2.1.3 

Gisela Lannig is an expert in temperature and stress physiology of marine vertebrates and 

invertebrates, in particular in organismic and cellular physiology where she has a broad 

methodological spectrum. In the framework of global change her work focuses on the combined 

effects of temperature and additional environmental stressors such as pollution or elevated pCO2 

on energy metabolism of marine ectotherms (Lannig et al. 2006). 

Thomas Brey is an expert for trophic relations and for energy budgeting at the organism, 

population and ecosystem level (Heilmayer et al. 2004). He is experienced in the 

sclerochronological analysis of biogenic carbonate archives, particularly mollusc shells (Epplé et 

al. 2006). 

Christian Bock is a biophysicist and expert in developing and applying Nuclear Magnetic 

Resonance (NMR) techniques, particularly for in vivo measurements of muscular and circulatory 

performance (Bock et al. 2008). His emphasis is the development and application of non-invasive 

in vivo techniques (NMR imaging and spectroscopy, optical techniques) in adaptive and 

comparative physiology of mainly marine animals. 

Olaf Heilmayer is an expert in experimental bivalve ecophysiology with emphasis on pectinid 

species. He is a marine biologist with a special interest in the ecology and physiology of marine 

shellfish thriving under conditions that set limits to life, namely in the deep-sea and polar context. 

His work on latitudinal adaptation mechanisms in pectinid bivalves was a key to understand 

biogeographic shifts in commercially important species (Heilmayer et al. 2004). 

Ragnhild Asmus is an expert in long term research on complex intertidal ecosystems, 

particularly in experimental work at various scales (Asmus & Asmus 2005). 


Subproject 2.1.1 Ocean Acidification and Reproduction: Is the beginning of Life in Danger? 

PI: Angela Köhler / Ulf Bickmeyer 


Sea urchin eggs are an excellent model to study early effects of environmental changes on vital 

biological processes during fertilisation and embryogenesis, mechanisms that are present across 

all eukaryotic species up to humans. We use Strongylocentrus sp and Psammechinus miliaris for 

analysing (i) calcium related signal transduction and enzyme activation during fertilisation 

processes, (ii) formation of a fertilisation membrane and (ii) embryogenesis at different CO2 

concentrations (280 to 1,900 ppm). Our fertilization studies will complement the studies 

proposed by Storch et al., Piatkowski, Melzner, Bleich et al., Tollrian as well as by Wahl 

(subprojects 2.2.1, 2.3.1, 3.1.3, 3.1.4, 3.2.2, 4.1.2) on embryonal development.. 

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- For exposure experiments gametes are harvested from female adults maintained in the 

aquarium. Oocytes will then be exposed to the suggested CO2/pH conditions and in vitro 

fertilisation experiments with appropriate replicates will be performed. 

- Calcium signalling: Oocyte calcium levels will be measured optically using fluorescent calcium 

chelators e.g. Fura 2 by means of CCD imaging and confocal microscopy (see Wertz et al. 2006, 

2007). L-type calcium channels seem to be present in oocytes (Tosti & Boni 2004). Calcium 

channels have been described to change function with changing pH; how small pH changes affect 

calcium channel function in echinoderm oocytes and calcium dependent second messenger 

cascades remains to be investigated in detail. Pharmacological tools like thapsigargin, 

nitrendipine and brominated pyrrole-imidazole alkaloids (Hassenklöver & Bickmeyer, 2006, 

Bickmeyer et al. 2007) are used to investigate calcium entry pathways. 

- In vivo imaging of enzyme kinetics during in vitro fertilisation of NADPH producing pathways 

and ovoperoxidase activation for metabolic at different CO2/pH and temperature regimes will be 

conducted to analyse the influence of pH on Km and Vmax values. (Use of non-destructive 

epifluorescence microscopy in a flow chamber for controlled gamete exposition). 

- Immunolocalisation. Many of the potential target proteins have been conserved throughout 

evolution. Therefore, a whole range of antibodies designed for mammalian studies can be applied 

in sea urchins, as we have tested already. Sequencing of the genome of Strongylocentrotus 

purpuratus is complete and supports immunocytochemical studies at the light, confocal and 

transmission electronmicroscopic levels which are highly informative with respect to amount and 

localisation of molecules at work and their changes in relation to environmental factors such as 

pH. 

Internal cooperation/networking with projects in other themes: 

Subproject 2.1.1 is strongly connected, predominantly to projects in theme 2 via methodological 

and scientific approaches (exchange of methodological expertise). Collaborative exchange with 

Melzner [3.1.3], Bleich [3.1.4] and Wahl [4.1.2] will address CO2 impact on performance at 

additional ontogenetic stages. Joint work is planned with Tollrian (3.2.2) in order to analyse CO2 

and pH effects on fertilisation processes and embryonal development across species. 

External cooperation/networking: 

Jacobs University: Prof Klaudia Brix, Prof. Andrea Koschinsky; Stanford University: D. Epel, 

A. Hamdoun; Brown University: G. Wessel; Bodega Marine Laboratory: G. Cherr. 

Schedule 


Collection and maintaining of organisms 

Set-up of culturing facility 

CO2 perturbation experiment, 

Measurements of calcium signals in living 

oocytes 

Metabolic mapping of enzymes during in 

vitro fertilisation 




Sample preparation for immunolocalisation 

(LM/TEM) 

Sample processing and microscopy 

(LM/TEM,AFM) 

Data analysis, statistical evaluation, data 

interpretation 





- Measurements of calcium signals in living oocytes 

and description of basic oocyte properties completed 

month 12 

- Metabolic mapping of NADPH producing enzymes during in vitro fertilisation 

1st set completed 

month 12 

- Sample preparation for immunolocalisation (LM/TEM) of NADPH producing 

enzymes/tyrosin crosslinking 1st set completed 

month 12 

- Measurements of calcium signals in living oocytes during CO2 changes month 24 

- Metabolic mapping of NADPH producing enzymes during in vitro 

fertilisation (CO2-T), 2nd set completed 

month 24 

- Cortical granule reaction to CO2, 2nd set completed month 24 

- Electron microscopy of protein localisation and cell pathology month 28 

- Measurements of calcium signals in living oocytes, 3rd set month 33 

- Data evaluation, manuscript preparation, presentation of results month 24-36 

Subproject 2.1.2 The response of zooplankton organisms to elevated CO2 concentrations 

PI: Barbara Niehoff / Franz-Josef Sartoris 


Our work will focus on three Calanus species, which dominate the zooplankton communities in 

Arctic Seas. C. finmarchicus is advected into the Arctic Ocean from the Norwegian Sea while C. 

glacialis and C. hyperboreus are endemic (Jaschnov 1970). All three species are adapted to the 

environmental conditions prevailing in the respective water masses (Conover and Huntley 1992). 

With increasing seawater temperature as climate models predict, the biogeographic boundaries of 

these key species will potentially shift (Hirche and Kosobokova 2007), and this will have 

consequences for the ecosystem functioning. 

Calanus finmarchicus is one of the copepod species studied best. Many aspects of its life cycle 

phenomena and their relation to environmental conditions are known (e.g. Marshall and Orr 

1955, Hirche 1998, Niehoff 2007), and this provides an ideal basis for studying the physiological 

response to CO2 in detail. In order to detect the species-specific responses to elevated CO2 and 

increasing temperature (links to 2.1.3, 2.2.1, 3.1.3, 3.1.4), we will also include C. glacialis and C. 

hyperboreus. All three species are large enough to perform measurements of acid-base 

physiology parameters. 

113

114 


Copepods will be collected in the field, either during sea going expeditions or at land based 

marine biological stations, e.g. in Kristineberg (Sweden) or Tromsø (Norway), and then 

transported to the aquarium facilities at AWI. In incubation experiments (link to 0.3.1), the 

animals will be exposed to different CO2 levels and temperatures according to values predicted 

from climate models as used within BioAcid. Algal food, i.e. diatoms and flagellates, will always 

be supplied in excess in order to avoid effects of food limitation. At start and during experiments, 

metabolic activity (citrate synthase, respiration rates measured by means of optical O2 

microelectrodes) as well as growth rates in terms of body weight (CN, protein content, dry 

weight) will be determined. In female copepods, egg production rates and hatching success will 

be studied to elucidate the combined effect of elevated CO2 concentrations and higher 

temperature on reproductive success. For studying the acid-base status, the extracellular pH in the 

copepods will be measured by optical pH microelectrodes suitable for small sample sizes (



- Aquarium facilities and algal food cultures established month 6 

- Copepods sampled for first incubation experiments month 6 

- Copepods sampled for second incubation experiments month 6 

- First experimental series and sample analyses (Calanus finmarchicus) completed month 12 

- Second experimental series and sample analyses completed (C. glacialis and C. month 24 

hyperboreus) 

- Data analysis and comparison of species completed month 30 

- Final BioAcid report month 36 

Subproject 2.1.3 Calcifying macroorganisms in acidifying & warming shallow waters 

PI: Gisela Lannig / Thomas Brey / Christian Bock / Olaf Heilmayer / Ragnhild Asmus 


- Analysing the physiological response is based on experiments using pectinid bivalves 

(Aequipecten opercularis, 38° - 62°N, and Chlamys islandica, 64° - 70°N) as a model organism. 

Pectinids grow comparatively fast and can swim, i.e. they can be exercised. Different populations 

will be sampled along the latitudinal/ temperature gradient. Animals will be exposed to present 

(control, 380 ppm) and future elevated CO2 levels (between 980 and 1,960 ppm) at ambient and 

predicted temperature regimes [link to 0.3.1]. After 4-6 weeks metabolic key processes will be 

determined in control and experimental animals: At the organism level (in vivo) we determine 

oxygen consumption and metabolomics during routine metabolism, after exhausted exercise and 

during recovery using flow-through respirometry in the NMR (Bailey et al. 2003), circulatory 

capacity (heart rate, hemolymph pO2) using Doppler perfusion system and implantable O2-micro 

sensors (Lannig et al. 2008), fitness parameters (contractile performance, lipid and glucose 

metabolism, growth and shell structure (see below) and acid-base & ion regulation (changes in 

pH and ion content in extrapallial fluid, hemolymph and tissues) using a force gauge/sensor 

combination and NMR techniques incl. MAS spectroscopy and chromatography, and 

pCO2/pO2/pH sensors (Bock et al. 2001, Brodte et al. 2007, Guderley et al. 2008) [link to 0.3.2, 

2.1.2, 2.2.1, 2.2.2, 2.3.2]. Direct incorporation of calcium and CO3 2- will be measured using 

labelling techniques (see Wheeler et al. 1975; Peck et al. 1996). At the cellular/organ level (in 

vitro) we determine temperature-dependent changes in energy allocation and estimate capacities 

for CaCO3 formation under internal milieu conditions inferred from organism level data and 

under changing CO2 and extracellular pH values [link to 2.3.2, 3.1.3, 3.1.4, 3.4.1, 3.5.1]. We will 

measure capacities and costs of key processes like ion regulation (Na + /K + -ATPase, Na + Proton 

exchanger, Na + -dependent HCO3 - /Cl - co-transporter), protein synthesis, RNA/DNA synthesis, 

ATP synthesis (mitochondrial efficiency) in gill, mantle and muscle tissue using CMOS cell 

chip-technology (cellular respiration and acidification), NMR/MAS spectroscopy and 

chromatography (see Wind et al. 2001, Lehmann et al. 2001, Mark et al. 2005, Bruno et al. 2006, 

Cherkasov et al. 2006). Capacities of carbonic anhydrase will be determined in gill and mantle 

tissue (Skakks & Henry 2002, Yu et al. 2006). 

115

116 


- The ecological effects will be evaluated by a “screening” approach that applies the whole 

organism approach to a wide range of taxa (molluscs, barnacles, sea urchins) from the German 

Bight. For this approach organisms will be maintained at defined temperatures and control and 

elevated CO2 concentrations (see above) for one whole seasonal cycle in mesocosms systems at 

the AWI field station Sylt. The selection of taxa will depend to a certain extent on the availability 

of the various candidate species in the Sylt/Rønne area that may vary greatly from year to year. 

Long-term effects of elevated CO2 levels on carbonate shell/skeleton properties are analyzed by 

standardized procedures at levels of macrostructure (mass, density, stability), microstructure (µm 

scale such nacre structure in bivalves, REM) and ultra structure (nm scale, crystallite structure, 

Atomic Force Microscopy) [link to 3.2.4, 3.3.1, 3.3.2, 3.4.1, 3.5.1]. Short-term experiments will 

be used to investigate synergistic effects of CO2 and temperature in early life stages (larvae, 

juveniles) of selected species [link to 2.2.1, 2.3.1, 4.1.2]. Organism fitness for all ontogenic 

stages is determined by standard measures such as metabolic activity, C, H, N, P content, and 

enzyme activities (see above). 

- The future scenario will combine our findings with existing evidence and with model 

predictions of the marine environment development during the next century. As an example, a 

predicted increase in wave action may be relevant for a species in which we find a certain 

reduction in shell strength owing to reduced CO3 2- levels. 

Internal cooperation/networking with projects in other themes: 

Subproject 2.1.3 is strongly interconnected, predominantly to projects in theme 2 via 

methodological approaches and incubation studies. Cooperation with projects in theme 3 will link 

our “physiological performance” data to “biomineralization success”: in collaboration with Bijma 

[3.3.1], Böttcher [3.4.1] and Immenhauser [3.5.1] we will determine CO2-dependent changes in 

shell dissolution rates and structure of our experimental animals. Collaborative exchange with 

Melzner [3.1.3], Bleich [3.1.4], Fitzke [3.2.4] and Wahl [4.1.2] will address CO2 impact on 

additional aspects of calcification and stress resistance. 

External cooperation/networking: 

University Bremen, DFG project in SSP 1158 “Sclerochronology & Recent Climate Change” (PI 

Heilmayer), EU project in EPOCA (PI Pörtner) 

Schedule 


Animal collection & culture 

Sample processing & measurement - 

organismic level 

Sample processing & measurement - 

cellular level 

Physiological data evaluation - 

organismic level 

Physiological data evaluation - cellular 

level 

Mesocosm long term experiments 




Mesocosm short term experiments & 

analysis 

Shell properties analysis & data 

evaluation 

Reporting & publication 

Theme workshop & conferences 


- Pectinid species culture established month 3 

- Pectinid exposure & mesocosm experiments started month 6 

- Data evaluation at organism level completed month 20 

- Shell property analysis completed month 28 

- Coupled cellular energy allocation & animal metabolic models month 30 

- Development of future scenarios month 33 

- Final BIOACID report month 36 



2.1.1 

2.1.2 

2.1.3 

Subtotal 

Consumables 

2.1.1 

2.1.2 

2.1.3 

Subtotal 

Travel 

2.1.1 

2.1.2 

2.1.3 

Subtotal 

Investments 

2.1.1 

2.1.2 

2.1.3 

Subtotal 

Other costs 

2.1.1 

2.1.2 

2.1.3 

Subtotal 

Total 


117


2.1.1 

118 


Personnel costs: 1 PhD position for 3 years 

Consumables: chemicals, antibodies, dyes, catching trawls 

Travel: €/y each for the PhD and the two PIs 

Investment: n/a 

Other costs: n/a 

2.1.2 

Personnel costs: 1 PhD position for 3 years 

Consumables: €/y for chemicals including ion concentration and enzyme activity 

determination, microelectrodes and aquarium equipment 

Travel: €/y for participation in expeditions, animal transport, BIOACID 

meetings and international conferences 

Investment: n/a 

Other costs: n/a 

2.1.3 

Personnel costs: 1 PhD position for 3 years – analysing the physiological response 

1 PhD position for 3 years – ecological effects & future scenario 

Consumables: chemicals incl. NMR consumables, sensors and metabolic chips SC1000, 

gas mixtures, aquarium equipment, sample preparation (REM, AFM) 

Travel: Animal transport ( €/transport), BioAcid meetings ( €/person), Int. 

Conferences (e.g. Symposium “The Ocean in a High CO2 World”, 

€/person), Transfer Sylt-Bremerhaven (ca. /trip) 

Investment: The MAS-NMR (Magic Angle Spinning NMR; €) spectra of intact 

tissue samples yield high-resolution “fingerprints” of component 

metabolites, while avoiding the risk of decomposition 

Other costs: Publication costs, e.g. American Journal of Physiology ( €)



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Project 2.2: Long-term physiological effects on different life stages of benthic 

crustaceans 

PI: Felix Mark 

i. Objectives 

This project will investigate CO2 sensitivity in different life stages using crustaceans as a model 

phylum to investigate common principles from the systemic to the molecular level. We intend to 

examine whether and how quickly various life stages can adapt to or tolerate increased CO2 

levels under exposition to carbon dioxide tensions from today’s up to the predicted extreme 

levels of OA. As populations within a latitudinal gradient display different thermal sensitivities, 

we will determine whether there is a synergy between CO2 and thermal challenges and what 

implication this has for the future spatial distribution of a species. To explore the synergistic 

effects of CO2 and temperature on the transcriptome, we will study whether and how relevant 

gene clusters and regulatory networks are differentially expressed within and between 

populations. We will use population genetics to include effects of genetic differentiation, and 

analyse whether populations of different scope for adaptability exist and whether the genetic 

variability of populations within a latitudinal gradient affect their physiological tolerance range. 

These molecular data will be combined with analyses of systemic ecophysiological parameters 

like behavioural and growth performance, acid-base regulation, oxygen delivery and 

consumption and calcification. Ultimately, this will enable us to develop an integrative view of 

how the necessity to regulate affects the animal´s energy budget. 

The above objectives will be investigated in two osmoconforming and calcifying reptant 

crustaceans (Anger 2001), that might be especially vulnerable to OA: Hyas araneus and Cancer 

pagurus. They are easily accessible, uncomplicated to rear and much is known about their 

biology (Anger and Jacobi 1985; Anger 2001; Truchot, 1980; Burnett and Bridges, 1981; 

Metzger et al 2007). They occur with great abundance over a wide latitudinal range (H. araneus: 

Barents Sea into the English Channel, C. pagurus: Norway to Northern Africa). The retreat in the 

Southern North Sea indicates that H. araneus is already affected by global warming whereas C. 

pagurus might benefit from the warming North Sea. This commercially exploited species is longlived 

and reaches sexual maturity at 10 years (Neal and Wilson, 2007). The juveniles spend most 

of their time near the intertidal with extreme daily fluctuations of CO2 tensions and temperature 

whereas the sexually mature adults inhabit deeper waters with less fluctuating abiotic factors. 


Ocean acidification and global warming present - in evolutionary terms - extreme challenges to 

environmental adaptation that summate onto the existing necessities of adaptation and ultimately 

require evolution of adaptations de novo. For some aquatic and terrestrial animals, an effect of 

global warming on the geographical distribution and even the risk of local extinction could be 

demonstrated (Parmesan and Yohe 2003, Pörtner & Knust, 2007). Based on seminal data on the 

decapod crustacean Maja squinado (Frederich and Pörtner, 2000), thermal tolerance windows of 

aquatic organisms have been defined by limits in oxygen supply through ventilation and 

circulation (Pörtner 2001, 2002). These limits exert their effects on the growth rate of individual 

specimens and the abundance of a population thereby shaping the biogeography of a species. 

Ocean acidification in combination with increasing temperatures thus even more adversely 

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affects species survival: upper thermal limits have been shown to decrease under hypercapnia in 

the decapod crustacean Cancer pagurus (Metzger et al 2007). 

It has been assumed that sensitivity to CO2 may be highest in the smaller life stages (Ishimatsu et 

al. 2004, 2005), such as eggs or the planktonic larvae, whereas thermal stress is felt especially by 

the largest individuals of a species (Pörtner & Knust, 2007), the benthic adult. Decapod 

crustaceans with complex life cycles develop through a planktonic larval and a benthic juvenile– 

adult phase. The larvae show dramatic growth and morphogenetic changes, which are affected by 

variation in environmental factors such as temperature (Anger, 1998) and salinity (Anger et al., 

1998). The Megalopa is the larval stage that might experience the greatest environmental 

fluctuations, as it is the transition stage from the pelagic to the benthic mode of life. Among the 

adults, brooding females might be especially affected by temperature extremes during cost 

intensive brooding care (Fernandez et al., 2000; Brante et al., 2003), the extent of which has an 

impact on the performance of the eggs and the hatching larvae. 

By influencing the acid-base status of seawater, CO2 severely affects ion and osmoregulation of 

higher marine animals and their life stages. This is of special importance in crustaceans, as they 

differ with respect to their ability to regulate the extracellular osmolality against the seawater, 

which significantly impacts species distribution: whereas osmoregulating species are able to enter 

coastal and estuarine environments, osmoconformer are usually limited to full seawater and more 

sensitive to varying salinity and are normally found in deeper water. The capability of 

osmoregulation can differ between early developmental life-history stages and adults in both 

directions (Charmantier 1998). Extracellular osmoregulation is associated with energy 

expenditure for active ion transport, which becomes apparent in higher Na + /K + ATPase capacities 

usually found in gills of osmoregulating species like Carcinus maenas, when acclimated to 

diluted seawater (Lucu & Flic, 1999; Henry et al., 2002). Thus the capacity of the sodium pump 

in branchial epithelia may be a suitable marker to determine limited acid-base regulatory function 

in certain species or life stages. In line with this assumption, the hypometabolic deep-sea decapod 

crustacean, Chionoecetes tanneri, displays higher sensitivity towards short-term, severe 

hypercapnia than the shallow-living Pacific Dungeness crab, Cancer magister, which is able to 

restore extracellular pH to normocapnic levels during 24 h hypercapnic challenge (Pane and 

Barry, 2007). Similar sensitivities may become especially effective in early life stages when 

development of the ion regulatory inventory may be incomplete. 

Classic examples of how osmolality, pH, CO2 and oxygen levels act in concert influencing 

central aspects of the energy metabolism, can be found in the respiratory pigment of the 

crustaceans, haemocyanin. As systemic oxygen supply and thus haemolymph oxygen levels 

strongly affect thermal tolerance windows (Frederich and Pörtner, 2000), respiratory protein 

function under hypercapnia and elevated temperatures is of central importance. CO2 can affect 

haemocyanin function by binding to the respiratory protein itself and by increasing the acid load 

of the haemolymph, both effects leading to decreased haemocyanin oxygen binding capacities 

(Bohr shift, cf. Mangum, 1980; Bridges, 2001). Decreasing levels of physically dissolved oxygen 

at warmer temperatures cause a further reduction in haemolymph oxygen tension. This can in 

part be compensated at the molecular level by expression and rearrangement of haemocyanin 

subunit composition. The expression of haemocyanin monomers in decapod crustaceans is thus 

very variable and influenced by ontogenetic stage (Durstewitz & Terwilliger, 1997, Terwilliger & 

Dumler, 2001) and many factors that exert physiological stress, such as pH, temperature, 

hypoxia. Recently, even microarray approaches have been developed to analyse the complex 

expression of a whole suite of haemocyanin functional units (Terwilliger et al. 2007). Other 

molecular mechanisms defining the limits of acclimation to environmental challenges have so far


been studied mainly by describing effects on single proteins. Genome wide transcriptome studies 

with their model-independent, inductive form of analysis contrast with the more conventional 

hypothesis-driven gene-by-gene or protein-by-protein form of analysis. Due to advantages in 

technologies, genomic studies become more and more attractive in non-model animals, where 

essentially no genetic information has been available and which were selected for special 

adaptation of their physiology to answer questions in an ecological, environmental or 

phylogenetic context (e.g. Fish: Gracey et al., 2001; 2004; frogs: Storey, 2004; turtles: Storey, 

2007). Recently, an EST library for the porcelain crab, Petrolisthes cinctipes, has been 

established, which serves as a basis for genomic studies in related crustaceans (Stillman et al. 

2006). An important advantage of these screening techniques is their ability to identify groups of 

proteins/enzymes that can have interrelated functions. Using profiling techniques based on 

categorisation of gene function, regulatory pathways and tissue-specific functions become visible 

and help to identify functions of so far unknown genes. Furthermore, novel (candidate) genes and 

pathways essential in the response to certain stressors can be detected (Gracey and Cossins, 2003; 

Cossins et al., 2006) and thus help to identify common molecular principles resulting in 

phenotypic plasticity of the whole organism. Since transcriptome studies cannot detect regulation 

at consecutive levels such as translation, protein turnover and post-translational modifications, 

the complex pattern of transcript responses has to be interpreted within the framework of known 

physiology to provide new hypotheses. Those have then to be tested for their functional 

significance with regard to common principles in evolutionary adaptation. 

During adaptation to their habitats in a climate gradient, marine animals encounter a wide variety 

of local conditions that over time lead to differentiation of populations within the distribution 

range of a species. Little is known whether genetic differentiation of populations is directly 

linked to the scope for physiological adaptability of a given species, but it may be assumed that 

population differentiation is key to fine tuning the match between genotype and environment 

along climate gradients. Adaptation to local environmental conditions on a molecular basis are 

typically identified by comparing expression patterns in populations experiencing known, 

different conditions. This indirect inference of the adaptive value of the observed differences in 

expression implicitly ignores other factors, most notably gene flow, that also structure the spatial 

distribution of alleles within a species. This argument can, however, be reversed in that the 

existence of different alleles and/or expression patterns among populations - despite high levels 

of genetic exchange - is a strong argument in favour of a stabilizing selection being responsible 

for maintaining the observed differences between them (Hemmer-Hansen et al 2008). Hence 

evidence, unbiased by colonization history, is needed, which corroborates the adaptive value of 

the candidate genes independent from interpretations derived from physiological pathways they 

are involved in. This can be obtained by contrasting the spatial distribution of alleles potentially 

involved in a response to OA with a baseline derived from selectively neutral microsatellite 

markers. 


2.2.1 

Daniela Storch, systemic ecophysiologist, has been working intensely on the thermal tolerance 

of crab larvae in a latitudinal gradient along the Chilean coast in the past years financed by the 

Alexander von Humboldt foundation (Storch et al., submitted a,b,c). As PI in a recently approved 

DFG funded project, she will investigate the thermal limits of early life history stages and their 

relevance for the biodiversity and biogeography of reptant decapod crustaceans. She is interested 

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in how behavioural and physiological performance of the different life history stages within the 

ontogeny of crustaceans may determine distribution limits. 

Christoph Held is head of a working group focussing on population genetics, phylogeny and 

phylogeography of marine crustaceans. By interpreting spatio-temporal patterns of molecular 

variation they investigate the processes driving diversification and speciation over ecological and 

evolutionary timescales (Held, 2000). They developed and successfully applied a novel approach 

for isolating microsatellite markers from unknown genomes (Leese & Held, 2008). They have 

also co-developed an integrated software environment aimed at facilitating high-throughput 

detection and classification of suitable microsatellites including multiplexed primer design (Held 

& Leese, 2007). 

Magnus Lucassen, molecular biologist, investigates the genetic basis of climate driven evolution 

in higher marine animals and aims to identify the underlying networks and signals involved. He 

focuses in his work on key processes of the central energy metabolism, in particular ion/pH 

regulation, aerobic/anaerobic capacities and oxygen carriers (Lucassen et al., 2006). Realtime 

PCR in combination with immunodetection of proteins and functional assays are being used to 

characterize key genes from fish, crustaceans and molluscs (Deigweiher et al., 2008). Currently, 

he investigates as PI in a DFG funded project the entire transcriptome in cold-adapted versus 

eurythermal fish in response to environmental challenges. These genomic approaches in different 

marine phyla are being introduced for the detection of unifying and specific regulatory networks 

essential for the adaptability to certain abiotic factors and their consequences for whole animal 

performance (Eckerle et al., 2008). 

Felix Mark, molecular physiologist, has a strong ecophysiological and biochemical background 

and has worked in various ecophysiological projects with fish, crustaceans and molluscs 

investigating energy metabolism at systemic (Mark et al., 2002), cellular (Mark et al., 2005) and 

molecular levels (Mark et al., 2006). As PI in a DFG funded project, he currently investigates 

haemocyanin adaptation in polar and temperate cephalopods and is specifically interested in how 

molecular structural design relates to physiological function (Melzner et al., 2007). 

2.2.2: 

The previous work of the group around Christopher Bridges has been extensively directed 

towards the study of ecophysiological adaptations in marine organisms in both the intertidal and 

sublittoral areas. Crustacean studies have led to the development of numerous techniques for 

catheterisation (Burnett and Bridges, 1981) and investigation of both oxygen and carbon dioxide 

transport (Bridges et al, 1979) in detail at the whole animal level. The properties of haemocyanin 

in terms of the structure and function of respiratory proteins and control of respiratory affinity 

have also been studied together with the influence of carbon dioxide (Bridges, 2001, Chausson et 

al, 2001). Ongoing studie involve the moulting of rock lobsters and extensive work on moulting 

hormones with colleagues in South Africa (Marco and Gade UCT).



H. araneus is a well-established model species at the AWI for examination of larval 

development. C. pagurus is a well-established model species in the Bridges lab in Düsseldorf 

with regard to acid-base regulation. The two species will be subject to the same BIOACID 

standard pCO2 levels of 380 (present, base line), 700 (2,5 * preindustrial, IPCC ‘business as 

usual’ for 2100), 1120 (4* preindustrial) and 1960ppm CO2 (7* preindustrial) and BIOACID 

standard temperatures in the range between 6 and 21°C. To ensure standardised experimental 

conditions, most long-term acclimations of both species will be carried out in the aquarium 

system in Bremerhaven. CO2 incubation of adult animals will take place in the AWI mesocosms, 

the hatching and rearing of larvae under CO2 in the recirculated AquaInno Pond-in-Pond systems 

[0.3.1: AWI infrastructure development plan]. Common parameters that will be measured in both 

subprojects 2.2.1 and 2.2.2 on H. araneus and C. pagurus include: 

- oxygen consumption (larval stages, juveniles and adults) 

- acid-base parameters (haemolymph pCO2, pH, total alkalinity, CCO2 and buffer titration) 

(juveniles and adults) 

- ion composition (larval stages, juveniles and adults) 

- haemocyanin oxygen affinity and subunit composition (larval stages, juveniles and adults) 

Subproject 2.2.1 Hyas araneus: Sensitivity, adaptive capacities and evolutionary 

consequences in populations from different latitudes 

PI: Daniela Storch / Christoph Held / Magnus Lucassen / Felix Mark 


The H. araneus model will look at the mechanisms from systemic (A) to molecular levels (B) on 

the background of population differentiation. 

(A) System physiology 

We will study the combined effect of acidification and temperature on the whole organism across 

populations including all life history stages from the brooding female and egg to the larval stages. 

Eggs are included since they may be affected in two ways: first, less oxygen supply by the 

females because of thermal stress and or higher costs for extra cellular acid-base regulation and, 

second, high CO2 sensitivity for the eggs. 

(1) Long-term accumulative effects of acidification 

Long-term exposure experiments of H. araneus females with and without eggs will be conducted 

to determine the costs of parental care at high temperature and different CO2 levels. Oxygen 

consumption will be measured to identify differences in the investment of brooding at varying 

CO2 levels at high temperature. Haemolymph flow of females and water flow through the embryo 

mass, as indicators for oxygen provision to the eggs, will be obtained by using magnetic 

resonance imaging techniques [2.1.3]. The loss of embryos, survival/development of the eggs and 

the hatching rates from egg masses of long-term CO2 incubated females will be detected. Larval 

performance of hatchlings from these differently incubated females will be monitored with regard 

to mortality rates during successive larval stages. Haemolymph ion composition in the various 

larval stages will be measured [2.1.2] and related to the Na + /K + ATPase function (see below). 

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126 


Additionally, oxygen-binding characteristics of haemolymph samples from the long time scale 

perturbation incubations of H. araneus and C. pagurus (sub-project 2.2.2) will be analysed 

spectrophotometrically at AWI and gas diffusion chambers (cf. work programme 2.2.2) in 

Düsseldorf. Purified haemocyanin samples of H. araneus and C. pagurus will be further analysed 

biochemically by 2D gel electrophoresis and native and SDS PAGE to examine putative changes 

in quaternary structure and expression of different isoforms (further characterised at the genetic 

level (B)). 

(2) Life history stages most vulnerable to acidification in synergy with temperature 

To meet this goal, larvae of populations along the latitudinal temperature gradient will be reared 

at varying CO2 levels and then acute temperature tolerance windows will be determined in the 

varying larval stages by measuring swimming performance, oxygen consumption rate and cardiac 

performance. 

(B) Molecular physiology and genetic basis of hypercapnic vulnerability 

(1) Functional characterisation of ion regulatory capacities 

To estimate ion regulatory sensitivities, Na + /K + ATPase functional capacities and mRNA 

expression will be compared in adults of different populations. Due to the small sample size 

Na + /K + ATPase abundance in larval stages will be assessed using antibodies, which have already 

been used in several crustaceans (Lucassen et al. unpublished). The data serves as baseline 

information as in several other projects [2.3.2; 3.1.3] and will be related to the acid-base 

parameters. 

(2) Isolation of key genes determining hypercapnic sensitivity/adaptability 

For analyses of the transcriptome we will generate a normalized cDNA library for one population 

under control conditions to ensure high genome coverage. Using suppression subtractive 

hybridization (SSH), further cDNA libraries will be constructed for the identification of 

differentially expressed genes within the time course of hypercapnic acclimation. This technique 

has been successfully used in our laboratory for copepods, cephalopods and fish. Additionally, 

the life stages that turn out to be most sensitive in the part (A), will be tested against the least 

sensitive stage. Sequencing of clones from the normalised and the subtractive libraries will be 

performed at the AWI, IFM-GEOMAR or service companies using pyrosequencing (normalized 

library) and conventional Sanger technologies (subtractive libraries), respectively. Genbank 

based assignment of protein function will allow identification of physiological processes 

involved in the response to hypercapnia. For unknown genes we expect several overlaps with 

other genomic studies [2.3.2; 3.1.3], so that at least the functional framework of the isolated gene 

can be attributed. 

Based on these approaches, oligonucleotide based micro-arrays will be developed for 

representative gene clusters that mirror physiologically important metabolic pathways and 

regulatory processes, with special emphasis to the ion regulatory inventory, aerobic/anaerobic 

metabolism and oxygen transport system. With this micro-array, the time course of the 

expression profile of all spotted genes will be determined under hypercapnia. Hybridisation, data 

collection and analyses will be performed at AWI (in coop. with Dr. Uwe John). For promising 

candidate genes differential expression will be validated using real-time PCR (ABI7500), and 

profiled in detail over the whole time series, for all acclimation conditions and in all populations


and life stages. We will thus be able to assess differences between populations (=genetic markers 

under selection) and to detect general fluctuations over time within populations. 

(3) Population genetics 

We will enrich and isolate candidate microsatellite loci from the genome of H. araneus using the 

reporter genome protocol (Nolte et al 2005; Held & Leese 2007). The high efficiency of the 

protocol allows careful selection of suitably variable loci for the question under study (Leese & 

Held, 2008). Contrasting presumably adaptive genetic differences between populations against 

selectively neutral microsatellites allows us to gather independent evidence for the adaptive 

nature of candidate genes identified using the functional and transcriptome approach. We will 

identify genetic evidence for natural selection by calculating pairwise differences for a matrix of 

n populations in the standardized F’ST for coding and neutral markers, respectively. Those point 

estimates for which the F’ST(coding) significantly exceeds the F’ST(neutral) are taken to be under natural 

selection. They will be tested to what degree environmental similarity (temperature) can explain 

genetic similarity - regardless of geographic proximity or background genetic similarity 

(Hemmer-Hansen et al 2007). These loci can then be considered prime candidate genes for indepth 

functional and physiological characterisation not only in crustaceans, but also in other 

animal phyla [fish: 2.3.2; molluscs: 3.1.3]. A nested clade analysis (Templeton 1998) will enable 

us to separate the impact of contemporary gene flow from the effects of events in the 

evolutionary history of H. araneus. The variability of microsatellites typically resolves more 

recent events (tens of thousand years), a period in which both the range shift in the German Bight 

took place presumably in response to ongoing climate change as well as a recolonization of 

higher latitudes after the last glaciation of the Northern hemisphere [4.1.2]. Thus, the combined 

approach of systemic to molecular approaches on the background of population genetics and 

history will allow for meaningful predictions of the future impacts of OA in this model species, 

which can be seminal for studies in other phyla. 

Schedule 


Animal collection, establishment of lab 

populations 

Incubation experiments, tissue/body 

fluid sampling, sample preparation 

Female performance with and without 

eggs (NMR) 

Larval performance (swimming 

activity, oxygen consumption cardiac 

parameters) 

Collaborative measurements of acidbase 

parameters, ion composition of 

body fluids, enzyme capacities, 

Western blots 

cDNA library construction/analysis, 

sequencing, gene identification, SSH 

libraries 

Gene expression analysis (micro array) 

and validation (real-time-PCR) 

Population genetics 


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results at conferences, synthesis 


- Implementation of incubation experiments, base-line data on acid-base + gas 

month 6 

transport status 

- Data set on female performance, base-line data on larval performance month 12 

- Data sets on activity and expression of ion regulatory proteins month 15 

- Data set on transcriptome month 21 

- Data set on synergistic effects of CO2 and temperature (first population) month 24 

- Data set on population structure month 27 

- Data set on larval performance (second population) month 29 

- Synthesis, evaluation of combined data sets, sensitivities and uncertainties month 35 

Subproject 2.2.2 Cancer pagurus: Chronic and acute responses – Adaptation versus 

Tolerance 

PI: Christopher Bridges 


In the C. pagurus model, three key physiological areas will be examined: 

1. Acid-base balance of the whole organism under varying carbon dioxide load (chronic and 

acute) together with varying temperature regimes 

2. Functionality of both oxygen and carbon dioxide transport at different life stages and the 

influence of both temperature and environmental carbon dioxide on this parameter. 

3. The influence of acidification, through higher carbon dioxide levels on both the hormonal 

control of moulting and at the same time calcium deposition within the carapace. 

Initially both juvenile and adult populations will be established together with suture tagging of 

individuals for experimental purposes and baseline parameters measured. These will involve 

acid-base status of the two populations (adult and juvenile), the gas transport properties of the 

haemolymph, their moulting stage, hormonal status and calcium incorporation rates. Acute (short 

time scale, 6 hrs) laboratory exposure to graded level(s) taken from the perturbation schemes will 

also be investigated if possible, probably using maximum exposure (1960ppm CO2; 7* 

preindustrial) only if time is limited. As out-lined above (work programme 2.2.1), long time scale 

perturbations experiments will then be commenced using the BIOACID standard levels of 380, 

700, 1120 and 1960ppm CO2. These will consist of parallel groups and also some specimens 

prepared for serial sampling from the base line groups. Sampling of both the haemolymph and 

acute exposure experiments for each perturbation level will be carried out at 6 and 12 month 

intervals. Acid-base status and gas transporting properties will then be examined using similar 

methodology as shown in subprojects 2.1.3 for bivalve molluscs and 3.1.3 for cephalopods. At 

the same time calcium incorporation rates will be studied for each of the groups. During 

perturbation incubation, growth and moulting will be regularly monitored by weighing and 

collection of exuvia.


A further set of synergy experiments will be carried out involving both carbon dioxide and 

temperature commencing 6 months after the start of the single parameter perturbations or 12 

months if space is limited. This will involve the use of standard carbon dioxide levels and 

standard temperature levels of 9, 15 and 21°C. Acid base status will be determined by standard 

measurements of haemolymph PCO2, pH, TA, CCO2 and buffer titration of the haemolymph for 

perturbated animals (Düsseldorf). CO2 transport properties can at the same time be established 

and oxygen transport properties of the haemolymph under various CO2 levels via the “diffusion 

chamber” (Bridges et al, 1979) technique (Düsseldorf). These analyses will also be applied to 

blood samples of H. araneus from parallel incubation experiments at AWI (cf. work programme 

2.2.1). The temperature sensitivity of oxygen transport can then be determined together with any 

specific effect of carbon dioxide on oxygen affinity. After the new base line determination from 

perturbated organisms these can be acutely exposed to the maximum levels of carbon dioxide 

perturbations (1960ppm CO2; 7* preindustrial) and again acid-base status and gas transport 

determined. Moulting hormone levels will be measured using standard HPLC methods and 

bioassays. For calcium incorporation, radioactive Ca 2+ incubations at set carbon dioxide levels 

will be measured and at the same time the role of carbonic anhydrase investigated using specific 

inhibitors to look at both the cellular and whole animal level (Düsseldorf) similar measurements 

will be made in project 2.1.3 for bivalve molluscs. 

Data from both matrices of carbon dioxide perturbation experiments and combined temperature 

carbon dioxide experiments with then be examined for synergistic effects. After completion of all 

sampling and analytical work, a possible scenario for C. pagurus acid-base balance, gas transport 

and calcium deposition in the face of long-term changes in environmental pH will be elaborated. 

The magnitude of the acute response before and after long-term perturbation will be assessed. 

The role of tolerance or adaptation in the physiological adaptation of C. pagurus, H. araneus and 

other crustacean species will then be considered for consequences on a larger scale. 

Schedule 


Establishment of juvenile and adult 

populations – Parallel and serial groups 

ID.-Tagging 

Collaborative analysis of base-line 

acid-base, gas transport and growth 

status 

Laboratory CO 2 Perturbations 

experiments 

Laboratory Combined CO 2 

Temperature Perturbations 

Sampling – Serial sampling + Parallel 

group 

Sample Analysis (Serial-) and acute 

test (parallel-group) 

Calcium incorporation experiments 

(Parallel group) 






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130 


- Establishment of populations and begin perturbation acclimation month 3 

- Baseline data on acid-base, gas transport and growth status month 6 

- Data set on CO2 perturbations experiments month 13 

- Data set on synergy of temperature and CO2 perturbations month 24 

- Completion of data evaluation and synthesis of model for crustaceans month 33 



2.2.1 

2.2.2 

Subtotal 

Consumables 

2.2.1 

2.2.2 

Subtotal 

Travel 

2.2.1 

2.2.2 

Subtotal 

Investments 

2.2.1 

2.2.2 

Subtotal 

Other costs 

Subtotal 

Total 


2.2.1 


Personnel costs: personnel cost stated here are for two PhD students over a period of three years 

to be paid at the official rate of . The success and feasibilty of this comprehensive 

subproject is dependent on the approval of the two PhD students as they will both be responsible 

for the time-consuming animal collection, perturbation experiments, rearing of the larval stages 

and maintenance of the adult populations aside from the experimental parts. One of the PhD 

projects will have a systemic, the other one a more molecular bias. Both PhD students will be 

supervised by all PIs involved, according to needs and expertise. This should lead to the 

presentation of two PhD theses in the form of published papers at the end of a period of 

approximately 3 years. 

Consumables: The annual consumables would involve the standard requirements for the 

laboratories: Animal collection and maintenance: ; Systemic physiology: optodes, gases, 

chemicals: molecular biology: library construction and sequencing: expression analyses:


Travel: Since some of the analyses will be based at the University of Düsseldorf, costs of trips to 

Düsseldorf would be included, also collecting trips for juveniles and adults (Helgoland, Millport, 

Norway (Bergen, Tromsö) and Spitsbergen). Support of travelling and subsistence is also needed 

for attending project meetings and workshops, for presenting project results on national and 

international conferences. 

Investment: None 

Other costs: None 

2.2.2 

Personnel costs: personnel cost stated here are for one PhD student over a period of three years 

to be paid at the official rate of . This person will be responsible for collecting and setting up of 

the juvenile and adult populations within the mesocosm system which will lead to the 

perturbation experiments over a period of six to 12 months. He/she will be responsible for 

sampling at the given six-month intervals and analysing both chronic and acute responses. This 

should lead to the presentation of a PhD thesis in the form of published papers at the end of a 

period of approximately 3 years. 

Consumables: the consumables outlined here are graded over a three-year period with a high 

during the second year. The consumables would involve the standard requirement for the 

laboratory e.g. gases, chemicals and isotopes, test kits, aquaria, electrodes, spare parts, animal 

supply and feeding 

Travel: Since most of the perturbation work will based in AWI (Bremerhaven), costs of trips to 

the centre would be included, also collecting trips for juveniles and adults (Millport, Roscoff, 

Helgoland). Support of travelling and subsistence is also needed for attending project meetings 

and workshops, for presenting project results on national and international conferences. 

Investment: Most of the necessary equipment is present and we have a well-equipped technical 

workshop, which can provide most of our needs. Specialized systems may have to be purchased 

for carbon dioxide measurement systems together with acid-base balance determinations and also 

an optode system. Again a graded use of these courses is proposed with a high investment in the 

first two years. 

Other costs: None 


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Templeton AR (1998) Nested clade analyses of phylogeographic data: testing hypotheses about gene flow and population history. Mol Ecol 

7(4):381-397 

Terwilliger NB, Dumler K (2001) Ontogeny of decapod crustacean hemocyanin: effects of temperature and nutrition. J Exp Biol 204:1013– 

1020 

Terwilliger NB, Ryan M, Phillips M R (2007) Crustacean hemocyanin gene family and microarray studies of expression change during ecophysiological 

stress. Integr Comp Biol 46: 991-999 

Truchot JP (1980) Lactate increases the oxygen affinity of crab hemocyanin. J Exp Zool 214:205-208 

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Project 2.3: Effects on top predators (fishes, cephalopods) 

PI: Catriona Clemmesen 

i. Objectives 

Early developmental stages of fish and cephalopods have been shown to be generally more 

susceptible to environmental toxicants than adults (McKim 1977, Ishimatsu et al. 2004) leading 

to the hypothesis that these are the life stages most sensitive to changes in oceanic pH and 

hypercapnia. At the same time, the allometry of thermal limitation shows that tolerance to 

temperature extremes is lowest in the largest specimens of a species (e.g. Pörtner and Knust 

2007). Exposure to combined temperature and CO2 scenarios are needed to show how OA and 

warming interact to shape fitness windows in the natural environment. This involves testing of 

the hypothesis that OA causes a narrowing of thermal windows (Pörtner et al. 2005, Metzger at 

al. 2007). 

Comparative analyses of the growth performance and biomineralization of calcified structures 

(otoliths, statoliths) of these two important groups (fish and cephalopods) will allow the 

comparison of effects of changing pH in the environment on processes involved in development, 

growth, metabolic reaction, energy budget allocations and calcification responses. These data will 

be compared to analyses of the capacity of ion and acid base regulation in order to evaluate their 

role in setting tolerance to the CO2 induced acidification of ambient media (Pörtner et al. 2005, 

2008). The mechanisms involved are specifically those setting and defending the parameters of 

acid-base status in extracellular and then intracellular compartment, which in turn strongly 

influence the levels of metabolic and functional performance of cells, tissues and the whole 

organism including growth. 

In addition to experimental studies a quantitative approach requires integrated mathematical 

modelling of the functioning of contributing acid-base exchangers as identified by experimental 

molecular and physiological analyses. The complexity of the model can be reduced to a 

manageable size by introducing parameters derived from experiments. At the same time the 

experimental design can be substantially simplified by the parallel development of a model, since 

the model defines exactly the parameters that are required for the comprehension of the 

processes. The project will concentrate on molecular and membrane biology functional studies, 

on patterns of extracellular and intracellular acid base regulation, on modelling the integrated 

functioning of the acid-bas exchangers as well as on acclimation and adaptation and will 

evaluate the reaction of the organisms with different performance indicators. 

For each species or life stage the conceptual model (Fig. 2.1) outlined above needs testing and 

complementation on various scales. Firstly, the mechanisms of acid-base regulation have to be 

identified. Long term incubations will reveal their immediate response and their capacity to 

acclimate under OA scenarios at various temperatures and depending on pre-adaptation to 

various climate regimes. Secondly, the relative contribution of these carriers to cellular, tissue 

and whole organism functioning and the level of energy turnover and performance needs to be 

explored under these scenarios. Thirdly, the response of the acid-base machinery to ocean 

physicochemistry requires mechanism based modelling as a basis for future quantitative 

scenarios of CO2 impacts at the whole organism level. As a perspective, process models may be 

used in a spatial ocean model, where past, present, and future scenarios are provided by model 

simulations of physical and biogeochemical states as reported by IPCC. This would allow 

examination of spatio-temporal scales for the tolerance of organisms to CO2 induced 

acidification.



Emerging knowledge indicates different sensitivities to ocean acidification in various marine 

invertebrate groups and marine fishes, with a key role for the capacity of ion and acid-base 

regulation in setting tolerance to the CO2 induced acidification of ambient media (Pörtner et al., 

2005, 2008, Fig. 2.1). The mechanisms involved are specifically those setting and defending the 

parameters of acid-base status, firstly in extracellular and then intracellular compartments, which 

in turn strongly influence the levels of metabolic and functional performance of cells, tissues and 

the whole organism during growth, reproductive periods or foraging (Fig. 2.1). The principles of 

how setpoints of acid-base regulation influence function and energetics at cellular, tissue or 

whole animal level, have been most comprehensively explored in the sipunculid, Sipunculus 

nudus, with a role for shifting use of individual acid-base exchangers (e.g. Pörtner et al. 2000). 

Data available for an Antarctic zoarcid indicate similar principles in operation in fish 

(Langenbuch and Pörtner 2003). However, our understanding of how individual transport 

proteins contribute to setting the levels of acid-base parameters and define their response to 

environmental change is still in its infancy. Similarly, detailed study is needed to quantify the 

feedback of acid-base status on cellular and tissue functioning as well as whole organism 

performance. 

Changes in whole organism performance are key to an understanding of stress effects at marine 

ecosystem level. A clear example is one where temperature extremes cause a loss in species 

abundance primarily through a weakening of performance at the limits of the window of oxygen 

and capacity limited thermal tolerance (Pörtner and Knust 2007). CO2 likely weakens the 

capacity of oxygen supply mechanisms further (Metzger et al. 2007). Sensitivity to CO2 may be 

thus highest at the limits of the thermal window of a species and, vice versa, sensitivity to 

temperature extremes may rise with increasing ambient CO2 levels (Pörtner et al. 2005, Metzger 

et al. 2007). 

During early life history the development of mechanisms involved in acid-base regulation may be 

a critical stage with respect to maintenance of growth and sensitivity to ocean hypercapnia. 

Larval growth is crucial for winter survival. Larval fish and cephalopod growth follow 

genetically determined patterns that are modified by environmental conditions including 

photoperiod, oxygen, temperature, prey availability and possibly CO2 induced changes in water 

physicochemistry. RNA concentration and RNA/DNA ratios in tissues of a wide variety of 

organisms have been related to growth rate and feeding condition. This approach appears to work 

particularly well for fish and cephalopod larvae that typically grow rapidly in weight mainly 

through high rates of protein synthesis during the larval period (Clemmesen 1994, 2003, 

Clemmesen & Doan 1996, Melzner et al 2005). Recent research findings indicate reduced 

translational activity (measured as a decrease in RNA/DNA ratio) in newly hatched herring 

larvae as a response to increased pCO2 during embryonic development (Franke, Clemmesen & 

Riebesell in prep.). 

Otoliths and statoliths are calciform ear stones of fishes and cephalopods, respectively. They are 

already available in early larval stages and involved in spatial orientation and movement, as well 

as in the perception of acceleration. They also function as life history recorders and under 

“normal environmental conditions” daily increments are added to their ring-like structure, with 

the amount of deposited material being dependent on growth and condition of the animals 

(Zumholz et al. 2006, 2007). Under stressful conditions like OA, deposition may decrease. 

Element incorporation into ear stones is mainly influenced by chemical parameters of the 

surrounding water, but it is unknown if and how changes in the pH in the marine environment 

will effect formation and daily incorporation of in these internal calcified structures. 

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136 


Inorganic carbonate species (CO2, HCO3 - ) in different body compartments mirror the response of 

acid-base physiology to changes in oceanic pH and hypercapnia. For instance, intracellular pH is 

controlled by the velocity of the interconversion of CO2 and HCO3 and the transport rates for 

carbonate species or protons across the compartment boundary (e.g. HCO3 - /Cl - antiport, CO2 

source via respiration, diffusive CO2 efflux). Mathematical modeling of the regulation of 

intracellular pH involving CO2 and ion exchange processes between body compartments and the 

environment rely on a sound understanding of carbonate chemistry in open buffered systems. 

Properties of the equilibrium state of the carbonate system in aqueous solution are well known. 

However, it can be shown that in cellular systems, where we have to deal with small 

compartments and short time scales the steady state of carbonate chemistry can deviate 

appreciably from equilibrium (Thoms et al. 2001, Thoms & Wolf-Gladrow in prep). Hence, for 

modeling intracellular pH regulation this chemical non-equilibrium is of particular interest. 

Hitherto there is little detailed work on the steady states of an open buffered carbonate system. 

The development of analytical and numerical techniques to determine the steady states of the 

carbonate system within body compartments is mandatory for the mathematical description of the 

acid-base status of the organism under different environmental conditions. 


2.3.1. 

Uwe Piatkowski is a zooplankton and fish ecologist with broad expertise in macrozooplankton 

community studies and in the biology of cephalopods focusing on reproductive and feeding 

ecology, on growth patterns, and on distribution and taxonomy. Recent studies investigated the 

influence of temperature and salinity on the trace element incorporation into statoliths of 

cephalopods and the reproductive adaptation of sepiolid cephalopods to an oceanic lifestyle 

(Zumholz et al. 2006, 2007a,b,c). He is a member of the Kiel University cluster “Future Ocean” 

and a partner in several EU projects such as EPOCA, the Marie Curie ITN network CalMarO, 

and the European network of excellence MarBEF and he has broad experience in 

interdisciplinary research. 

Catriona Clemmesen’s primary research interest has been studying environmental effects on the 

distribution and composition of fish communities, and the biology and importance of early life 

history stages of fish. The applicant has extensive knowledge and experience in using 

zooplankton and ichthyoplankton studies to interpret changes in the dynamics of marine 

ecosystems based on climate change and in biochemical and otolith analysis in relation to growth 

performance and animal well-being (Caldarone et al., 2006, Clemmesen, 1994, 1996, Clemmesen 

et al., 2003). The proponent is a member of the excellence Cluster “Future Ocean”, partner in the 

EU project EPOCA and the Marie Curie ITN network CalMARO and has initiated the 

installation of the CO2 manipulation system at IFM-GEOMAR. 

2.3.2. 

Magnus Lucassen is a molecular biologist with broad experience in the isolation and 

characterization of the responses of key genes in marine fishes, crustaceans and mollusks to 

temperature, CO2 and oxygen, using quantitative realtime PCR in combination with the 

immunodetection of proteins and functional assays. He investigate the genetic bases of climate 

driven evolution and responses to environmental challenges in marine animals, by focusing on 

key processes in energy metabolism as well as ion- and acid-base regulation, specifically on


aerobic/anaerobic pathways, oxygen carriers and transmembrane ion exchangers (Lucassen et al. 

2006; Eckerle et al. 2008; Deigweiher et al. 2008). Genomic approaches are currently being 

introduced for the detection of unifying and specific regulatory networks essential for the 

response to certain abiotic factors and their consequences for whole animal performance. 

Ulf Bickmeyer is a cell biologist experienced in optical (CCD-imaging, confocal laser scanning 

microscopy) and electrophysiological techniques (patch clamp and others) partly combined with 

cell and tissue culture (vertebrate and invertebrate cells). Interests: Properties and modulation of 

ion channels, especially calcium channels. Toxicity and cellular effects of secondary metabolites 

(natural products) from marine organisms. Intracellular signal pathways using cAMP and calcium 

as second messenger. pH sensitive dyes. Chemoreception in marine invertebrates (Bickmeyer et. 

al.2002; 2007; Wertz et al. 2006). 

Gerrit Lohmann is experienced in large scale modelling as well as the setting up of modelling 

tools including statistical analysis of observational and proxy data. His working Group 

‘Paleoclimate Dynamics’ at AWI has a long experience in simulating the different components of 

the Earth system in various climate stages. In addition, geostatistical methods are developed and 

applied for the analysis of environmental and climate data. Emphasis is on the ocean circulation 

and data-model comparison. Statistical analysis of paleo-environmental data and direct 

simulation of the data are essential for the integrated use of models and data. Models of different 

complexity are developed and used, ranging from conceptual, models of intermediate complexity 

to full circulation models (Lohmann, 2003; Knorr and Lohmann, 2003; Lohmann et al. 2008). 

Hans O. Pörtner has a long standing history in addressing the physiological bases of ecological 

processes and ecosystem functioning, particularly the roles of climatic factors like temperature, 

CO2 and oxygen in animal evolutionary history and ecology as well as biogeography. Together 

with his division he elaborated the concept of oxygen and capacity limited thermal tolerance 

across animal phyla, as well as its ecosystem implications. He is also an expert in quantitative 

studies of acid-base regulation. Current interests cover the interaction between climatic factors, 

the mechanisms shaping cellular and whole-animal energy budgets under various thermal and 

carbon-dioxide regimes, and the molecular mechanisms of environmental adaptation and 

limitation (Pörtner et al. 1998, 2000, 2005; Pörtner and Knust, 2007; Metzger et al., 2007; 

Pörtner, 2008). The proponent has coordinated the EU project CLICOFI and is currently a work 

package leader in the EU project EPOCA. 

Silke Thoms is a theoretical physicist with broad experience in interdisciplinary work and in 

modelling of biochemical processes involved in NADPH/ATP generation and carbon acquisition 

in (photoautotrophic) marine organisms. She has introduced the use of mathematical models to 

understand the role of spatial compartmentalization of chloroplasts for the carbon concentrating 

mechanism (CCM) in eukaryotic algal cells and to simulate phytoplankton growth under different 

light and atmospheric CO2 conditions. Currently, she is principle investigator of a DFG-project 

that has the aim to contribute to a better understanding of the regulation of elemental fluxes 

(carbon (C), nitrogen (N), divalent metal-ions (Ca 2+ , Mg 2+ , Sr 2+ ) on the level of individual (algal) 

cells. In the future, we are developing improved parametrizations of biochemical cellular 

processes as a platform from which a higher - level ecosystem simulation model can be 

constructed (Thoms et al 2001; Engel et al. 2004; Kroons and Thoms, 2006). 

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138 



Subproject 2.3.1: Effects of changes in ocean pH on the development, growth, metabolism 

and otolith/statolith formation and composition of fish and cephalopod early life stages: a 

comparative approach 

PI: Uwe Piatkowski / Catriona Clemmesen 


Perform laboratory and mesocosm experiments with marine fish (herring, cod and gobiids) and 

cephalopod (European cuttlefish, long-finned squid) eggs, larvae and juveniles at different pH 

levels in the range of expected changes from climate forecasting models (0.3.1). A CO2 

manipulation system has just been installed at IFM-GEOMAR and will allow for controlled and 

stable CO2 concentrations to be applied, which will enable to use the same pCO2 scenarios e.g. 

present (380), 2x pre-industrial (560), 2.5x pre-industrial, business as usual scenario for 2100 

(700), 3.5x pre-industrial (980) and 5x pre-industrial (1400) and temperature scenarios according 

to local climate. It is postulated that gill chloride cells are involved in pH regulation in marine 

species and exceptionally CO2 tolerant larval fish have been shown to have a high density of 

chloride cells (Ishimatsu et al., 2004). Therefore histological studies on the development and 

abundance of these gill cells in juvenile fish in relation to pH changes will be performed (in 

cooperation with EPOCA workpackage 6). 

Examine synergistic effects of changes in temperature, oxygen and pH on development, growth 

and survival of larval and juvenile fish and cephalopods, with the temperature tolerance being 

expected to decrease under lower pH levels. Detailed studies: 

• Measure fertilization rate, egg development (2.1.1, 4.1.2), changes in the morphology of 

fish/cephalopod eggs and embryos, size at hatch, size of the otolith/statolith at hatch and 

during larval development. 

• Determine RNA/DNA ratios as a biochemical indicator for protein metabolism in early fish 

and cephalopods (larvae and juveniles) and determine which stages are most susceptible to 

pH changes (3.1.4, 2.3.2) 

• Determine the effects of different pH levels on the formation and elemental composition of 

otoliths and statoliths in larval and juvenile fishes and cephalopods (3.3.1). 

• Study short- and long-term effects of low pH on larval fish and cephalopods to define the 

vulnerability (2.1.3, 2.3.2, 5.3). 

Elementary composition of statoliths and otoliths will be determined by using modern Laser 

Ablation techniques (LA-ICP-MS) available in IFM-GEOMAR Research division Marine 

Biogeochemisty in cooperation with Prof. Eisenhauer within the Marie Curie ITN Network 

CalMarO. 

Internal cooperation: 0.3.1, 0.3.2, 0.4, 2.1.1, 2.1.3, 2.3.2, 3.1.3, 3.1.4, 3.3.1, 4.1.2, 5.3

Schedule 




calibration 

Collection and rearing of organisms 

CO 2 perturbation experiment with fish 

& cephalopods 


Combined CO 2/temperature 

perturbation experiments with fish & 

cephalopods 









- Experimental data set on CO2/pH sensitivity on development, growth 

and metabolism of fish & cephalopods 

month 18 

- Statolith and otolith composition of cephalopods and fish month 24 

- Data set on synergistic effects of CO2 and temperature 

of fish and cephalopods 

month 30 

- Evaluation and comparison of combined data sets on 

fish & cephalopods 

month 33 

- Completion of work, submission of manuscripts month 36 

Subproject 2.3.2 Mechanisms setting and compensating for animal sensitivity to ocean 

acidification: functional capacities, thermal interactions and mechanism-based modelling 

PI: Magnus Lucassen / Ulf Bickmeyer / Gerrit Lohmann / Hans O. Pörtner / Silke Thoms 


The project will focus on teleost fish and their life stages from various climates, specifically 

Atlantic cod (Gadus morhua). Comparative data will be elaborated for an elasmobranch, e.g. 

Scyliorhinus canicula. For some principle questions and as a preliminary test of unifying 

principles in acid-base metabolism across phyla, data should be contrasted to those available for 

animal models characterized by incomplete acid-base compensation, calcifiers like Mytilus edulis 

or non-calcifiers like S. nudus. Accordingly, the project will coordinate with others with respect 

to such comparative analyses and the principle investigations of adult and juvenile life stages 

[0.3.1, 2.1.1, 2.1.3, 2.2.1, 2.2.2, 2.3.1, 3.1.3, Coordination with EPOCA, work package 6]. 

Experiments will use the BIOACID standard levels of 380, 700, 1120 and 1960ppm CO2 at 

variable temperatures considering the thermal windows of the species analysed. 

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140 


Central hypotheses driving the comparative approach: 

• Marine animals display various sensitivities to elevated CO2 levels according to the 

degrees of acid-base compensation in association with their phylogenetic constraints, 

metabolic capacities, life history stage, and the capacities of contributing transporters, 

as influenced by thermal acclimation and adaptation including the thermal influence on 

membrane structure. 

• Fish are less sensitive to ocean acidification than marine invertebrates due to a 

quantum leap in acid-base regulation capacity and efficiency. Early life stages are more 

sensitive than adults especially during the transition phase to adult physiologies. Adults 

may become more sensitive than juveniles at the edges of their thermal windows. 

• There is a climate dependent setting of thermal windows which has its bearing on the 

capacity and efficiency of acid-base regulation and thus, sensitivity to elevated CO2 

levels is higher at the edge of thermal windows; vice versa, thermal sensitivity is higher 

at elevated CO2 levels. 

A combination of molecular, physiological and modelling approaches will be applied in order to 

test these hypotheses under controlled conditions of water physicochemistry and temperature. 

Responses of larvae and adults will be titrated at various CO2 levels comprising those expected 

from IPCC emission scenarios. Detailed studies include: 

Studies of molecular and membrane biology coordinated with functional assays 

• identification of transporters according to sequence information and inhibitor effects: 

The presently about 150,000 ESTs published from G. morhua transcriptome projects 

will serve as a basis for identification and characterisation of transport proteins. 

Subtractive libraries (SSH) [2.2.1, 3.1.3] and determination of full-length sequences 

(RACE; see Mark et al. 2006, Deigweiher et al. 2008) will focus on gill transporters. 

• assessment of pH dependent gene expression capacity and protein concentrations of 

individual transporters using real-time PCR: The corresponding protein concentration 

will be monitored with available antibodies specifically developed for cod. Labelling 

the transporter protein with specific inhibitors will be another method of choice [2.2.1, 

3.1.3]. 

• analyses of the interaction of pH and temperature effects on gene expression: 

Transporters identified as essential for acclimation due to their mode of regulation will 

be further characterised through functional studies. After determination of the fulllength 

sequence, the gene will be cloned into an appropriate vector system for 

heterologous expression and functional assays applying optical and 

electrophysiological techniques [3.1.4]. 

• assay of membrane structures (changes in lipid composition) and functional 

consequences for ion and acid-base regulation depending on thermal acclimation and 

adaptation (climate regime) 

Functional studies, coordinated with molecular work 

• whole organism patterns of extra- and intracellular acid-base regulation at thermal 

optima as well as at low and high thermal extremes depending on acclimation (and 

adaptation) [0.3.2, 2.1.2] 

• influence of acclimation on the capacity of cellular pH regulation


• analysis of the capacity of cellular regulation mechanisms using pH sensitive 

intracellular dyes. Transporters will also be functionally characterised in isolated gill 

preparations (Deigweiher et al. unpubl.) using specific inhibitors. 

• pH dependent changes in the contribution of ion exchangers involved in transepithelial 

proton equivalent ion exchange, and in associated energy turnover in isolated perfused 

gills, following the rationale of Pörtner et al. 2000. 

• analysis of CO2 dependent performance indicators (e.g. protein synthesis, growth, 

larval development) [0.3.1, 2.1.1, 2.1.3, 2.2.1, 2.3.1, 3.1.3] and regulators (e.g. 

adenosine, neurotransmitters) at low and high temperatures and thermal optima, at 

various levels of acid-base parameters to define the vulnerability of important fish 

species [5.3]. 

Mechanism based modelling, building on functional studies 

Mechanism based modelling of the complex acid-base regulatory system and responding 

metabolic and functional characters will support predictions of set-points in acid-base regulation 

and their relevance in whole organism functioning. In thinking generally about the effects of 

temperature and CO2 on acid-base status, it is helpful to start by modeling a single compartment, 

which represents either particular cells or all cells collectively. We intend to investigate acid-base 

regulation in relation to aerobic / anaerobic metabolism and CO2/ion exchange between the 

organism and the environment. The model is described in terms of rate equations, being a system 

of coupled ordinary differential equations (ODEs). Based on this model we will use the metabolic 

control analysis (MCA) theory to examine the regulatory aspects of the acid-base status, 

including a shift of extracellular pH, and to infer the systemic control properties of CO2 and 

temperature variations on the level of the functional protein. 

• model of transport of inorganic ions (e.g. Na + , H + , Cl - , HCO3 - ) based on the transport 

kinetics of identified transporters and on analyses of energy turnover attributed to 

individual ion transport mechanisms as well as pH dependent changes in such energy 

turnover 

• model of acid-base regulation on short time scales, based on an open CO2 system and 

starting with a one compartment model comprising all known biochemical reactions 

(including buffering and net proton equivalent ion exchange) 

• simulation of acid-base regulation on long time scales: introduction of feedback 

phenomena caused by long term acclimation of the organism to environmental stresses 

in terms of adjusted model parameters reflecting the short term behaviour 

Internal cooperation: 

Projects 0.3.1, 0.3.2, 2.1.2, 2.1.3, 2.2.1, 2.2.2, 2.3.1, 3.1.3, 3.1.4, 5.3 

External cooperation: 

University Bremen, DFG project “Calcification” (Heilmayer) 

141

Schedule 

142 



Set up of aquarium and culture 

facilities 

Sampling of fish, larval cultures 

Incubation experiments, in vivo data, 

tissue sampling 

Isolation of transporter genes, library 

construction 

Tissue specific expression 

Analysis of gill ion exchanger 

activities in gill tissues and after 

heterologous expression 

Set-up of mechanistic model for 

integrative analysis of acid-base status 

Evaluation of physiological data and 

running of mechanism based model 

Reporting & publication 

Theme workshop & conferences 



- Aquarium facilities and animal cultures established month 6 

- Completion of first incubation experiments in vivo month 6 

- Set-up of tissue and cell preparations for functional studies month 9 

- Isolation of genes and proteins of relevant transporters month 9 

- First experimental series and sample analyses completed month 18 

- Set-up of integrated model of carbonate chemistry and ion transport month 24 

- Second series and sample analyses completed month 30 

- Data analysis and comparison of species completed, publication month 36 

- Simulation of acid-base regulation and sensitivity studies, publication month 36 

- Final BioAcid report month 36 



2.3.1 

2.3.2 

Subtotal 

Consumables 

2.3.1 

2.3.2 

Subtotal 


Travel 

2.3.1 

2.3.2 

Subtotal 

Services 

2.3.1 

2.3.2 

Subtotal 

Other costs, studentships 

2.3.1 

2.3.2 

Subtotal 

Total 


2.3.1 



Personnel costs: 1 PhD position for 2 years for the studies on fish, 

1 PhD position for 2 years for the studies on cephalopods. The missing two years for both PhD 

students will be supplied from the EU-Project EPOCA 

Consumables: €/year for chemicals (biochemical analysis), aquarium equipment… 

Travel: €/year for animal collection and transport, meetings and conferences… 

Services: €/year for the measurements of the elementary composition of statoliths and otoliths 

using Laser Ablation techniques 

Other costs: Student help: €/year for assistance in the rearing of larval fish and cephalopods, 

otolith/statolith preparation 

2.3.2 

Personnel costs: 3 x (3 PhD-students, total per year). Different skills and 

specializations are needed for each of the three fields of molecular and membrane physiology, of 

acid-base and functional studies up to organismal levels and of mechanism based modelling. 

Consumables: Chemicals, molecular biology (e.g. Sequencing: ,-€; expression studies: 

,- €; antibodies €, inhibitors etc., total: per year) 

Travel: field work, animal collection and transport; meetings etc. per year 

Investment: NA 

Other costs: NA 

143


144 


Caldarone EM, Clemmesen CM, Berdalet E, Miller TJ, Folkvord A, Holt GJ, Olivar MP and Suthers IM (2006) Intercalibration of four 

spectrofluorometric protocols for measuring RNA/DNA ratios in larval and juvenile fish. Limnol Oceanogr Methods 4: 153-163. 

Clemmesen C (1994) The effect of food availability, age or size on the RNA/DNA ratio of individually measured herring larvae: laboratory 

calibration. Mar Biol 118:377-382 

Clemmesen C and Doan T (1996) Does the otolith structure reflect the nutritional condition of a fish larva? - A comparison of otolith structure 

and biochemical index (RNA/DNA ratio) determined on cod larvae. Mar Ecol Prog Ser 138:33-39. 

Clemmesen C, Buehler V, Carvallo G, Case R, Evans G, Hauser L, Hutchinson WF, Kjesbu OS, Mempel H, Moksness E, Otteraa H, Paulsen 

H, Thorsen H and Svaasand T (2003):Variability in condition and growth of Atlantic cod larvae and juveniles reared in mesocosms: 

environmental and maternal effects. J Fish Biol 62:706-723. 

Deigweiher, K, Koschnick, N, Pörtner HO and Lucassen M (2008). Adaptation of ion regulatory capacities in gills of marine fish under 

hypercapnic acidosis. submitted. 

Franke A, Clemmesen C and Riebesell U. The effect of changes in ocean pH on the development and early larval phase of herring. Manuscript 

in prep. 

Ishimatsu A, Kikkawa T, Hayashi M, Lee K-S and Kita J (2004) Effects of CO2 on marine fish: larvae and adults. J Oceanogr 60, 731-741. 

Kroon, B. and Thoms, S. (2006). From electron to biomass: a mechanistic model to describe phytoplankton photosynthesis and steady state 

growth rates. J. Phycol. 42: 593-609. 

Langenbuch, M and Pörtner, HO (2003). Energy budget of hepatocytes from Antarctic fish (Pachycara brachycephalum and Lepidonotothen 

kempi) as a function of ambient CO2: pH-dependent limitations of cellular protein biosynthesis? J. exp. Biol. 206: 3895-3903. 

Larsen, BK, Pörtner HO and Jensen FB (1997) Extra- and intracellular acid-base balance and ionic regulation in cod (Gadus morhua) during 

combined and isolated exposures to hypercapnia and copper. Mar. Biol. 128: 337-346. 

Lucassen, M, Koschnick, N, Eckerle, LG and Pörtner, HO (2006). Mitochondrial mechanisms of cold adaptation in cod (Gadus morhua L.) 

populations from different climatic zones. J. Exp. Biol. 209: 2462-71. 

Mark FC, Lucassen M and Pörtner HO (2006) Thermal sensitivity of uncoupling proteins in polar and temperate fish Comp. Biochem. Physiol. 

D 1, 365–374. 

McKim JM (1977) Evaluation of tests with early life stages of fish for predicting long-term toxicity. J Fish Res Board Can 34:1148-1154 

Melzner F, Forsythe JW, Lee PG, Wood JB, Piatkowski U and Clemmesen C (2005) Estimating recent growth in the cuttlefish Sepia 

officinalis: Are nucleic acid based indicators for growth and condition the method of choice? J Exp Mar Biol Ecol 317:37-51. 

Metzger R, Sartoris FJ, Langenbuch M and Pörtner HO (2007) Influence of elevated CO2 concentrations on thermal tolerance of the edible 

crab Cancer pagurus. J. therm. Biol. 32: 144-151. 

Michaelidis, B, Ouzounis, C,Paleras, A and Pörtner HO (2005) Effects of long-term moderate hypercapnia on acid-base balance and growth 

rate in marine mussels (Mytilus galloprovincialis). Mar. Ecol. Progr. Ser. 293: 109-118. 

Michaelidis B, Spring ,A and Pörtner HO (2007). Effects of long-term acclimation to environmental hypercapnia on extracellular acid-base 

status and metabolic capacity in Mediterranean fish Sparus aurata. Mar. Biol. 150: 1417-1429. 

Pörtner, HO, Reipschläger, A, and Heisler, N (1998) Metabolism and acid-base regulation in Sipunculus nudus as a function of ambient carbon 

dioxide. J. exp. Biol. 201: 43-55. 

Pörtner, HO, Bock, C, and Reipschläger, A (2000) Modulation of the cost of pHi regulation during metabolic depression: a 31 P-NMR study in 

invertebrate (Sipunculus nudus) isolated muscle. J. Exp. Biol. 203: 2417-2428. 

Pörtner, HO, Langenbuch, M and Michaelidis, B (2005) Synergistic effects of temperature extremes, hypoxia, and increases in CO2 on marine 

animals: From Earth history to global change, J. Geophys. Res. 110: C09S10 

Pörtner, HO, and Knust R (2007) Climate change affects marine fishes through the oxygen limitation of thermal tolerance. Science 315: 95 - 

97. 

Pörtner HO (2008) Ecosystem effects of ocean acidification in times of ocean warming: a physiologist’s view. Mar. Ecol. Progr. Ser. in review 

Thoms, S, Pahlow, M, and Wolf-Gladrow, DA (2001). Model of the carbon concentrating mechanism in chloroplasts of eukaryotic algae. J. 

Theor. Biol. 208: 295-313. 

Thoms, S and Wolf-Gladrow, DA Relaxation time conctants of the carbonate system at steady states. Manuscript in prep. 

Zumholz K, Hansteen TH, Klügel A and Piatkowski U (2006) Food effects on statolith composition of the common cuttlefish (Sepia 

officinalis). Mar Biol 150:237-244 

Zumholz K, Hansteen TH, Piatkowski U and Croot PL (2007a) Influence of temperature and salinity on the trace element incorporation into 

statoliths of the common cuttlefish (Sepia officinalis). Mar Biol 151: 1321-1330 

Zumholz K, Hansteen, T, Hillion F, Horreard, F and Piatkowski, U (2007b) Elemental distribution in cephalopod statoliths: NanoSIMS 

provides new insights into nano-scale structure. Rev Fish Biol Fisheries 17: 487-491 

Zumholz K, Klügel, A, Hansteen T and Piatkowski U (2007c) Statolith microchemistry traces the environmental history of the boreoatlantic 

armhook squid Gonatus fabricii. Mar Ecol Prog Ser 333: 195-204


11.4: Theme 3: Calcification - sensitivities across phyla and ecosystems 


Marine calcification processes, especially those of dominant planktonic autotrophs (e.g. 

coccolithophores), have a considerable impact on the marine carbon cycle . While heterotrophic 

calcifiers are less important in terms of their contribution to the ocean carbon cycle, they play a 

crucial role in structuring ecosystems, mainly by creating complex three-dimensional habitats 

(‘ecosystem engineers’: reef building warm- and cold-water corals, bivalve beds). Others (e.g. 

sea urchins; pteropods) control ecosystems by means of grazing (e.g. on kelp forests or 

phytoplankton of high latitudes). 

The acidification of the ocean will have a variety of effects on its inhabitants and their metabolic 

performance, particularly on the calcifying organisms. Most of these effects will occur through 

the shifts in the carbonate system, where acidification leads to an increase of CO2 and a decrease 

in CO3 2- . 

CO2 + H2O ↔ H + + HCO3 - ↔ 2H + + CO3 2- 

Marine photosynthesis and organic matter mineralization are closely linked to calcification and 

CaCO3 dissolution: 

⇐dissolution ⇐ respiration 

Ca 2+ + 2HCO3 - ↔ CaCO3 + H2O + CO2 ↔ CaCO3 + O2 + biomass (CH2O) 

calcification ⇒ photosynthesis ⇒ 

In words, calcium carbonate precipitation buffers to some extend the pH increase due to 

photosynthetic CO2 fixation, and decalcification buffers the pH decrease due to respiratory CO2 

release. 

The marine calcification process is almost entirely biogenic and, in the absence of 

photosynthesis, releases CO2 to the atmosphere. Calcification can occur when the concentration 

product ([Ca 2+ ]x[CO3 2- ]) exceeds the saturation constant. The saturation constant decreases with 

temperature, i.e. warmer water is stronger oversaturated at the same calcium and carbonate 

concentrations. The state of seawater with respect to calcium carbonate saturation is often 

conveniently expressed as Ω, below 1 indicates under-saturation and above 1 over-saturation. 

Seawater of pH 8.2 is oversaturated for calcite, at 25 o C ca. 7 times (Ω=7), at 5 0 C ca. 3 times 

(Ω=3), under which condition no precipitation occurs. For precipitation a higher over-saturation 

is needed, either driven by shifts in the carbonate system or by active transport of protons and 

Ca 2+ by ion-pumps at the calcifying sites. Altered carbonate chemistry of future oceans will 

probably change calcification rates in most ecosystems. Calcification rates in many marine 

invertebrates correlate well with declining sea water Ω, while some are surprisingly tolerant of 

acidification induced changes in Ω (see Fig. 3.2, Ch 3.1). Conversely, calcium carbonate 

dissolution can only occur in under-saturated seawater, but the kinetics are even less well known 

and depend on surface area. 

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This theme is composed of projects studying calcification and decalcification mechanisms, 

signals in biogenic carbonates to obtain insights in formation, as well as evidence from past and 

recent events and the effects of pH stress on calcification both on ecosystem- and organism level. 

The projects on calcification mechanisms will study the pH-sensitivity of the transport of ions 

across membranes of marine calcifying organisms, and the sensitivity of the coupling between 

benthic microbial photosynthesis and calcification. By both mechanisms the over-saturation must 

be enhanced, but whereas benthic photosynthesis leads to a simple shift in carbonate equilibria, 

the biochemical mechanisms will include a more complete control by the organisms over the 

local chemistry of the calcifying site. The mechanism of calcification is studied from the biogenic 

carbonate side as well. Calcifying organisms maintain their own distinct trace metal homeostasis, 

which results in characteristic and species specific elemental ratios as well as in peculiar isotope 

fractionation (“vital effect”) which is significantly different from any inorganic-thermodynamic 

expectations. This “vital effect” is most likely due to the development of biochemical 

mechanisms to keep the trace metal homeostasis of the most important divalent cations, and other 

trace elements, e.g. strontium, in narrow limits in order to meet certain physiological needs. It is 

thus expected that trace metal concentrations in the biogenic carbonates will hold clues on its 

formation, and is effected by the oceanic pH. The effect of pH decrease on several highly 

sensitive calcifiers will be studied on the organism level. Since the calcium carbonate saturation 

constant is temperature dependant, a future sub-saturation with respect to aragonite will begin in 

the Southern Ocean and in sub-Arctic regions. Therefore, the local calcifying key-organisms, 

pteropods, may be most vulnerable for ocean acidification. Beside the vulnerability of subpolar 

regions also tropical coral reef ecosystems are globally threatened by a twofold effect of the 

increasing CO2 concentration in the atmosphere: Global warming as a major cause of coral 

bleaching and ocean acidification. Although it is generally assumed that corals are negatively 

affected at a certain level of pCO2, differential effects in corals and other calcifiers indicate that 

the picture may be much more complicated than previously thought. Of particular interest is how 

different ontogenetic stages respond to pH stress. 

The ongoing acidification appears not to be a unique event, but is documented for at least 4 times 

in the period between 52.5 till 58.4 Ma BC. These events were marked by global warmings, 4x 

current CO2 concentrations and a low seawater pH. The hypothesis will be tested that 

nannoplankton is less sensitive to acidification as bivalves, and compared to fossil records of 

these periods. Research directed towards decreasing the CO2 concentrations has focused on 

increasing marine or terrestrial net productivity (e.g. the oceanic iron fertilization experiment). 

However, also oceanic calcification is a potential atmospheric CO2 modifier: reduced 

calcification may at least partially compensate the increase of CO2, provided the decreased 

calcification does not lead to a decreased organic carbon burial. Decalcification leads to a pH 

increase and thus can buffer the effects of increasing CO2. This hypothesis will be tested by 

experiments with Antarctic- and North Sea sediments. 


Calcification and decalcification mechanisms 

3.1 Cellular mechanisms of calcification 

4 subprojects. PI: Melzner, co-PIs: Bleich, Form, Schulz 

Within this project we will study the physiological mechanisms of calcification in a variety of 

calcifying marine organisms: echinoderms, bivalves, cold-water corals, coccolithophores, ranging 

from temperate to arctic regions. The methodology includes molecular biological, biochemical


and cell physiological approaches, and the groups will strongly profit from each others expertise. 

The project is internally well integrated as explained in Ch 3.1.iv. The project will study 

calcification mechanisms from the organism side, i.e. ion transport to the calcifying sites. It is 

thus complementary to project 3.3, which focuses on mechanistic research on the solid phase. 

3.2 Calcification under pH-stress: Impacts on ecosystem and organismal levels 

4 subprojects. PI: Tollrian, co-PIs: Riebesell, Richter, Fietzke 

In this project we will study the effect of pH stress on organismal level. Investigated will be reef 

survival under pH stress (on reef and colony level), pH effects on coral recruitment, calcification 

of pteropods during critical development stages, and calcification in red coralline algae. A wide 

range of methods will be used, ranging from organisms physiological techniques, imaging of 

individuals and colonies, and solid phase chemistry and physics. A strong synergy will be 

obtained from collaboration with project 3.4 and 3.3, e.g. by comparing studies on solid 

chemistry, metabolic rate and microenvironment assessments using microsensors. Organisms will 

be studied from temperate and tropical areas. 

3.3 Ultra-structural changes and trace element / isotope partitioning in calcifying organisms 

(foraminifera, corals) 

2 subprojects. PI: Bijma, co-PI: Eisenhauer 

This project will investigate the isotope composition and physical parameters of the shells of 

corals and foraminifera to obtain information on the mechanism of calcification. The 

methodology is based on a state of the art solid phase analyses. The organisms will be sampled 

from temperate and tropical regions. The solid phase analyses will strongly support project 3.4, 

and the studies towards the mechanisms clearly interlink with Project 3.1. However, different 

from project 3.1, mechanistic information is derived primarily from the biogenic carbonates. 

3.4 Microenvironmentally controlled (de-)calcification mechanisms 

3 subprojects. PI: Böttcher, co-PIs: de Beer, Hoppema 

Within this project we will study benthic calcium and carbonate cycling, controlled by the 

metabolic activity of microorganisms, and in how far this is influenced by water column 

chemistry. The benthic conversions are subjected to a transport resistance in the sediments and 

boundary layer, which has profound effects on the sediment and porewater chemistry. The 

methodologies are mainly geochemical, including solid phase physics and chemistry, porewater 

composition, and measurements of transport- and metabolic rates. A modeling approach will be 

implemented and applied in several of the subprojects. Collaborative microsensor experiments 

are planned with Melzner and Form (3.1), Fietske and Richter (3.2). Joint solid phase analyses 

are planned with Eisenhauer (3.3). The studies will be done on benthic systems from tropical, 

temperate and Arctic regions. 

3.5 Impact of present and past ocean acidification on metabolism, biomineralization and 

biodiversity of pelagic and neritic calcifiers 

3 subprojects. PI: Immenhauser, co-PIs: Mutterlose, Meier. 

This project aims to assess traces of past pH events in bivalve shells, foraminifera and 

coccolithophores. The fossil record from past hyperthermal and acidification events combined 

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with analyses on experimentally grown shells will give insight in how environmental parameters 

influence the shell chemistry and physics. This project includes solid phase chemistry and 

physics, thus a collaboration with the partners in project 3.3 is evident. The samples originate 

from temperate and tropical areas. 

Project 3.1: Cellular mechanisms of calcification 

(PI: Frank. Melzner) 

i. Objectives 

Research in four sub–projects will focus on elucidating the mechanisms and sensitivities of 

calcium (Ca 2+ ) and dissolved inorganic carbon (DIC) transfer to calcification sites. Emphasis will 

be placed on identifying the role ion transporters and channels play therein, both in autotroph 

(coccolithophores) and heterotroph (corals, molluscs, echinoderms) taxa. Comparing relatively 

sensitive with tolerant calcifiers will help identify the limiting components in biogenic 

calcification machinery. In addition, the differing DIC requirements of calcification vs. 

photosynthesis will be investigated in autotroph model organisms. 

Figure 3.1: An overview of how the proponents combine expertise and resources to characterize cellular calcification 

responses, from the gene regulatory level to cell and tissue function.



Fig. 3.2: The dependence of calcification on seawater 

calcium carbonate saturation (Ωarag) in marine 

invertebrates: Long-term coral reef data set recorded in 

the Biosphere 2 mesocosm (Langdon et al. 2000; 

Milliman 1993), the bivalve Mytilus edulis calcification 

(Gazeau et al. 2007), the cephalopod Sepia officinalis 

(Gutowska, Pörtner, Melzner, submitted). Control 

calcification rates were set at 100%. 

Altered carbonate chemistry of future oceans will probably change calcification rates in both, 

marine auto- and heterotroph taxa. Coral, echinoderm and bivalve calcification rates correlate 

extremely well with declining sea water calcium carbonate saturation state, Ω (Fig 3.2), while 

others (cephalopods) are surprisingly tolerant of acidification induced changes in Ω. It is not 

known yet as to why, in many species, Ω gives the best correlation with calcification 

performance, as the carbonate ion is not considered the biologically active form of dissolved 

inorganic carbon (DIC): Rather, CO2 can cross biological membranes, and protons, as well as 

bicarbonate ions (HCO3-), are chief substrates of ion–transport proteins important for cellular 

homeostasis and carbon acquisition (e.g. Na+/H+ exchangers, Na+ dependent Cl-/HCO3- 

exchangers etc). It is still a matter of debate, which transport proteins are involved in Ca2+ 

transport towards the calcification site and if the paracellular or transcellular route is used. 

Involvement of Ca2+ ATPases and calcium channels has been demonstrated at least in some 

groups (Allemand et al. 2004). However, studies that investigate the ion transporters involved in 

combination with the functional properties of epithelia and whole cells in detail are scarce. 

(1) Autotrophs: calcification sensitivity might be influenced by photosynthesis 

Unicellular coccolithophores are complicated model systems at the base of the marine food web, 

as both calcification and photosynthesis have a high demand for DIC. They have been shown to 

actively take up CO2 and HCO3 - , operating a so-called carbon concentrating mechanism (CCM) 

(Rost et al. 2003; Schulz et al. 2007). However, similar to heterotrophs, differing sensitivities 

towards elevated CO2 have been observed in this taxon: While cellular calcification in Emiliana 

huxleyi was found to linearly decrease with increasing CO2 levels (Riebesell et al. 2000), 

Coccolithus pelagicus was hardly affected by changes in the carbonate system, and Calcidicus 

leptoporus showed an optimum curve with maximum particulate inorganic carbon (PIC) 

production rates at present day CO2 (Langer et al. 2006). A species specific response in 

particulate organic carbon (POC) production with varying CO2 was also observed, urgently 

calling for a process based understanding of CO2 dependent carbon acquisition for both 

photosynthesis and calcification in coccolithophores (see sub project 3.1.1). 

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(2) Basal heterotrophs: coral models without photosynthetically active symbionts. 

Complex interactions between calcification and photosynthesis complicate the study of ion 

sources and supply for calcification in hermatypic warm–water corals (Allemand et al. 2004; 

Gattuso et al. 1999). During biomineralisation, corals have to supply calcium and inorganic 

carbon to the calcification site (to the extracellular calicoblastic fluid, ECF) but also need to 

eliminate protons resulting from CaCO3 production (McConnaughey & Whelan 1997). Transport 

of molecules across the calicoblastic membrane into the ECF could be achieved by two 

mechanisms: a paracellular pathway (driven by a chemical or electrochemical gradient: 

diffusional), or a transcellular pathway through the cell (active transport with specific membrane 

transporters such as carriers and pumps). While there is some evidence for carrier mediated (e.g. 

Ca 2+ /H + ATPase, L-type Ca 2+ channels) calcium transport across the calicoblastic epithelium in 

warm-water corals (Tambutte et al. 1996; Zoccola et al. 2004), the pathways for DIC are largely 

unknown. Inhibitor studies point to the involvement of anion exchangers (e.g. HCO3 - /Cl - 

exchangers) in these epithelia (Tambutte et al. 1996). Carbonic anhydrase (CA) is also a key 

component of the calcification machinery, inhibition of this enzyme resulted in an 80% reduction 

of calcification rate in tropical corals (Tambutte et al. 2007). Within calicoblastic cells, it may 

primarily mediate between metabolically produced CO2 and HCO3 - destined for the ECF. A 

significant part of the DIC incorporated by coral calcification probably stems from metabolically 

produced CO2, either produced by calicoblastic cells or by photosynthetically active symbiont 

cells(Furla et al. 2000). Cold-water corals, such as Lophelia pertusa, do not contain 

photosynthetic symbionts, making them ideal study objects to solely consider calcification related 

ion transport. Experiments with tropical reef building corals demonstrated that a lowering of the 

carbonate ion concentration significantly reduces coral calcification and growth (Langdon et al. 

2003; Marubini et al. 2003; Schneider & Erez 2006) (see Fig 3.2). 

In the middle of this century, many tropical coral reefs may well erode faster than they can 

rebuild. Cold-water corals are living in an environment (high latitude, cold and deep waters) with 

carbonate saturation already close to a critical carbonate saturation (i.e. Ω=1) below which 

CaCO3 will dissolve. Projections indicate that about 70% of the currently known Lophelia 

pertusa reef structures will be exposed to sub-saturating conditions by the end of the century 

(Guinotte et al. 2006). This illustrates the necessity to learn more about the processes that 

influence calcification in these organisms (see sub project 3.1.2). 

(3) Complex heterotrophs: calcification sensitivity gradients within a phylum 

Certain molluscs may react similarly sensitive towards acidified sea water, especially the class 

bivalvia (see Fig 3.2). Exposed to elevated seawater pCO2, the genus Mytilus displays (1) an 

inability to fully compensate hemolymph pH, (2) decreased somatic growth rates, (3) decreased 

rates of calcification, (4) an inability to control blood Ca 2+ concentration, which might be an 

indicator of internal shell dissolution at low hemolymph pH (Gazeau et al. 2007; Michaelidis et 

al. 2005). Quite the contrary, the cephalopod mollusc Sepia officinalis does not suffer from any 

of these physiological perturbations when exposed to comparable sea water pCO2 (Gutowska, 

Melzner, Pörtner et al., submitted, Melzner et al. unpublished). In fact, S. officinalis is the only 

marine invertebrate studied so far, that does not show a decrease in calcification rate at elevated 

pCO2 values predicted for the next 300 years (Gutowska, Pörtner, Melzner, submitted). A high 

ion–regulatory capacity in combination with a certain natural pre–adaptation to transiently high 

pCO2 values, as can be observed in cephalopods, might be the key to tolerance. Little is known 

about the exact ion transport mechanisms related to calcification in molluscs. A higher 

organismic complexity (with respect to coccolithophores and corals) led to the evolution of


specialized ion exchange organs, the gills. Several candidate genes / proteins with a relevance to 

calcification processes have already been identified in both, Mytilus spec. and S. officinalis gills 

(e.g. Na + /K + ATPase, Na + /HCO3 - cotransporters (NBC) and HCO3 - /Cl - exchangers, CAs, Ca 2+ 

ATPases, Ca 2+ channels (Medakovic 2000; Piermarini et al. 2007; Melzner, Lucassen, Gutowska, 

Hu unpublished), their expression patterns in response to high pCO2 have, however, not yet been 

considered. A recent study on CO2-induced acclimation processes relevant for acid-base 

regulation in fish gills demonstrated that several important gill ion transporter mRNAs were 

differentially expressed during a six week time-course experiment (e.g. Na + /K + ATPase, NBC, 

AE, NHE, Deigweiher et al., submitted). Elevated Na + /K + ATPase and NBC capacities at the end 

of the experiment may be reflected in elevated costs of ion and acid base regulation under 

elevated CO2. As several of the differentially regulated transporters also play a role in cellular 

DIC accretion, these results indicate how intricately calcification and general pH regulatory 

processes are intertwined, and suggests that their metabolic machinery should be studied 

simultaneously (see subproject 3.1.3). 

(4) Larval heterotrophs: ontogenetic calcification sensitivity gradients 

Similarly, sea urchin embryos and larvae are suited for studies that investigate both, calcification 

and pH homeostasis due to their use as model organisms for the past 100 years. There is a large 

amount of data available on developmental processes and gene regulatory networks in this taxon 

(Poustka et al. 2007). Still, functional data examining pH and ion regulation on a cellular level 

are limited. Sequencing of the genome of Strongylocentrotus purpuratus has now been 

completed (Sodergren et al. 2006) and renders molecular genetic approaches highly attractive to 

identify candidate genes for proteins involved in homeostatic processes for ion composition, 

volume regulation and pH. Larval stages of marine ectotherms have been shown to react much 

more sensitively towards ocean acidification than their respective adult stages (Ishimatsu et al. 

2005). This indicates that compensatory mechanisms which are recruited in adults are not 

functionally active in early development. In several sea urchin species, stark decreases of larval 

calcification rate with water pCO2 have been recorded (Fig 3.5, Kurihara & Shirayama 2004. 

Stumpp & Melzner, unpublished). Mechanisms leading to retarded growth are largely unknown, 

as are the major pathways for Ca 2+ and DIC transport in echinoderms. How these pathways 

function and how they are being modified in the course of ontogeny, thereby altering sensitivity 

towards high water pCO2, constitutes an important research challenge (see subproject 3.1.4). 

In summary, the ion transport pathways relevant for calcification are poorly understood in many 

marine phyla, indicating that basic research efforts have to be undertaken to (i) identify gene 

products that code for important ion transport proteins, (ii) monitor levels and / or activities of 

mRNAs and proteins in response to abiotic stress, (iii) study the function of cells and epithelia 

where these proteins are active, (iv) study fluxes of Ca 2+ and DIC from the sea water into 

organisms and cells and, finally (v) estimate the capacity of key model organisms to acclimate 

their ion transport systems to an altered carbon system (see Fig 3.1). 


3.1.1: Kai Schulz combines all the expertise necessary to successfully study the proposed 

hypothesis, e.g., coccolithophore culturing, carbonate system manipulations, mass spectrometry 

(Schulz et al. 2006; Schulz et al. 2007; Schulz et al. 2004). Furthermore, employed as a research 

scientist at IFM-GEOMAR in Kiel he has full access to unique infrastructure, essential for an 

effective experimental implementation (centralized CO2 aeration system, membrane-inlet-massspectrometer 

lab). 

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3.1.2: During the last three years Armin Form has established and improved a world-wide 

unique cultivation facility for high latitude marine organisms and has developed many new 

methods for laboratory deep-sea coral research (Form et al., in prep.). Based on these methods 

the proponent's primary research focus has been on measurements of calcification rates of coldwater 

corals (Form and Riebesell, in prep.) and arctic coralline red algae (Form and 

Büdenbender, in prep.) in relationship to elevated CO2-levels and temperatures. 

3.1.3: Frank Melzner and Magdalena Gutowska have considerable expertise in physiological 

studies on mollusc stress physiology and the effects of ocean acidification on marine ectothermic 

animals (Melzner et al. 2006a; Melzner et al. 2006b; Melzner et al. 2007a; Melzner et al. 2007b; 

Melzner et al. 2007c), Melzner et al. submitted, Gutowska, et al., submitted). Melzner leads the 

Junior Research Group ‘Ocean Acidification’ within the Kiel Excellence Cluster ‘Future Ocean’ 

and has access to state of the art culturing and CO2 manipulation facilities. Gutowska currently 

finishes her PhD thesis at the AWI and will thereafter move as a Postdoc to Markus Bleich’s lab. 

Magnus Lucassen is an expert in molecular physiology and functional genomics of marine fish 

and invertebrates (molluscs, crustaceans), with a strong background in isolation and quantitative 

expression analyses of key genes and their functional impact under different environmental 

challenges (Eckerle et al. 2008; Heise et al. 2006; Lucassen et al. 2006; Lucassen et al. 2003), 

Deigweiher et al. submitted; see also subproject 2.3.2). Hans-Otto Pörtner has a long standing 

history in studying acid-base regulation and proton equivalent ion exchange of marine ectotherms 

in relation to ambient water conditions, as well as the interaction of acid-base parameters with 

metabolic processes (e.g. Pörtner 2002; Pörtner et al. 2000; Pörtner et al. 1990; Pörtner et al. 

2005; Pörtner et al. 2004; Pörtner et al. 1993, see also subproject 2.3.2). Current interest 

comprises the interaction between CO2 levels and other climatic factors, and the mechanisms 

shaping cellular and whole animal energy budgets. 

3.1.4: Markus Bleich has a long standing expertise in experimental ion transport physiology 

(Bleich et al. 1990; Kosiek et al. 2007; Schroeder et al. 2000). In previous projects on 

mammalian cells as well as on marine organisms, electrophysiology of cellular transport 

processes has been characterized from the single ion channel to the systemic level (Bleich et al. 

1999; Hou et al. 2007). Equipment and technology for patch clamp analysis of cells and isolated 

cell membranes is in place. In addition, microfluorimetry is used for the measurement of 

intracellular ion concentrations (H + , Ca 2+ , Na + , Cl - ) and membrane voltage. From human and 

experimental pharmacology in close collaboration with pharmaceutical industry a collection of 

pharmacological tools is available to manipulate ion channels and transporters which are directly 

involved in transport or which are responsible for the generation of driving forces (Bleich & 

Greger 1997; Reuter et al. 2008). Bleich and co-workers have significantly contributed to 

genome research projects for the identification of disease related genes (Barth et al. 2005) and 

used molecular biological techniques to identify, clone and characterize ion channels in epithelial 

transport (Waldegger et al. 1999). Kerstin Suffrian, PhD student in Bleich’s working group, has 

a profound expertise in microfluorimetric experiments on nano- and micrometric marine 

organisms (Suffrian, Bleich et al., unpublished) and will help train the proposed PhD student. 

Frank Melzner and working group (see also 3.1.3) have established a sea urchin culture at IFM- 

GEOMAR and recently performed first experiments on embryonic and larval development of the 

sea urchin Strongylocentrotus droebachiensis under elevated pCO2 (Stumpp & Melzner, 

unpublished).



Collaborative research within project 3.1 

While the four subprojects will each focus on one important aspect of cellular calcification 

mechanisms, some collaborative experiments will be performed simultaneously by all PIs (1 st 

quarter of years two and three) to gather comparative biochemical indicators for calcification 

performance for all studied taxa. Recent findings indicate that carbonic anhydrase (CA) activity 

might be a meaningful indicator for ion-regulatory capacity in marine organisms and, possibly, 

sensitivity towards ocean acidification (Fabry et al. 2008, in press). CA is a crucial enzyme 

important both for general acid-base regulation, calcification and especially photosynthesis, in 

auto- and heterotrophic organisms (see e.g. Henry 1996). Experiments will be conducted using 

the state of the art membrane inlet mass spectrometry (MIMS) setup in the lab of Ulf Riebesell 

and Kai Schulz. At present, meaningful CA measurements can only be conducted using MIMS 

technology, as classic biochemical CA assays operate under un-physiological conditions (cold 

temperatures, low pH). CA capacity analysis will also be strongly related to studies of in vivo 

acid base physiology carried out in theme II. 

In addition, gene expression studies will be performed in the central molecular biology labs of 

IFM-GEOMAR and Magnus Lucassen’s lab at the AWI. Using transcripts that might be of 

importance for calcification processes in all organisms (CA, Na + /K + ATPase, Ca 2+ ATPase and 

Ca 2+ channels), we will try to answer the question, whether ocean acidification activates a 

common, compensatory gene response and how their regulation capacity defines organisms 

sensitivity. Which transcripts will be monitored following the first year will depend on the results 

of the transcriptomic work conducted in subproject 3.1.3. Gene expression will be studied using 

real-time PCR techniques. Instrumentation is available at both AWI and IFM-GEOMAR, 

technical support can be provided by Frank Melzner’s working group (e.g. lab technician). 

Eventually, we will correlate calcification performance of the various taxa with the 

genomic/enzymatic indicators, to develop a conceptual framework of how sensitivity to future 

acidification might be related to the phenotypic flexibility of the metabolic machinery. Output of 

this collaborative effort will be a synthesis / review type article, co-authored by all PIs. 

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Subproject 3.1.1: Inorganic carbon acquisition for calcification and photosynthesis in 

marine coccolithophores: towards a unifying theory 

(Kai Schulz, IFM-GEOMAR Kiel) 


Within the framework of the project 3.1 'Cellular Mechanisms of calcification', the apparently 

different modes of carbon acquisition for particulate inorganic and organic carbon production 

(PIC/POC) in marine key coccolithophores, 

such as Emiliania huxleyi, Coccolithus 

pelagicus and Calcidiscus leptoporus will be 

investigated. The experiments planned will be 

based on the hypothesis that the seemingly 

different calcification and photosynthesis 

dependence on CO2 in various 

coccolithophores are the result of species 

specific thresholds for DIC demand and pH 

sensitivity. Therefore we hypothesize all 

species to possess certain CO2 and pH optima 

for both processes. On one side of the optimum 

curves photosynthesis and calcification would 

Fig. 3.3: Schematic diagram of the proposed effects 

of seawater and pH on photosynthesis and 

calcification of coccolithophores. The process 

sensitivity observed for a certain coccolithophore 

crucially depends on the experimental CO 2 / pH 

range and can even change its sign. 

be limited by the availability of DIC. While on 

the other, low pH values would hinder their 

maximum functioning (Fig. 3.3). Hence, 

coccolithophores will be cultured under a broad 

range of well defined carbonate chemistry 

conditions, extending the core CO2 levels between 280 and 980 ppm towards both ends. This 

allows assessment of individual CO2 sensitivities of photosynthesis and calcification. 

Furthermore, employing sophisticated carbonate chemistry manipulations, it will be possible to 

separate the usually tight coupling of CO2, 

respective DIC concentration, and pH. Standard 

measurement techniques (POC, PIC, inorganic 

nutrient, growth rate determinations) will be 

combined with membrane-inlet-mass 

spectrometry (MIMS), allowing for carbon flux 

measurements on the cellular level (Fig. 3.4). 

Moreover, the MIMS setup will be used to 

quantify cellular carbonic anhydrase (CA) 

activity, a key component of many CCMs. The 

MIMS setup will also be available for other 

project partners interested in CCM activity 

measurements such as subproject 1.1.3, 1.1.4, 

1.2.5, 3.4.2. Frequent experience exchange and 

discussions with project 4.2.2 will ensure high 

data quality. Collaboration with subproject 3.5.3 

Fig. 3.4: Schematic representation of a 

coccolithophorid cell taking up dissolved 

inorganic carbon. Internally this carbon is fixed as 

CO2 in the chloroplast (green) while it is 

precipitated as CaCO 3 in the coccolith production 

vesicle (grey). 

will give additional information regarding the nature of the calcification response towards 

changing carbonate chemistry by automatic quantification of coccolith numbers, calcite contents


and morphology which allows for comparison with results obtained in 3.5.2. Finally, the results 

on coccolithophorid photosynthesis and calcification in response to increasing CO2 will directly 

feed into the biogeochemical modelling projects 1.3 and 5.2. 

Together, the results will greatly enhance our understanding of the effects of ocean acidification 

on calcification and photosynthesis. This in turn is a prerequisite for predicting the future of 

marine coccolithophores in a high CO2 ocean. 

Work Schedule 


Calcification / Photosynthesis 

dependence of the three 

coccolithophores on CO 2 / DIC 

Identifying their pH sensitivities 

Assessment of inorganic carbon fluxes 

under selected CO2 / pH conditions 

(MIMS) 

Data assessment and identification of 

potential optima and thresholds 

Collaboration within 3.1 



- Identifying the calcification and photosynthesis dependencies of Emiliania 

huxleyi, Coccolithus pelagicus and Calcidiscus leptoporus on CO2, respective 

DIC concentration. 

- Identifying the pH sensitivity of particulate inorganic (PIC) and organic 

carbon (POC) production in these three species. 

- Assessment of dissolved inorganic carbon uptake fluxes under selected CO2 / 

pH conditions. 

Month 18 

Month 24 

Month 30 

- Data assessment and identification of potential optima and thresholds Month 36 

- Collaborative work: Gene expression patterns / MIMS Month 30 

155

156 


Subproject 3.1.2: Transepithelial calcification processes in the hermatypic cold-water coral 

Lophelia pertusa (Scleractinia) 

(Armin Form, IFM-GEOMAR) 


The hermatypic cold-water coral (CWC) Lophelia pertusa (Fig. 3.5) will be cultivated under 

standardized conditions and challenged by different pCO2manipulative 

long- and short-term experiments (IFM-GEOMAR, 

Kiel): 

Long-term experiments (according to the BIOACID pCO2 core levels): 

- Phase I: 12 months: 380, 560, 700 and 980 ppm CO2 at 7°C 

- Phase II: 12 months: 380, 560, 700 and 980 ppm CO2 at 11°C 

Short-term experiments (to accommodate synergistic aspects): 

- 4 phases ranging from weeks to max. 3 months: various pCO2 

levels and temperatures 

Physiological functioning of the calicoblastic epithelium from the corals 

cultivated under these conditions will then be determined in ex-vivo 

measurements in Ussing chamber and patch-clamp experiments 

(collaboration with M. Bleich & K. Suffrian, Institute of Physiology - 

CAU, Kiel). These techniques allow the determination of transepithelial 

Fig. 3.5: Lophelia pertusa 

during calcification 

measurements under 

elevated pCO2's using 

modified alkalinity 

anomaly technique 

voltage, transepithelial resistance and equivalent short-circuit currents as characteristics of 

epithelial and/or cellular function (for methodical details see subproject: 3.1.4). Using 

pharmacological tools and changes in the solute composition on either site of the epithelium, we 

aim to uncover the transepithelial pathways for calcium and carbon during the calcification 

process (milestone 2, M2) and their sensitivities to ocean acidification by identifying the involved 

ion transport proteins for the corals (M3). In this context we will further address the issues of 

how corals can regulate pH in the calicoblastic cells, and how they remove protons produced 

during the calcification process, respectively. 

In addition to the core questions, the experimental setup is designed for a maximum of crosscollaborations 

with several BIOACID subprojects outside 3.1: 

- For characterizing the pH-gradients and Ca 2+ -concentrations on the different sites of the 

calicoblastic epithelia, microsensor measurements will be done in collaboration with Dirk de 

Beer (MPI Bremen, subproject 3.4.2). These measurements should complete our 

understanding of the epithelial functioning (M2 & M3), and are invaluable precursors for 

the Ussing chamber and patch-clamp experiments. 

- During the long-term experiments carbonate substratum (coral skeleton) will be built 

through the calcification process. This material will be analysed for microchemical and 

structural changes of crystal growth (M5) in a collaboration with Jan Fietzke and Thor 

Hansteen (IFM-GEOMAR Kiel, subproject: 3.2.4). 

- In a collaboration with Alban Ramette (MPI Bremen, subproject 4.1.4) whole coral 

branches from long- and short-term experiments will be analyzed for their microbial 

community composition and their sensitivities to alterations of environmental parameters 

(pH, CO2 concentration, temperature). Molecular techniques will be used to obtain a high


resolution description of microbial community shifts in the different experimental 

treatments (M6). 

- During short-term experiments the synergistic effects between elevated pCO2-levels and 

hydrostatic pressures of 0.1 – 5 MPa to coral physiological performance (PP) will be 

investigated in a joint study with Laurenz Thomsen and Giselher Gust (Jacobs University 

Bremen, subproject 1.2.5) (M7). 

For complementary comparisons between cold- and warm-water corals, results of this project 

will be set in context with those of Ralph Tollrian (Ruhr University Bochum, subproject 3.2.2), 

Jelle Bijma (AWI Bremerhaven, subproject 3.3.1) and Anton Eisenhauer (IFM-GEOMAR Kiel, 

subproject 3.3.2). 

All these collaborations in addition to the subproject internal findings should result in a 

comprehensive integrated view about the effects of ocean acidification on the main hermatypic 

cold-water coral Lophelia pertusa (M8). 

Schedule 


Method development & improvement 

Long-term experiments (Phase I & II) 

Short-term experiments on selected 

aspects 

Collaborative Research within project 

3.1 (MIMS, real-time PCR) 

Sample processing & Measurements 

Data analysis and interpretation 

Joint publication of results 



- Transfer and modification of physiological methods/tools (M1) Month 06 

- Characterization of coral epithelial ion transport mechanisms (M2) Month 18 

- Quantification of changes in ion transport mechanisms in response to prolonged 

elevated CO2 (M3) 

Month 36 

- Collaborative work: gene expression patterns / MIMS (M4) Month 30 

- Quantification of microchemical and -structural changes due to prolonged 

elevated CO2 (M5) 

Month 33 

- Quantification of microbiological community sensitivity due to elevated CO2 

(M6) 

Month 33 

- Characterization of synergistic effects between pressure and elevated CO2 on PP 

(M7) 

Month 36 

- Comprehensive data set on the response of cold-water corals to ocean 

acidification (all experiments) (M8) 

Month 36 

157

158 


Subproject 3.1.3: Calcification & ion homeostasis in the phylum mollusca in response to 

ocean acidification 

(Frank Melzner, (IFM-GEOMAR), Magdalena Gutowska, Magnus Lucassen, Hans O. Pörtner 

(AWI Bremerhaven)) 


We will study model organisms from both, classes bivalvia and cephalopoda, integrating 

genomic and physiological approaches. According to areas of expertise, three work packages 

have been assembled, which will be targeted by two PhD students that will be jointly supervised 

by the WP–leaders (Frank Melzner (FM), Magdalena Gutowska (MG), Magnus Lucassen (ML), 

Hans Pörtner (HP). 

A good tolerant model species is available with the 

cephalopod S. officinalis, currently in culture at the 

AWI Bremerhaven (see Fig. 3.6). A subtracted cDNA 

library (elevated pCO2 vs. control) has recently been 

established for S. officinalis gill tissue (Melzner, 

Lucassen, et al., in prep.), further sequence 

information will be obtained using the next-generation 

sequencing technology available to the Kiel 

Excellence Cluster ‘Future Ocean’. Two Baltic Sea 

bivalve species (Mytilus edulis and Arctica islandica) 

are presently emerging as sensitive model organisms 

within the Excellence Cluster working groups of F. 

Melzner (Ocean acidification, A1) and P. Rosenstiel 

(Marine Medicine, B2). For both organisms, large 

fractions of the transcriptome are presently being 

sequenced (Rosenstiel, Phillip, Melzner, work in 

progress). Availability of large amounts of genomic 

information is an absolute prerequisite for meaningful 

hypothesis-driven research targeting selected crucial 

processes. Incubations of animals for extended periods 

of time under realistic pCO2 (e.g. 380 – 1400 ppm) 

and temperature regimes will be performed using the 

Fig. 3.6: Growth and calcification in the 

cuttlefish Sepia officinalis incubated under 

∼6,000 ppm CO2 (red) and control 

conditions (black). CaCO 3 accretion shown 

as bars, calcified structure shown shaded 

grey in the schematic drawing. 

IFM-GEOMAR / AWI CO2 incubation systems and wet labs and in close collaboration with 

several projects of theme 2 (i.e. 2.1.1, 2.1.3, 2.2.1, 2.2.2, 2.3.1, 2.3.2), as the calcification 

machinery strongly depends on intracellular ion- and pH homeostasis. 

WP I: Gene expression profiling (ML, FM): Based on available sequence information, 

representative transcripts will be monitored in relevant tissues (gill, calcifying tissues of the 

mantle margin) of model organisms exposed for various periods (time course series of up to 8 

weeks) to various intensities of abiotic stressors (CO2, T). Main processes targeted using 

techniques such as realtime PCR and in situ hybridization will be the (i) ion regulatory 

transcriptome in the gills (e.g. Na + /H + exchanger (NHE), Na + /K + ATPase, Cl - /HCO3 - -exchanger 

(AE1), Na + /HCO3 - -cotransporter (NBC1) etc.), (ii) mRNA of proteins that are important for Ca 2+ 

transport / calcification, in both, gills and calcifying epithelia (e.g. Ca 2+ ATPases, Ca 2+ channels, 

AEs, etc.). Based on their plastic expression patterns, essential transporters involved in the 

response to high CO2 / T stress will be identified. Regulated transcripts then will be studied in


more detail on the protein / cellular level (see below). A second approach will be more 

explorative and focus on differentially expressed genes using approaches such as cDNA library 

generation via subtractive suppression hybridization (SSH). This will help to identify (novel) 

essential ion regulatory proteins involved and aid in developing novel hypotheses on how 

elevated pCO2 might affect yet unconsidered physiological processes. Results of this approach 

will also be verified by realtime PCR and further (functional) characterization. There will be a 

strong collaborative link to sub-project 2.3.2, as similar approaches will be used in both projects. 

WP II: Ion regulatory proteins and epithelia function (MG, ML, FM): In a first step, possible 

changes in ion transport protein composition of important epithelia (gill, calcifying interfaces) 

will be assessed using Western Blots and immuno-histochemical methods (e.g. Deigweiher et al. 

submitted, Melzner et al., submitted). Further, enzyme ion-transport capacity (Ca 2+ ATPase, 

Na + /K + ATPase) will be determined employing photometric enzyme tests (FM, ML). In a second 

step, important epithelia will be isolated, and their functional characteristics will be determined 

using Ussing-chamber techniques (MG, in close collaboration with Markus Bleich and his 

working group, as well as with partners from sub-projects 2.1.1, 2.1.3, 2.3.2). Finally, changes in 

mRNA, transporter protein concentration and transporter activities can be correlated with 

functional changes in ion-transporting epithelia. 

WP III: Ionic composition, acid – base physiology, calcification (HOP, FM): The third work 

package will focus on how the ion regulatory machinery studied in WPs I and II affects wholeanimal 

performance, namely (i) ion composition of body fluids (all major cations and anions in 

hemolymph, coelomic fluid, renal fluid, extrapallial fluid) (HOP, FM), (ii) acid base parameters 

of extra- and intracellular compartments (HOP), (iii) oxygen transport protein physiology (FM, 

link to , 2.2.1 and 2.2.2), and (iv) calcification rates at various time points during acclimation to 

elevated pCO2 (FM, collaboration with sub-project 4.1.2). Calcification rates will be determined 

using the ∆TA method, or by directly determining CaCO3 contents of shells in fast calcifiers 

(cephalopods). Calcification performance in our model organisms will be compared to that of 

other molluscs (pectinids, 2.1.3, pteropods 3.2.1). In a collaboration with Dirk DeBeer (see 

3.4.2), it is further planned to measure the ionic composition of extrapallial fluid directly at the 

calcification site using microsensors. This work package will provide information on the degree 

of acidification that can be compensated by the ion-regulatory and physiological machinery. 

Schedule 


Incubation experiments, tissue / body 

fluid sampling, calcification rates, 

sample preparation 

cDNA library construction / analysis, 

sequencing, gene identification 

Gene expression analysis (qRT PCR) 

Protein functional properties (Enzyme 

activities, Western blots, immuno - 

histochemistry) 

Epithelial function (Ussing chambers) 

Acid – base parameters (intra / 

extracellular), ion composition of body 

fluids 


159

160 



Collaborative work within the project 

(MIMS, Gene expression studies) 







- Dataset on calcification rates, tissue / body fluid samples month 09 

- Dataset on gene expression patterns month 15 

- Dataset on protein functional properties month 21 

- Dataset on acid - base / pH regulation month 21 

- Dataset on epithelial function month 33 

- Completion of PhD theses / defense month 36


Subproject 3.1.4 Sea urchin membrane transport mechanisms for calcification and pH 

regulation 

Markus Bleich, Kerstin Suffrian (CAU Kiel), Frank Melzner (IFM-GEOMAR) 


Preliminary work for the project: 

Animals: We have collected Strongylocentrotus droebachiensis from the western Baltic Sea and 

maintain them for continuous use in the aquarium facilities of the IFM-GEOMAR in Kiel. In 

addition continuous supply of sea urchins has been organised using routine cruises of IFM- 

GEOMAR research vessels to substitute the existing stock. Fertilized eggs are routinely 

generated and different embryonic stages and larvae are available. The maintenance of adult as 

well as embryonic to larval sea urchins can be performed under any CO2 partial pressure between 

380 and 1,400 ppm in a newly established facility. 

Genomics: In a collaborative project within the cluster of excellence “Future Ocean” (Frank 

Melzner, Philip Rosenstiel), sequencing maps of S. droebachiensis are currently aligned with the 

genome of S. purpuratus and provide the data base for the search of homologous genes coding 

for membrane transport proteins involved in H + , HCO3 - and Ca 2+ transport, and for proteins 

involved in the generation of membrane voltage as a key driver of electrogenic transport 

processes. 

Infrastructure: One lab is available to the group and equipped for all measurements described in 

the work plan. A setup is ready for the installation of the imaging equipment which has been 

applied for. 

Key questions: i. which membrane proteins are responsible for ion transport? ii. which ion 

channels are pH sensitive? iii. which mechanisms are involved in pH and Ca 2+ homeostasis? iv. 

does chronic environmental CO2 incubation change the functional properties of the cells? v. does 

chronic environmental CO2 incubation change the expression of transport proteins? 

Methods: 

Patch clamp analysis: Whole cell conductance measurements will be performed on isolated 

cells from embryonic stages to determine the basic electrophysiological properties and to 

characterize the ion transport mechanisms involved in pH homeostasis, Ca 2+ metabolism and 

membrane voltage generation. The sensitivity of the respective ion channels towards changes in 

pH/CO2 will be investigated on the single channel level in isolated membrane patches. 

Microfluorimetry: Cells from embryonic stages will be challenged by acute changes in ambient 

pH/CO2. They will be monitored via pH sensitive fluorescent dye indicators for their ability to 

counter-regulate as a measure for the expression of pH regulatory mechanisms. The same 

experiments will be performed after chronic incubation at increased CO2 partial pressures. 

Fluorescence measurements of cytosolic Ca 2+ will reveal the status of Ca 2+ metabolism with 

respect to its role for calcification and as a second messenger molecule. 

Molecular biology: Realtime PCR of membrane transport protein candidate gene mRNAs will 

be performed to validate the results suggested from functional studies and database mining. The 

influence of chronic CO2 elevation on the expression level will be investigated. 

161

162 


Networking with projects and sub-projects in other themes: 

The projects under themes 1-5 provide an excellent platform for interactive networking on a 

methodological as well as on a data driven basis. Obvious contact points are within project 1.1 

(acclimation versus adaptation; subproject 3/4) with respect to the analysis of genetic and 

proteomic changes in calcifiying organisms on long-term exposure to CO2. Project 2.1 is 

especially well suited for a strong interaction since similar questions are asked and similar 

methods are used here for the analysis of sea urchin oocytes, while subproject 3.1.4 takes over 

after fertilization. We expect exchange on cellular mechanisms and technical progress. Also 

project 2.3 has one focus on cellular mechanisms of CO2/pH sensitivity on development, growth 

and metabolism and finally interaction with subproject 4.1.2 could provide added value in the 

discussion how survival and performance of early life stages are affected. 

Schedule 


Set-up of facilities and instrumentation, 

hiring of PhD student (M1, M2) 

Patch clamp experiments (M3) 

Microfluorimetry (M4) 

Data base mining and gene 

identification (M5) 

Quantification of specific membrane 

transport mechanisms (M6) 

Gene expression analysis (M7) 


data interpretation (M3 – M8) 





- Setup of a new microfluorimetric imaging equipment (M1) Month 03 

- Characterization of embryonic cells by patch-clamp technique (M2) Month 12 

- Characterization of embryonic cells by microfluorimetry (M3) Month 18 

- Database mining to identify candidate genes for the respective transporters and 

channels (M4) 

- Quantification of specific membrane transport mechanisms in embryonic cells 

after chronic pre-incubation at different CO2 concentrations (M5) 

- Quantification of the expression level of the candidate genes which might be 

functionally relevant at different CO2 concentrations (M6) 

- Generation of a cell model for membrane transport mechanisms in embryonic cells 

(M7) 

Optional: Expansion of measurements to cells from other developmental stages 

according to M2-M7 

Month 21 

Month 33 

Month 36 

Month 36 

Month 36




3.1.1 PhD student 

3.1.2 Postdoc position 

3.1.2 Student contracts 

3.1.3 PhD student AWI 

3.1.3 PhD student IFM- 

GEOMAR 

3.1.4 PhD student 

Subtotal 

Consumables 

3.1.1 

3.1.2 

3.1.3 AWI 

3.1.3 IFM-GEOMAR 

3.1.4 

Subtotal 

Travel 

3.1.1 meetings 


3.1.3 meetings AWI 

3.1.3 meetings IFM- 

GEOMAR 


Subtotal 

Investment 

3.1.1 MIMS maintenance 

3.1.4 

Subtotal 

TOTAL 


163


3.1.1 

164 


Personnel costs: Most of the work will be done by a PhD candidate, to be employed for three 

years. 

Consumables, chemicals, gases, analyzes: Money for consumables will be needed for various 

analyzes such as elemental composition analyzes on a mass-spectrometer, nutrient analyzes, 

chemicals, gases and isotopic analyzes. 

Investments and maintenance: The membrane-inlet mass-spectrometer will need frequent (at 

least annual) maintenance to ensure high data quality. 

Travel costs and publication charges: Travel of the PhD student and one supervisor to an 

international conference and publication charges. 

3.1.2 

Personnel costs: 

Postdoc position for Armin Form: Due to the complex nature of the proposed 

research program and the very challenging animal model, an experienced researcher is needed for 

this project. Armin Form is currently finishing his Ph.D. thesis on effects of elevated pCO2-levels 

to cold-water corals and arctic coralline red algae and has thus in-depth understanding of both, 

the relevant biogeochemical processes and the cultivation methods (see 3.1.2.iii). 

Student contracts (HiWi): Student helpers are needed to support the extensive cultivation work as 

well as for the routine analysis in the lab (e.g. water chemistry monitoring). They will also be 

trained to assist the experiments and will help with sample preparation and processing. For 

adequate assistance, two continuous contracts per long-term experiment will be necessary 

. 

Consumables: 

Coral cultivation: The costs for coral cultivation are estimated to be per year, including 

mainly purchases for food and water treatment (such as filter materials and adsorbers). Additional 

funds of per year will be needed for culturing experiments (aquarium supplies, incubation 

vessels, gas, sea salt). The costs for chemicals are estimated to be per year. These costs 

include purchases of gases, acids, enzyme inhibitors. 

Travel: One international conference per year will be visited by the Postdoc in order to 

disseminate the project results to the scientific community. 

3.1.3 

Personnel costs: Two PhD students will be needed owing to the complexity of the proposed 

research projects. We envision one PhD student to focus on the bivalve and one on the 

cephalopod model. Formally, one student will be based at IFM-GEOMAR, the other at the AWI. 

However, work plans will be arranged in parallel, as to facilitate collaborations between the 

students. 

Consumables: For both model organisms, one sequencing run on the next generation sequencing 

/ 454-platform (P. Rosenstiel, CAU Kiel) needs to be performed on gill tissue extracts to gain


sufficient amounts of transcript information to devise meaningful gene expression experiments. 

One run will cost approximately and will generate 200.000 sequences of 200-300 bp 

length. Reagents for realtime PCR experiments / in situ hybridization need to be purchased 

(approximately ). Significant funds will be used purchasing molecular biological 

reagents (ca. ). Additional funds will be needed for culturing experiments 

(aquarium supplies, incubation vessels, gas, sea salt), for Ussing-chamber experiments (ion 

channel / transporter inhibitors, chemicals) and tissue acid-base analysis (chemicals, pH 

electrodes). 

Travel: Travel of the PhD students to an international conference in years two and three. 

Additional costs for travel between the Institutions (AWI Bremerhaven, CAU Kiel / IFM- 

GEOMAR) will be covered using the home institution’s funds. 

3.1.4 

Personnel costs: The experimental approach utilizes difficult and complicated techniques which 

need a substantial training period. The supervision and training is provided by the PIs. A three 

year contract guaranties a sustained and effective exploitation of the techniques by the PhD 

student. 

Consumables: All consumables covered by the given sum are necessary to perform the planned 

experiments. The higher needs in second and third year are caused by molecular biological 

experiments. 

Travel: Travel of the PhD student and one supervisor to an international conference. Costsharing 

with home institution. 

Investment: The lab is already equipped with a complete experimental setup, which, however, 

has to be upgraded for microfluorimetric measurements. The availability of a separate setup for 

microfluorimetric measurements is critical for the success of the project since about 50% of the 

data will be produced using this equipment. Due to the high workload, time sharing at the 

existing setup is not feasible. 


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Project 3.2: Calcification under pH-stress: Impacts on ecosystem and organismal 

levels. 

(PI: Ralph Tollrian) 

i. Objectives 

Anthropogenic CO2 emissions are reducing ocean pH-values and carbonate saturation state, with 

strongest effects in high-latitude surface waters. In this project we will compare the responses of 

organisms from pteropods, scleractinian corals and red coralline algae, which have been 

identified as key groups for the understanding of community responses of ocean acidification 

(Kleypas et al. 2006). These calcifying organisms are vulnerable to acidification and because of 

their relevant role in their ecosystems these impacts have the potential to cause cascading effects 

and to alter ecosystem stability and functioning. Pteropods dominate zooplankton communities in 

polar regions of both hemispheres, scleractinian corals form the physical structure of the tropical 

reef ecosystems and coralline red algae are cosmopolite in the ocean and found at all depths 

within the photic zones. The thresholds and tipping-points where the different organisms and 

systems respond to acidification, the magnitude of the response and the impact on the 

communities will be determined for different ontogenetic stages. 


Subsaturation with respect to aragonite, the metastable form of calcium carbonate, could begin in 

the Southern Ocean and in sub-Arctic regions before the end of this century (Orr et al. 2005). In 

these cold regions, the only pelagic aragonite-producers are shelled pteropods, e.g. the bipolar 

species Limacina helicina. These shelled pteropods will be among the first organisms 

experiencing carbonate saturation states


the field. Although it is generally assumed that corals are negatively affected at a certain level of 

pCO2 (Hoegh-Guldberg et al. 2007), differential effects in corals (Lough and Barnes 2000) and 

other calcifiers (Iglesias-Rodriguez et al. 2008, Fabry et al. 2008) indicate that the picture may be 

much more complicated than previously thought. Of particular interest is how different 

ontogenetic stages respond to the same stress factor. Possibly, recruitment will be influenced 

before an effect on growth of adult colonies can be detected. In scleractinian corals, settling of 

larvae and early colony formation are critical steps where a reduced calcification will threaten the 

recruit because it lives in competition with overgrowing turf algae, is preyed on by predators and 

is exposed to herbivores which feed on algae and scratch off the fragile young colonies. But also 

coral community compositions may change as a consequence of increasing pCO2. Corals have 

two main modes of larval production: “brooding”, where larvae develop within the colony and 

are released at a relatively large size, or “broadcast spawning”, where eggs and sperm are 

released simultaneously and larvae develop in the water column. The larger larvae of brooding 

species possibly will have an advantage under decreased ph-conditions because they contain 

more reserves, which allow faster initial growth. 

The health of reefs critically depends on recruitment, especially when reefs are devastated by 

coral bleaching or storms. Coral reefs have been shown to possess two alternative stable states 

with coral- or algae-dominated climax conditions and with a limited ability to return into a coral 

dominated state once the tipping point has been exceeded (Mumby et al. 2006). Ocean 

acidification is supposed to influence corals negatively, while macroalgae might even benefit 

from higher CO2-levels. Additionally coralline red algae may decline with a negative effect on 

coral recruitment. Thus, ocean acidification has the potential to influence survival of early stages, 

change community compositions and processes and on a higher level, lead to phase shifts in 

whole ecosystems and alter ecosystem functioning and stability. 

Despite the broad abundance of coralline red algae so far little is known about their calcification 

mechanisms. Recently some studies focussed on the use of coralline red algae as environmental 

recorders (Halfar et al. 2007, Kamenos et al. 2007), showing the strong response on 

environmental changes, displayed in systematic variations in the chemical composition of the 

precipitated carbonates (high magnesium calcite). This observation suggests that coralline red 

algae are promising candidates to study the responses of important calcifiers on ocean 

acidification in a wide range of habitats. 

In a recent short (7 weeks) mesocosm experiment study (Kuffner et al. 2008) at low latitudes, the 

recruitment rate and growth of red algae was shown to be severely inhibited at elevated pCO2. No 

research was however undertaken to determine whether changes in the carbonate structure or 

microchemistry or adaptations occurred over a longer period (i.e. over an annual growth cycle). 

Such changes in the chemical composition would clearly affect the solubility of the carbonates 

formed by those algae. So far no study was done addressing pH thresholds for reproduction and 

growth/calcification of coralline red algae or possible physiological responses to deal with 

decreased ambient pCO2 levels. 

Finally, solubility changes due to chemical modifications of the carbonates directly affect the pH 

buffering capacities of shelf regions where coralline red algae contribute significantly to the 

carbonates deposited in the sediments. 


3.2.1 The group of U. Riebesell has been among the first to study direct effects of CO2-induced 

seawater acidification (Riebesell et al. 1993, Riebesell et al. 2000). Presently, research in his 

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group addresses a variety of OA-related aspects from the cellular to the community level 

(Riebesell 2004, Riebesell et al. 2007). This includes studies on the CO2/pH sensitivity of a 

variety of marine calcifying groups, ranging from coccolithophores and rhodoliths to bivalves, 

and cold water corals. In preparation for the proposed work on pteropods calcification in 

BIOACID, a pre-study is being carried out with Limacina helicina in Ny Alesund, Svalbard, in 

May/June 2008. The group of I. Werner has expertise in cold-life culturing and experimental 

approaches on measuring metabolic rates such as respiration, grazing, and predation of Arctic 

marine invertebrates (Werner et al. 2002, Werner and Auel 2005). Of direct relevance to the 

proposed project are previous studies on the seasonal dynamics of the Arctic shelled pteropod 

species Limacina helicina in epipelagic, partly ice-covered waters in the Fram Strait area and 

Kongsfjorden (Werner 2006). 

3.2.2 The research team has long experience in raising coral larvae. R. Tollrian had successfully 

established coral reef research systems in Munich and conducted settlement and growth 

experiments with coral larvae (Petersen and Tollrian 1991) and population genetic studies in 

corals (Maier et al. 2001, 2005). This work provided the tools for the SECORE (Sexual coral 

reproduction) initiative. D. Petersen initiated the SECORE program and is a leading expert in, 

and developed methods for, breeding and raising corals in closed aquaria systems (e.g., Petersen 

et al. 2006). E. Grieshaber. and W. Schmahl are experts in crystallography. Their methods will be 

applied to studies of the coral skeleton structure. A. Eisenhauer is the head of the isotope 

geochemistry lab at the IFM-GEOMAR. The work proposed here is interdisciplinary and 

involves biology, biodiversity, geo-chemistry, mineralogy, crystallography and material sciences. 

3.2.3 The group of C. Richter has been working on coral reef ecology over the last 12 years, 

with a regional focus on the Red Sea and South-East-Asia (Indonesia, Thailand and China). 

Major findings using novel endoscopic tools were the discovery of biomass-rich and diverse 

communities of sponges in Red Sea coral reef crevices fuelling a significant part of reef 

production (Richter et al. 2001, Richter and Abu-Hilal 2006). The group also contributed to a 

seminal paper on the interaction of currents and plankton explaining the enrichment of plankton 

near coral reefs (Genin et al. 2005). Ongoing research addresses the effect of solibores on coral 

communities of the Similan Islands in the Andaman Sea. The findings of significant differences 

in coral cover and growth between soliton-exposed and soliton-protected communities, and the 

development of a micro-CaveCam form the scientific and technological basis of the present 

proposal. A complementary project on the supply and early recruitment of corals has recently 

been approved within the EU-ITN CalMarO. 

3.2.4 J. Fietzke (IFM-GEOMAR) has a strong research record in developing and applying mass 

spectrometric techniques for trace and ultra-trace elements (e.g. Fietzke et al., 2004, 2006, in 

press). His work focused recently on issues of biomineralisation with marine biogenic carbonates 

(e.g. Heinemann et al. 2008, Rüggeberg et al. 2008). The research group has a strong background 

in carrying out microanalytical studies in biogenic carbonates. T. Hansteen is a specialist in 

microanalytical techniques such as Synchrotron X-ray Fluorescence microprobe (SYXRF; 

Hansteen et al. 2000), electron microprobe (EMP), NanoSIMS (e.g. Zumholz et al. 2007 a, b), 

and has co-supervised a PhD in Marine Ecology (K. Zumholz; Zumholz et al. 2006, 2007c).


iv. Work Programmes, Schedules, and Milestones 

3.2.1 Impact of ocean acidification and warming on sub-polar shelled pteropods 

(U. Riebesell) 

The aim of this project is to study the synergistic effects of ocean acidification and rising 

seawater temperatures on a key component of polar and sub-polar epipelagic communities, the 

shelled pteropods. In a second step, ecological consequences of these effects for different lifestages 

will be assessed. This will be achieved through a collaboration involving experimental 

marine biology, physiology, biochemistry and marine ecology. Specifically, the project will 

address the following aspects: 

1. Calcification/dissolution response 

2. Metabolic response (oxygen consumption, swimming performance) 

3. Life cycle responses (egg and larval development, growth and reproduction) 

4. Ecosystem response (feeding rates, predation, survival) 

Pteropods of the dominant and bipolar genus Limacina (L. helicina or L. retroversa) will be 

collected in Arctic waters (Kongsfjord, Svalbard) and will be reared in the Kings Bay Marine Lab 

under ambient conditions. Pteropods will be held in specially designed, V-shaped aquaria 

equipped with a circulating current system and filled with filtered, cold seawater. Food will be 

provided in form of algal cultures. Ambient pH/pCO2 and temperature will be altered in 

temperature-controlled incubation rooms equipped with a new gas-producing system. 

Methodologies/approaches for aspects 1-4 include: 

1. Calcification rate: 45 Ca incubations, shell weight, fluorochroms; shell composition: 

Mg/Ca, Sr/Ca, possibly boron isotopes; shell dissolution: SEM 

2. Respirometer measurements, digital camera system 

3./4. If culturing attempts turn out successful, life cycle experiments and grazing experiments 

of low and high CO2 exposed Limacina will be started in year two of the project. Grazing 

experiments will be conducted with a major predator, the gymnosomate pteropod Clione 

limacina, which was successfully cultivated previously. 

Links to other projects: The pteropod work in this sub-project will be closely linked to studies 

on mollusc larvae (3.5.1) and mollusc phyiology (3.1.3). Results of this sub-project with 

relevance to the ballast effect will also feed into sub-projects 1.2.5. Estimates of OA-induced 

changes in pteropod calcification will be provided to sub-projects 1.3 and 5.2. Samples of 

pteropod shells will be provided to sub-project 3.2.4 for SEM and electron backscatter diffraction 

(EBSD) analysis of the skeletal architecture as well as high-resolution geochemical analyses 

applying electron microprobe, NanoSIMS, LA-ICP-MS, SYXRF. 

Schedule 


Design & construction of experimental 

set-ups for field- and lab-based studies 

Pteropod CO 2 perturbation experiments 

at field station Ny Alesund 


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Attempts to culture pteropods in IFM- 

GEOMAR culturing facilities 

CO 2 perturbation exps. in IFM- 

GEOMAR culturing facilities 


Combined CO 2 and temperature 

perturbation exps. at Ny Alesund 

Follow-up exps. in IFM-GEOMAR 

culturing facilities 







- Experimental facilities implemented month 06 

- Data set on CO2/pH sensitivity of adult pteropods month 12 

- First practice in culturing of juvenile and adult pteropods month 21 

- Data set on synergistic effects of CO2 and temperature month 24 


3.2.2 Impact of ocean acidification on reproduction, recruitment and growth of 

scleractinian corals (R. Tollrian). 

Our main goals are: 1) Test survival of coral larvae, early colony establishment and growth of 

different species from several geographic regions, to find species specific thresholds and 

geographic differences. 2) Assess competitive ability against algae. 3) Test the hypotheses that 

brooding species will be less influenced compared to broadcast spawning species. 4) Test for 

differences in reproductive investment and growth in brooding corals under different ph 

conditions to assess sublethal stress effects. 5) Compare amount and quality of calcite formation 

of fragments under different ph-conditions and quantify changes in ultrastructure, 

biogeochemistry and material properties. 

Aim 1) The SECORE (sexual coral reproduction) working group has successfully 

established a recruitment program for coral conservation where larvae are collected during mass 

spawning events in the Caribbean, transported to public and research aquaria and raised in these 

closed systems (Petersen et al. 2006). We will be able to obtain larvae from broadcast spawning 

species from different locations during the highly predictable yearly spawning events: a) 

Maldives (March), b) Okinawa/Japan (May), c) Curacao/Caribbean (August/September), d) 

Mozambique (October), e) Great Barrier Reef (December). Experiments studying fertilisation, 

larval survival and early colony formation will be conducted during field studies. For long-term 

lab studies, larvae will be collected and transported to our aquaria facilities following an 

improved method (Petersen et al. 2005a). Larvae will not settle in filtered seawater because they


recognize suitable substrates based on relevant biofilms. Specifically designed ceramic tiles will 

be offered as settlement substrate (Petersen et al. 2005b). The larvae will be raised under five 

different pH conditions in closed aquaria systems. Each treatment will be fully replicated. We 

will measure larval survival, settlement, transformation and early colony growth and will identify 

threshold conditions. A share of the larvae will be settled according to the SECORE protocol 

under standard conditions and experiments with different pH treatments will be started after the 

young colonies have been established. This experiment will allow separating OA effects on larval 

survival from effects on juvenile growth. We will compare the species composition of successful 

settlers in the different treatments to test whether species differ in their tolerance to low pH 

conditions. By repeating the experiments with larvae from different locations we will be able to 

cover a wide range of different species and to compare thresholds within and between geographic 

regions. 

Aim 2) The competition with algae can be tested with the same experimental design but 

with different amounts of turf algae in the systems. 

Aim 3) The brooding coral species Pocillopora damicornis and Favia fragum are 

available in the aquaria facilities in Rotterdam and Munich where they reproduce in closed 

systems. Colonies and larvae can be obtained from Rotterdam. The experiments above will be 

repeated with larvae from brooding species to test for different thresholds and differences in 

vulnerability to lower pH levels. 

Aim 4) Sublethal effects on adult colonies may be measurable in reduced energy 

availability for growth, reproduction or both. To test for sublethal effects, adult colonies of P. 

damicornis and F. fragum will be raised in aquaria with different ph-levels to measure 

investments into reproduction and growth. Colonies of F. fragum are ideal for this purpose, 

because they reach maturity extremely fast, after just one year. 

Aim 5) Another mode of recruitment in corals is fragmentation. Fragments from 

individuals can be exposed to different treatments and will allow to test the effects of OA on 

growth without confounding effects of genotypic variation. We will use fragments of fast 

growing, branching Acropora species for the experiment (larvae of the same species will be used 

in the experiments with sexually produced larvae). The experiment will run 12 month but can be 

prolonged if necessary. Within 12 month the branches will grow up to 10 cm under good 

conditions. This growth should provide enough new skeleton material, even under low pH 

conditions, for isotope analysis and analysis of material properties. Additionally we will analyse 

material of the corals raised from larvae. Microsensor technology will allow measuring Ca 2+ , CO2 

and CO3 2- in the cell regions where calcification takes place. These analyses should reveal 

whether corals sacrifice growth rate, skeleton stability or both under stress conditions 

Links to other projects: 

2.1.1: In this project responses of early life stages of different organisms to acidification will be 

analysed. In collaboration we will study responses of coral eggs to trace mechanisms of 

acidification effects on the earliest ontogenetic stages. Our study will provide additional 

information for theme 2 (Performance characters: reproduction, growth and behaviours in animal 

species). 3.1.2: We will compare calcification mechanisms and responses to acidification 

between tropical and cold water corals. 3.2.3: Our project with a focus on early live stages of 

corals directly complements project 3.2.3 where the focus is on responses of corals in nature. 

3.2.4: The measurement of minor and trace elements (e.g. Sr, Mg, Ba, S) at micron scale 

resolution using electron microprobe (EMP) and LA-ICP-MS provides a tool to analyze changes 

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in chemistry within carbonate material such as coral skeletons and will be done in conjunction 

with project 3.2.4. We will deliver coralline red algae from our coral incubation experiments. 

3.3.2: For the analysis of the biogenic carbon composition we will collaborate with A. Eisenhauer 

and will apply his isotope techniques. 3.4.2: For the analyses of the cellular mechanisms of 

calcification under different ph-levels we will collaborate with D. de Beer and will apply his 

microsensor techniques. 4.1.2: this project will study effects on larvae of selected organisms and 

results will be compared to our results on coral larvae. 4.1.3: Our study includes competition 

experiments between early coral colonies and macroalgae. These results will be compared with 

the results on tropical macroalgae of project 4.1.3 and will provide relevant information about 

regime-shifts for theme 4 (Species interactions and community structure: will ocean acidification 

cause regime shifts?). 

Schedule 


Design & construction of experimental 

set-ups for field- and lab-based studies 

Experiments with brooding corals in 

Rotterdam and transport to Bochum 

Experiments with coral larvae from 

spawning species 

Competition experiments between 

coral larvae and macroalgae 

Field study during coral spawning 

Analysis of reproductive investment of 

adult brooding corals 

Microsensor analysis 

Comparative analysis of coral 

skeletons 







- Experimental facilities implemented month 06 

- Data set on CO2/pH sensitivity of larvae from brooding corals month 12 

- Data set on CO2/pH sensitivity of larvae from spawning corals month 21 

- Data set on CO2/pH sensitivity of adult brooding corals month 24 

- Microsensor analysis of calcification in corals grown at different CO2/pH month 28 

- Analysis of ultrastructure, crystallography and material properties of corals grown 

at different CO2/pH 

month 33


3.2.3 Coral calcification in marginal reefs (C. Richter) 

The aim of this project is to study the effect of natural oscillations of the aragonite saturation 

horizon, via the shoaling and breaking solitary internal waves, on coral calcification in marginal 

reefs. Marginal reefs mark the transition between flourishing coral reefs on the one hand, and 

rocky bottom devoid of macroskeletal organisms on the other. Such reefs occur under conditions 

where calcium carbonate accretion balances corrosion/erosion. In the Similan Islands, in the 

Andaman Sea of Thailand, the transition from reef to rock occurs over scales of only 100 m, 

between the solibore-protected East and the solibore-exposed West sides of the island. Marginal 

reefs at the flexion point provide a unique opportunity to assess tipping points in aragonite 

saturation state with regard to net coral calcification, and to explore the synergies between 

aragonite saturation, and other parameters (e.g. temperature, nutrients) associated with solibores. 

Specifically, the project will address calcification/decalcification and photosynthesis/respiration 

of corals using in situ microsensors and in situ chambers (Funke chamber). The work will involve 

three Steps: (1) technological development, (2) testing/deployment instrumentation in the field 

and (3) field measurements. 

As aragonite saturation co-varies with temperature, the fine-scale vertical and horizontal 

temperature field will be monitored with an array of Tidbit temperature loggers (Onset comp. 

Corporation). Two moored multiparameter-loggers (T, S, O2, pH, Fluorescence, OBS), a Rapid 

Access Sampler, RAS (McLane Inc.) and ZPS (McLane Inc.) available at AWI will allow 

autonomous pre-programmed sampling of alkalinity and nutrients. A novel in situ respirometer 

developed by the group (Funke chamber) combined with RAS will be used to measure timeseries 

of photosynthesis, respiration and calcification in situ, where calcification will be measured 

using the alkalinity anomaly technique. Oxygen, calcium, pH and novel CO3 2- microsensortechniques 

developed at MPIMM (D. de Beer) will be combined with the micro-CaveCam 

developed by the AWI group to measure photosynthesis and calcification in the immediate 

vicinity of corals along a spatio- temporal gradient of aragonite saturation state. 

Research will concentrate on the massive scleractinian coral Porites lutea, but to assess the 

general validity of our findings we will investigate also other abundant genera, including 

Acropora and Pocillopora, as well as purely heterotrophic forms such as Tubastraea micranthus. 

To test the equipment and laboratory runs, coral specimens will be collected in Koh Racha island 

off Phuket and reared in the Phuket Marine Biologial Laboratory (PMBC) under ambient 

conditions in a through flow system. Variable pH/pCO2 and temperature will be supplied in 

temperature-controlled incubators equipped with a CO2 bubbling system developed according to 

the blue-print of the Bioacid consortium. The analytical techniques and calculations for the 

incubation experiments follow Schneider and Erez (2006), for the microsensor set-up cf. Weber 

et al. (2007). 


Methodology of combining micro-CaveCam and microsensors will be carried out in close 

cooperation with the microsensor group of D. de Beer (MPIMM) (project 3.4.2). Field work will 

be coordinated with the other teams working in tropical waters, including R. Tollrian (project 

3.2.2.) and K. Bischof (project 4.1.3.), using common field stations as a logistic basis. High 

resolution records of pH (boron isotopes), and nutrients (P:Ca) in relation to linear growth and 

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skeletal density of Porites skeletons will be examined with J. Fietzke (project 3.2.4.) and A. 

Eisenhauer (project 3.3.2.). Results of this project will have important ramifications for the 

membrane studies in coldwater corals by A. Form (project 3.1.2), and provide baseline data for 

theme 2 and theme 4. 

Schedule 


Recruitment of staff, technological 

development 

Collection of corals and laboratory test 

runs 

In situ microsensor and incubation 

experiments 


Data analysis 





- In situ instrumentation developed and tested month 06 

- Laboratory data set on CO2/pH sensitivity of corals month 12 

- First field data set on CO2/pH sensitivity of corals (end NE monsoon) month 18 

- Second field data set on CO2/pH sensitivity of corals (end SW monsoon) month 24 

- Data set on synergistic effects of CO2, temperature and nutrients month 24 

- Data analysis, submission of theses and manuscripts month 36 

3.2.4 Impact of ocean acidification on coralline red algae (J. Fietzke) 

Coralline red algae are important carbonate forming species both at low and high latitudes. In the 

former, they are important for the development and stability of the coral reefs. In the latter, they 

are the dominant benthic marine carbonates. This project therefore will investigate the response 

of different species of coralline red algae in these two contrasting environmental settings to ocean 

acidification. In particular, this will include: 

1. Culturing experiments with CO2 perturbations using a high-latitude species 

2. Micro-scale skeletal architecture and geochemical analyses to compare cultured with insitu 

specimens. 

3. Comparison of the response of coralline red algae and corals from the same location at 

low latitudes, which has a high natural pH variability (linked to project 3.2.3). 

4. Investigation of the responses in long-lived coralline red algae to the anthropogenic 

driven CO2 increase since the beginning of the industrial revolution.


5. Comparison between different pH responses of macroalgae and macroorganisms (linked 

to project 2.1.3; 4.1.1; 4.1.3). 

To achieve these objectives, Lithothamnium topiphorme 

will be cultured from existing specimens available at 

IFM-GEOMAR. The samples will be grown for 12-14 

months, using the five standard pH levels within the 

BIOACID project. This will be done in conjunction with 

U. Riebesell and A. Form (project 3.2.1 and 3.1.2 

respectively). The light irradiance and temperature values 

used would be similar to those occurring at the collection 

site. In-situ analyses of the calcification process will be 

carried out (in conjunction with project 3.4.2). The 

skeletal architecture of naturally grown sample material 

will be analysed using SEM, electron backscatter 

diffraction (EBSD). This will be complemented by high-resolution geochemical analyses 

applying electron microprobe, NanoSIMS, LA-ICP-MS, SYXRF. After the completion of the 

culturing, this sample material will be subjected to the same analytical tools, to investigate the 

effects of the pH treatment. The architectural and geochemical analyses will be available for 

other cultured material (project 3.2.1 and 3.1.2) to understand how different calcifying organisms 

react to different pH. 

To further understand existing adaption to natural pH variability within coral and red algae 

communities in tropical reefs, samples from both taxa, grown in close proximity, will be used for 

geochemical comparison. This material and accompanying temperature and pH data will be made 

available through collaboration with project 3.2.3. Architectural and geochemical analyses of 

new coral and coralline red algae growth will be carried out (on material from project 3.2.2) to 

determine how changes in pH are affecting newly establishing colonies. 

Finally, using samples of long-lived species e.g. Clathromorphum nereostratum, we will study 

whether ocean acidification since the industrial revolution has impacted on the micro-architecture 

and geochemistry of coralline algae over decadal timescales. Thus, the rate of adaptation and 

changes from short-term studies can be compared to changes over extended timescales in order to 

predict the long-term effects of progressing ocean acidification. 


Electron microprobe map of sulphur 

variation in coralline red algae 

3.2.3: Coral and coralline red algae samples from the same habitat will be compared with respect 

to their architectural and microchemical responses on variable pH and temperature conditions. A 

comparable approach focussing on cultured specimen from pH controlled coral larvae 

recruitment experiments will be used in collaboration with 3.2.2. Both collaborations (3.2.2 and 

3.2.3) will contribute to the understanding the pH influence on the initial growth of coral 

colonies. Experience of U. Riebesell and A. Form (project 3.2.1 and 3.1.2 respectively) will be 

utilised in culturing the coralline red algae. In return our experience of geochemical and 

architectural analyses will be utilised to understand the effects of pH on pteropod and cold water 

coral samples from this two projects. 4.1.1 and 4.1.3: Joint experiments on the calcification 

processes in coralline red algae and other macroalgae will be undertaken focussing on the 

competition between different algae groups. 3.4.2: Joint experiments to study the calcification 

process on a cellular level will be carried out using microsensor techniques. T. Brey (project 

2.1.3) will provide time-series data for the coralline red algae samples from Svalbard, Norway. 

177

Schedule 

178 



Set-up of culturing facility 

CO 2 perturbation experiment 

Analyses of skeletal architecture from 

in-situ material 

Geochemical analyses of in-situ 

material (incl. archive material & 

corals) 

Analyses of skeletal architecture from 

cultured material 

Geochemical analyses of cultured 

material 







- Experimental set-up for CO2/pH implemented month 03 

- Baseline skeletal architecture and microchemistry month 09 

- Geochemical comparison between coexisting red algae and corals month 12 

- Long-term data set on centennial archive month 15 

- Collection of cultured samples month 15 

- Skeletal architecture and microchemistry of cultured samples month 21 

- Data set on CO2/pH sensitivity of cultured samples month 24 

- Completion of manuscripts month 30 



3.2.1 

3.2.2 

3.2.3 

3.2.4 

Subtotal 

Consumables 


3.2.1 

3.2.2 

3.2.3 

3.2.4 

Subtotal 

Travel 

3.2.1 

3.2.2 

3.2.3 

3.2.4 

Subtotal 

Investments 

3.2.1 

3.2.2 

3.2.3 

3.2.4 

Subtotal 

Other costs 

3.2.1 

3.2.2 

3.2.3 

3.2.4 

Subtotal 

TOTAL 


3.2.1 



Personnel costs: 1 Ph.D. position (Jan Büdenbender, per year), student helpers (€ per year) to 

assist with Svalbard experiments and lab culturing of pteropods and food algae 

Consumables: synthetic gases, Ca isotopes, Mg,, Sr and Ca analyses, fluorochromes, SEM 

analyses, reagents for artificial media, nutrient and Winkler analyses (€ in years 1 and 2) 

179

180 


Travel: 2 trips for 3 persons (Ph.D. student + 2 student helpers) to Ny Alesund, Svalbard each), 

meals and accommodation /person/day) and lab fee ( /person/day) at field station Ny Alesund 

for 2 visits of 28 days each for 3 persons: total , 2 trips for 2 persons (Ph.D. student + Co-PI) per 

year to international conferences), attendance of BIOACID workshops and annual meetings. 

Investments: Culturing facilities for pteropod maintenance (€), 12 experimental laminar-flow 

incubation vessels for CO2/temperature controlled incubations (x) 

Other costs: transport of equipment to Svalbard (€). 

3.2.2 

Personnel costs: 1 Ph.D. position (Sebastian Striewski, per year), student helpers (per year for 

year 1 and 2, for year 3) to assist with field experiments, lab experiments, calibration and 

control of the CO2-supply systems and chemical parameters. 

Consumables: synthetic gases, isotope analyses, ultrastructure analyses, reagents for artificial 

sea water, supply for culturing facility, nutrient and Winkler analyses, settling substrates for coral 

larvae (€ in year 1, in year 2, in year 3) 

Travel: 2 trips for 3 persons (PI, Ph.D. student + student helper) for field experiments to Lizard 

Island (Australia) ( each + bench fees and accommodation for 35 days: AUD 113/person and 

day), 3 short trips for 2 persons to field stations in different geographic regions for larvae 

collections after mass spawning events, 2 trips for 2 persons (Ph.D. student + PI) for year 2 and 3 

to international conferences ( each), attendance of BIOACID workshops and annual meetings (€ 

per year). Attendance of microsensor workshop, collaborative works in Rotterdam, Kiel and 

Munich for Ph.D.. Total: € . 

Investments: Culturing facilities for coral larvae and adult colonies. Separate systems for each 

CO2-level, including tanks, pumps, lamps, water purification, systems for CO2- and temperaturecontrolled 

incubations (total € ). 

Other costs: Coral larvae rearing tanks with circular flow, mobile CO2- and temperature-control 

systems for the field (total € ). Air freight (year 1 and year 2; total € ) total € 

3.2.3 

Personnel costs: 1 Ph.D. position (N.N., € per year) is needed to carry out the combined micro- 

CaveCam-microsensor work in situ and in the lab, on natural and simulated in situ variations of 

aragonite saturation state on coral calcification in marginal reefs. The student is to be trained in 

the BIOACID consortium using the expertise available in the various institutions, e.g. in 

methodology (CO2-experiments, IFM-GEOMAR) and instrumentation (microsensors, MPIMM). 

Student helpers (6 mo. x 80 h, € per year) are needed to assist with Thailand experiments and 

chemical analyses of samples. 

Consumables: microsensor materials, reagents for chemical analyses, glass- and plastic ware for 

coral maintenance, CO2 gases (€ in years 1 and 2; € in year 3) 

Travel: 1 flight (year 1) and 2 flights (year 2), each for 3 persons (co-PI, Ph.D. student, 1 student 

helper) to Phuket, Thailand (€ and € ); contribution to Accommodation/Living expenses (€ 

/person/day for 3 visits á 20 days for 3 persons); attendance of BIOACID workshops and annual 

meetings (€ per year, total € );


Investments: Microsensor set-up (€ ; year 1); in-situ respirometer (custom-built, € , year 1) 

Other costs: Transfers Phuket-Koh Racha/Similans (shared cost € /boat/day for 3 visits á 10 

days; total € ), Air freight (year 1 and year 3; total € ). 

3.2.4 

Personnel costs: Postdoc for 30 months (Laura Foster, € per year). In-depth experience of 

skeletal architecture and geochemical analyses is essential for this project. The named postdoc 

has a strong background in this type of research (e.g. Foster et al. 2008a+b, Foster et al. in 

review). She has extensive experience with a large suite of microchemical techniques e.g. SIMS, 

LA-ICP-MS, EMP, XAFS as well as studying changes in architecture within biogenic carbonates 

e.g. critical point drying and SEM. 

Consumables: During the first year € is required for culturing equipment with € for 

analyses of pre-existing material: electron microprobe (EMP) ~€ /day, LA-ICP-MS ~€ /day 

(in house prices); additional costs for preparation and SEM analyses. 2 nd year costs will be for 

analytics (LA-ICP-MS, SEM, EMP) with chemical analysis of the culturing water (quadrupole 

ICP-MS, ~€ ). 3 rd year completion of LA-ICP-MS, SEM, EMP analyses. 

Travel: International conference attendance; work at Bremen (NanoSIMS and microsensors); 

Glasgow (for EBSD analyses and meetings); SYXRF at Hamburg and attendance at BIOACID 

workshops. 


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9:1215-1218 

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Fietzke J, Liebetrau V, Günther D, Gürs K, Hametner K, Zumholz K, Hansteen TH and Eisenhauer A (accepted) An alternative data 

acquisition and evaluation strategy for improved isotope ratio precision using LA-MC-ICP-MS applied for stable and radiogenic strontium 

isotopes in carbonates. J An AtSpectrom 

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standard MC-ICP-MS. Geochem Geophys Geosys doi:10.1029/2006GC001243 

Fietzke J, Eisenhauer A, Gussone N, Bock B, Liebetrau V, Nägler Th.F, Spero HJ, Bijma J, and Dullo C (2004) Direct measurement of 

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Ca/ Ca ratios by MC-ICP-MS using the cool plasma technique. Chem Geol 206:11-20 

Foster LC, Allison N, Finch AA, Andersson, C (in review) Strontium distribution in the shell of the aragonite bivalve Arctica islandica. 

Geochem Geophys Geosys 

Foster LC, Allison N, Finch AA, Andersson C, Clarke LJ (2008a) Mg in aragonitic bivalve shells: seasonal variations and mode of 

incorporation in Arctica islandica. Chem Geol, accepted 

Foster LC, Andersson C, Høie H, Allison N, Finch AA, Johansen T (2008b) Effects of micromilling on δ 18 O in biogenic aragonite. 

Geochemistry Geophysics Geosystems doi:10.1029/2007GC001911. 

Genin A, Jaffe JS, Reef R, Richter C, Franks PJS (2005) Swimming against the flow: a mechanism of zooplankton aggregation. Science 308: 

860-862 

Halfar J, Steneck R, Schöne B, Moore GWK, Joachimski M, Kronz A, Fietzke J, Estes J (2007) Coralline alga reveals first marine record of 

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Hansteen TH, Sachs PM, Lechtenberg F (2000) Synchrotron-XRF microprobe analysis of silicate reference standards using fundamentalparameter 

quantification. Euro J Mineral, 12:25-31 

Heinemann A, Fietzke J, Eisenhauer A, Zumholz K (2008) Modification of Ca isotope and trace metal composition of the major matrices 

involved in shell formation of Mytilus edulis. Geochem Geophys Geosys doi:10.1029/2007GC001777 

Hoegh-Guldberg, O et al. (2007) Coral reefs under rapid climate change and ocean acidification. Science 318:1737-1742 

Jackson CR (2004) An Atlas of Internal Solitary-like Waves and their Properties, 2nd Edition. Office of Naval Research, Global Ocean 

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Kamenos NA, Cusack M, Moore PG (2008) Coralline algae are global paleothermometers with bi-weekly resolution. Geochim cosmochim 

acta 72:771-779 

Kleypas JA et al. (2006) Impacts of ocean acidification on coral reefs and other marine calcifiers: a guide for future research. 88 p. Boulder, 

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Mumby, PJ et al. (2006) Fishing, trophic cascades, and the process of grazing on coral reefs. Science 311:98-101 

Orr JC et al. (2005) Anthropogenic ocean acidification over the twenty-first century and its impact on calcifying organisms. Nature 437:681- 

686 

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Project 3.3: Ultra-structural changes and trace element / isotope partitioning in 

calcifying organisms (foraminifera, corals) 

(Jelle Bijma (PI und co-PI 3.3.1), Anton Eisenhauer (co-PI 3.3.2)) 

i. Objectives 

For uni-cellular organisms like foraminifera, it has been shown that the carbonate chemistry of 

the ocean not only affects the shell weight (Bijma et al., 2002) but also shell chemistry (e.g. 

Bijma et al., 1999). Similar observations have been made for multi-cellular organisms like reef 

building corals. The similarity in the ultra-structures of different calcifying organisms suggests a 

common ancestral calcification mechanism. Here we propose to investigate the links between the 

calcifying mechanism, the ultra-structure and the chemical composition. One group of the project 

(3.3.1: co-PI J. Bijma) will investigate the ultra-structure of foraminifera and corals cultured 

under different pCO2 conditions, whereas the second group (3.3.2: co-PI A. Eisenhauer) will 

focus on trace element and isotope partitioning. In addition, the physico-chemical environment 

within the cell will be determined to identify physiological processes that control the ultra 

structure as well as the chemical and isotopical composition of the biogenic carbonate. The 

overall aim of this study is to further develop a process based understanding of biocalcification 

for uni- (foraminifer) and multicellular (corals) organisms, which would allow to predict the 

consequences of ocean acidification on marine calcifying organisms and how this affects the 

structural and chemical properties of their skeletons. 


To understand the trace element partitioning and isotope fractionation in foraminiferal tests and 

corals, used for paleo-climate reconstruction, extensive research on the calcifying mechanism 

itself was performed during the last decades. For foraminifers this investigation led to a model 

which builds on the following major mechanisms. Seawater is taken up by the organism 

(endocytosis) and transported to the site of calcification. During this transport the composition of 

the solution inside the vesicle is modified (concentrating Ca 2+ while possibly removing other 

divalent cations (notably Mg 2+ ) and at the same time pumping H + to “acidic” vesicles, thereby 

increasing pH and CO3 2- in the “calcification” vesicles). This explains, on the one hand, why 

foraminifers are among the best recorders of paleo-proxies (because calcification is based on 

ambient seawater). On the other hand, but by the same token, it also explains why foraminifera 

are so sensitive to ocean acidification. A similar model has been developed for corals (Erez, 

personal com.). Physiological responses within the foraminiferal and coral calcification pathways 

will determine their success as calcifiers in an acidified ocean. These responses, however, are 

completely unknown and subject of the current proposal. 

Uni- as well as multi-cellular marine calcifying organisms maintain their own distinct trace metal 

homeostasis which results in characteristic and species specific elemental ratios as well as in 

peculiar isotope fractionation (“vital effect”) which is significantly different from any inorganicthermodynamic 

expectations. This “vital effect” is most likely due to the development of 

biochemical mechanisms to keep the trace metal homeostasis of the most important divalent 

cations, magnesium (Mg) and calcium (Ca), and other trace elements, e.g. strontium (Sr) in 

narrow limits in order to meet certain physiological needs. In particular, the uptake of Ca into the 

cytoplasm is strongly limited due to its cell-poisoning effect. Furthermore, uni- and multi-cellular 

calcifying organisms apply biochemical mechanisms in order to actively lower their Mg 

concentrations and the Mg/Ca ratio in cell vesicles below a certain threshold before calcification 

starts in order to precipitate calcite instead of aragonite. 

183

184 


In this regard, the function of Ca 2+ -selective channels and Ca 2+ -ATPases embedded in the cellular 

membranes have been recognized to be very important gateways to control trace metal fluxes 

from an outside solution via the cytoplasm to the site of calcification. However, information 

about their function for the transport of trace metals from seawater to the site of calcification is 

limited. In particular, the partitioning of trace metals between seawater and biogenic CaCO3 of 

the major divalent cation fluxes (e.g. Ca, Mg, Sr) and their dependency on external 

environmental factors like seawater pH, salinity and water temperature has remained unexplored. 

In order to overcome this lack of knowledge on the function of channels and ATPases we 

propose to study divalent cation (Ca, Mg, Sr) partitioning between seawater and coral aragonite 

as well as foraminiferal calcite as a function of those important environmental parameters in the 

marine environment expected to change as a function of global increase in pCO2. The results of 

this study will provide a better understanding of the calcification mechanisms as well as provide 

quantitative information on trace metal ratios and isotope fractionation as a function of pH, 

salinity and temperature. This will put distinct quantitative constraints on the modeling of the role 

of ion selective channels and ATPases for the trace metal fluxes across membranes and the 

marine calcification mechanisms. 


Jelle Bijma established the interdisciplinary section Marine Biogeosciences at the AWI. He 

holds a PhD degree in Marine Biology and Actuo-micropaleontology. Since almost 25 years he 

and his group work on biological and paleoceanographic aspects of calcifying organisms. He is a 

specialist in foraminiferal ecology, stable isotope and trace element geochemistry and 

paleoceanography. His particular strengths for this project are his experience in interdisciplinary 

work and expertise in foraminiferal calcification and its stable isotope and trace element 

geochemistry. 

Anton Eisenhauer is a physicist and has a long scientific track record in the field of lowtemperature 

and isotope geochemistry and their application to reconstruct past and present 

environmental conditions in the marine realm. The proponent is the head of the isotope 

geochemistry laboratory of the IFM-GEOMAR which is among the leading facilities for the 

analysis of traditional and non-traditional isotope systems (e.g. δ 25 Mg, δ 44/40 Ca, δ 88/86 Sr, δ 11 B). 

The proponent manages the IFM-GEOMAR laboratories which are equipped with state-of-the-art 

mass-spectrometers and laser-ablation systems. The proponent has initiated studies on inorganic 

precipitation experiments and biomineralization studies to examine the partitioning between the 

bulk solution and the CaCO3. 


The subproject 3.3.1 addresses the ultra-structure of the shells (foraminifera, pteropods, 

ostracods) or skeletons (corals). Specifically, it focuses on the impact of the carbonate chemistry 

on the calcifying mechanisms by investigating the resulting changes in the ultra-structure as well 

as by following the fate of the “calcifying vesicles” using fluorescent probes. 

The subproject 3.3.2 focuses on the function of ion selective channels and pumps by measuring 

trace element and isotope partitioning between artificially altered seawater and the CaCO3 

skeleton precipitated by selected marine calcifying organisms. During the course of the project 

trace element partitioning and isotope fractionation will be studied by applying Ussing chamber 

experiments. These kinds of experiments allow to study the role of Ca-activated channels and 

pumps for trace element partitioning and isotope fractionation.


Joint culture experiments (3.3.1 & 3.3.2) 

Planktonic foraminifera 

The basis for our investigations are culture experiments under controlled laboratory conditions. 

Our groups have routinely carried out such experiments for many years. We use established 

procedures for maintaining planktonic foraminifera in laboratory culture. Scuba divers 

hand-collect live specimens of planktonic foraminifera from water depths of 2 to 6 m. Specimens 

are brought back to the laboratory, where they were identified, measured with an inverted 

microscope, and transferred to 0.8 µm filtered sea water in 115 ml culture jars. The jars are 

sealed without an air space and placed into thermostated water tanks maintained at constant 

temperatures (three groups: 23, 26 and 29°C). 

Illumination is provided by F24T12/CW/HO fluorescent bulbs on a 12:12 hr light:dark cycle. 

Symbiont bearing species are maintained under high light (“HL”, >380 µmol m -2 s -1 ), which 

corresponds to maximum symbiont photosynthetic rates (Pmax) , and low light (“LL”, 20-30 

µmol m -2 s -1 ), which is below the compensation light level . For comparison, ambient field light 

levels are >2000 µmol m -2 s -1 . The foraminifera are fed one brine shrimp nauplius every other day 

until gametogenesis. Empty shells are then rinsed in purified water and archived in covered slides 

for later analysis. The carbonate chemistry of the culture water is changed by keeping total 

alkalinity, DIC or pH constant. 

Corals and benthic foraminifera 

Scleractinian and soft corals as well as benthic foraminifera will be cultured under controlled 

laboratory conditions. In order to simulate global change on laboratory scale scleractinian and 

soft corals (e.g. Pavona clavus) as well as benthic foraminifera will be cultured as a function of 

selected pH (equivalent to atmospheric pCO2 of 180, 280, 380 and 700 ppmv), salinity (between 

33 to 40) and temperature (benthic: 5, 10 and 15°C; corals: 23, 26 and 29°C ). To provide enough 

material for analysis two field seasons (each lasting 3 to 6 months) of culturing at the Hebrew 

University of Jerusalem (corals) and the Inter-university Institute (IUI) at Elat, Israel (e.g. 

planktonic foraminfera and corals) are scheduled. In addition, the culturing facilities of IFM- 

GEOMAR (e.g. soft corals and benthic foraminifera) and of the AWI (ostracods and benthic 

foraminifera) will also be used in the framework of this study. 

185

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3.3.1 Impact of ocean acidification on the calcification mechanisms in marine calcifying 

organisms and on ultra structural changes of biogenic calcite 

Ultra structure 

We will investigate the impact of ocean acidification on biogenic calcification, where nucleation 

on organic membranes is a key process (Fig. 3.7). 

Fig. 3.7: SEM micrographs. The left micrograph is showing the polished cross section of a foraminifer embedded into 

a resin. The right micrograph is showing the close-up of the test wall. A special etching technique reveals structural 

details, like the presence of layers, interpreted as organic linings. 

By using AFM (Fig. 3.8) we will investigate how the test wall ultra-structure of (hyaline) 

foraminifera (build up of units only a few nm in size, surrounded by organic layers) is affected by 

changes in the carbonate chemistry of the culture medium. Such structures have also been 

demonstrated in bivalves and corals. This consistency may point towards a universal mechanism 

of calcification among these groups. Therefore we will investigate the ultra-structure in other 

groups of calcifying organisms (pteropods, ostracods). High resolution AFM (i.e. below 1 nm) 

will also be used to investigate ultra-structural changes of the calcifying units due to growth in 

different carbonate chemistries. 

Fig. 3.8: left: AFM scan of the three prominent lines shown in Figure 3.8 (right micrograph). Right: 500 nm x 500 nm 

AFM scan, done between two of the lines shown in the left figure.


In addition, AFM force curves on the “organic membranes” will be performed to determine 

functional groups active in biomineralisation. This will be complemented by inorganic calcite 

growth experiments in AFM fluid cells which will allow direct monitoring of crystal growth in 

the presence of organic molecules identified in foraminiferal tests. In this way, mimicking 

processes assumed during the growth of biogenic CaCO3 will provide new insights into controls 

on calcification and its relation to sea water chemistry that cannot be observed in vivo. 

Vacuolisation and calcification 

High-resolution fluorescence microscopy and con-focal laser microscopy will allow us to follow 

the complete pathway from sea water vacuolization until calcification (Fig. 3.9). The incubations 

will be extended by combining HPTS-incubations with other fluorescent probes to investigate the 

role of organelles and analyze the contents of the high-pH vesicles (like Ca 2+ ). 

Fig. 3.9: Recently, the fluorescent probes 

HPTS and Fluo-3AM have been applied to 

foraminifera and can now be used to visualize 

foraminiferal intracellular pH and Ca 2+ 

respectively. Results with HPTS dissolved in 

natural seawater (pH=8.2) indicate that before 

a new chamber is formed in C. lobatulus, 

vesicles with an elevated pH are formed in the 

outer chambers and migrate towards the site of 

calcification, where they produce a zone of 

high (>9.0) pH in which CaCO3 precipitates. 

Investigating the pH change along the calcification pathway and at the site of calcification is of 

vital importance to understand the physiological response of foraminifera to ocean acidification. 

To do so, foraminifera will be cultured under a range of different sea water pH's with dissolved 

fluorescent probes. At different stages during calcification, the intracellular pH will be 

determined. 

3.3.2 The effect of decreasing pH, salinity and temperature on the trace element 

partitioning between marine calcifying organisms and seawater. 

Based on existing models on the function of ion selective channels (c.f (Gussone, Eisenhauer, et 

al., 2003)) and pumps we will approach the problem by the measurement of trace element (Mg, 

Ca, Sr) and isotope (δ 25 Mg, δ 44/40 Ca, δ 88/86 Sr, δ 11 B, δ 13 C, δ 18 O) partitioning between artificially 

altered seawater and the CaCO3 skeleton precipitated by selected marine calcifying organisms 

(scleractinian and soft corals, benthic foraminifera). According to existing models pH variations 

should be reflected by the strength of the isotope fractionation process. These variations will then 

help to improve our quantitative understanding of biocalcification as well as the impact of pH 

variations on the rate of calcification. 

Trace element and isotope analysis 

All trace element (Mg/Ca, Sr/Ca, B/Ca, etc.) and isotope (δ 25 Mg, δ 44/40 Ca, δ 88/86 Sr, δ 11 B, δ 13 C, 

δ 18 O) measurements will be performed at the mass-spectrometer facilities of the IFM-GEOMAR, 

Kiel, Germany, using state-of-the-art mass-spectrometer technique (c.f. Heuser, Eisenhauer et al, 

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2002). Beside the Mg-, Ca-, and Sr-isotopes the boron isotope systematic (δ 11 B and B/Ca) will 

also be monitored due to its sensitivity to pH- and CO3 2- -concentration. 

Spatial resolution and shell inhomogenities 

In order to test spatial trace element and isotope inhomogenities we will apply the Laser-Ablation 

system at the IFM-GEOMAR which allows a spatial resolution of about 10 µm and even better. 

The Laser-Ablation system is designed to directly inject the sputtered material in the MC-ICPMS 

for in situ determination of trace element concentrations and isotope fractionation. 

Identification of transport enzymes 

The identification of the Ca 2+ -binding enzymes (cf. Fig. 3.10) will be performed in close 

collaboration with Prof. M. Bleich (Physiologisches Institut, CAU, Kiel). Furthermore, in order 

to test the influence of Ca-selected channels and pumps on Ca isotope fractionation we will 

perform experiments by using Ussing chambers. These kinds of experiments will be done in 

close collaboration with M. Bleich and his staff members. 

Links to other subprojects 

There are several links to other BIOACID projects. A major link will be to the subproject 0.4 

concerning “Training and transfer of know-how”. Together with U. Riebesell and M. Meyerhöfer 

(subproject 0.4) and J. Fietzke (3.2.4) we will organize a workshop on MC-ICP-MS-, TIMSmeasurements 

and isotope fractionation during biomineralization. Such an event is important in 

order to inform interested BIOACID scientists about instrumental and analytical progress. 

There are close links concerning the study of the calcification mechanisms to subproject 2.1.3 

which is focused on calcifying processes in macroorganisms and their relationship to changing 

environmental conditions in the ocean. Further links exist to project 2.3.1 which will examine the 

effects of changing ocean conditions on the otolith/statolith formation. Latter approach is also 

directly relevant to our study. 

Direct links exist to the projects 3.1.2, 3.1.3 and 3.1.4 which focus on calcification processes in 

cold water corals, molluscs as well as on the membrane transport mechanisms for calcification 

and pH-regulation. 

Further links concerning the application of trace element and isotope partitioning effects exist 

with subproject 3.2.1 (U. Riebesell). In collaboration with this subproject we will examine the 

effect of ocean acidification and warming on pteropods from subpolar regions.


Fig. 3.10: ‘‘Epithelial’’ model of Mg 2+ removal from the privileged space in perforate foraminifera. The parent 

solution is based on seawater which arrives to the privileged space via vacuoles from the apical side. From the 

privileged space Mg 2+ diffuses into the cell via Mg 2+ channels (bold arrow); the diffusion is favored by both the 

concentration gradient and the membrane potential (_40 mV). In the cell, the free Mg 2+ is buffered by binding to 

negatively charged cytosolic molecules, mainly ATP, and by sequestration into cellular compartments such as 

mitochondria and endoplasmic reticulum (ER). Simultaneously, active extrusion of the excess Mg 2+ by ion exchangers 

and pumps is exerted at the apical side. Note that all the above Mg 2+ transport processes may also be employed on the 

seawater vacuoles during their intracellular pathway to the calcification site (from Bentov and Erez, 2006). 

Schedules 3.3.1 and 3.3.2 


Literature study, preparation field 

season 

Set-up of culturing facility; first CO2 perturbation experiment 

Sample processing and 

measurements: AFM + 

(convocal)microscopy 

Set-up of culturing facility; second 

CO2 perturbation experiment 

Sample processing and 

measurements: AFM + 

(convocal)microscopy 



Manuscript preparation, presentation 

of results at conferences 


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- 1st experiment successfully carried out month 09 

- 1st experimental data set on CO2/pH sensitivity (180 and 380 ppmv) month 18 

- 2nd experiment successfully carried out month 21 

- 2nd experimental data set on CO2/pH sensitivity (280 and 700 ppmv) month 30 

- Evaluation of combined data set; Sensitivities and uncertainties month 33 


Literature study, preparation field 

season 

Set-up of culturing facility; first CO2 perturbation experiment 

Sample processing and measurements: 

TIMS MC-ICPMS 

Set-up of culturing facility; second 

CO2 perturbation experiment 

Sample processing and measurements: 

TIMS MC-ICPMS 







- 1st experiment successfully carried out month 09 

- 1st experimental data set on CO2/pH sensitivity (180 and 380 ppmv) month 18 

- 2nd experiment successfully carried out month 21 

- 2nd experimental data set on CO2/pH sensitivity (280 and 700 ppmv) month 30 

- Evaluation of combined data set; Sensitivities and uncertainties month 33 



3.3.1 

3.3.2 

Subtotal 


Consumables 

3.3.1 

3.3.2 

Subtotal 

Travel 

3.3.1 

3.3.2 

Subtotal 

Other costs 

3.3.1 

3.3.2 

Subtotal 

Investments 

3.3.1 

3.3.2 

Subtotal 

TOTAL 


3.3.1 



Personnel costs: A post doctoral position for 30 months (6 months financed via different 

funding). We request a post doc for this project because the scheduled scientific program is much 

too ambitious for a Ph.D.-student. 

Consumables: microelectrodes, fluorescent probes, AFM tips, chemicals, glasware, etc. for 

experiments 

Travel: travel for PI and post doc to conferences and to Elat for fieldwork. 

Other costs: The total costs of carrying out fieldwork in Elat will amount to about k€ per field 

season (bench-fees, use of boat and divers for collection, accommodation and subsistence). The 

AWI contribution to this is € k€. 

3.3.2 

Personnel costs: The personnel costs for the project comprise 1.5 post-doc years. We request a 

post doc for this project because the scheduled scientific program is much too ambitious for a 

Ph.D.-student and request an experienced post-doc. We intend to hire Dr. Isabell Taubner who 

has already started to perform culturing of the requested sample material in the frame of an 

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excellence cluster project in Israel. Part of the sample material for the intended Bioacid study will 

be available already end of 2008 when the cluster project is terminated. Further material will then 

be cultured in the frame of this project Although the intended 18 post-doc months in the 

subproject 3.3.2 will not cover the whole project time the continuity in data interpretation and 

networking with other Bioacid projects is further guaranteed by the project Co-PI A. Eisenhauer. 

Consumables: The costs for consumables cover the expenses for lab rent in Eilat (in 

collaboration with Prof J. Erez, HUJI) and Jerusalem as well as the expenses for chemicals, lab 

ware for both laboratories and transportation from Israel to Germany. Furthermore, costs for 

chemical preparation of samples and for the mass-spectrometer measurements at the massspectrometer 

facilities of IFM-GEOMAR by applying state-of-the-art equipment (Laser-MC- 

ICP-MS, TIMS, MC-ICP-MS, etc) will also be covered by the consumables. 

Travel: Culturing experiments will be performed in the first project year in Israel. The requested 

travel costs cover the transfer of personal from Germany to Israel and will also cover the daily 

expenses for the post-doc during his stay in Israel. The travel costs will also cover expenses for 

renting a flat to save hotel costs. Furthermore, the costs will also cover the costs for the 

occasional renting of a car to transport Gulf of Elat water from Elat to Jerusalem for the culturing 

experiments. 


Bijma JH, Spero J, et al. (1999) Reassessing foraminiferal stable isotope geochemistry: Impact of the oceanic carbonate system (experimental 

results). Use of Proxies in Paleoceanography: Examples from the South Atlantic. G. Fischer and G. Wefer. Berlin, Heidelberg, Springer- 

Verlag: 489-512. 

Bijma JH, Hönisch B et al. (2002) Impact of the ocean carbonate chemistry on living foraminiferal shell weight: Comment on "Carbonate ion 

concentration in glacial-age deep waters of the Caribbean Sea" by W. S. Broecker and E. Clark - art. no. 1064." Geochemistry Geophysics 

Geosystems 3: 1064-1064. 

Bentov S, Erez J (2006) Impact of biomineralization processes on the Mg content of foraminiferal shells: A biological perspective. Geochem. 

Geophys. Geosyst. 7(Q01P08). 

Griffith EM, Paytan A, Kozdon R, Eisenhauer A, Ravelo AC (2008) Influences on the fractionation of calcium isotopes in planktonic 

foraminifera, Earth Planet Sci Lett., 268, 124-136. 

Gussone N, Eisenhauer A et al. (2003) Model for Kinetic Effects on Calcium Isotope Fractionation (δ 44 Ca) in Inorganic Aragonite and 

Cultured Planktonic Foraminifera. Geochim Cosmochim Acta 67(7), 1375-1382. 

Heuser A, Eisenhauer A et al. (2002) Measurement of Calcium Isotopes (δ 44 Ca) Using a Multicollector TIMS Technique. International Journal 

of Mass Spectrometry 220, 385-397.


Project 3.4 Microenvironmentally controlled (de-)calcification mechanisms 

(PI: Michael Böttcher,) 

i. Objectives: 

Our plan is to determine the effect of changing seawater pH and the dissolved carbonate species 

on calcification and decalcification in marine sediments and microbial mats. In these matrices 

microbial activity and mass transfer resistances control the pore water chemistry and thus the 

microenvironments where calcification and decalcification occurs. One hypothesis we will test is 

that the microenvironment in the top mm of such systems is buffered against pH and CO2 

changes, due to mass transfer resistances. Secondly, we will investigate in how far sediment 

processes can buffer the pH in the water column, depending on exchange rates and the reactivity 

of the biogenic carbonates. We will further investigate how the water column chemistry of the 

Wadden Sea reflects exchange with the North Sea and surface sediments. 

ii. State of the Art: 

Effect of ocean acidification on benthic calcification 

Whereas much research is done towards calcium cycling in ‘classical’ calcifying marine systems 

such as corals, foraminifera and coccolithophores, and much progress towards its understanding 

has been obtained, much less is done on calcification driven by photosynthesising 

microorganisms. Examples of the latter are calcification in stromatolites and tufas, calcareous 

sediments, and beachrock (Krumbein, 1979). The fundamental difference with the main marine 

calcifiers is that calcification is a side process of metabolic activity, rather then a well controlled 

process leading to species-specific structures, such as shells and skeletons. Essentially, 

photosynthetic CO2 fixation leads to a shift in the pH, increasing the oversaturation of CaCO3. 

This may be a globally important process for all shallow coastal areas with sediments in the 

photic zone. Microbially driven calcification in reef sediments was estimated to be of the same 

order of magnitude as by corals (Werner et al., 2007), calcified structures developing in hardwater 

creeks, tufas, were shown to be formed by photosynthesis (Bissett et al., 2007), and also in 

stromatolites and hypersaline mats microbial photosynthesis was shown the driving force for 

calcification (Ludwig et al., 2005). Degradation of EPS by sulphate reduction is thought to play a 

role in calcification in stromatolites and cyanobacterial mats (Arp, Reimer & Reitner, 1999; 

Visscher et al., 1998). It is thought that microbial degradation of these calcium ion binding 

polymers plays an initiating and structuring role in calcification. Calcification in benthic 

communities can be studied by direct budgeting methods, including tracer studies, using 

radioactive 45 Ca 2+ or 14CO3 2- (Al-Horani et al., 2005), alkalinity shifts (Gattuso, Allemand & 

Michel, 1999; Gattuso et al., 1996), Ca 2+ fluxes, the general characterization of the calcifying 

sites and driving microbial processes can be measured by microsensors (Bissett et al., 2007; de 

Beer et al., 2000; Werner et al., 2007; Wieland et al., 2001). 

Dissolution of carbonates in sediments of the North Sea and the Southern Ocean 

Decalcification leads to a pH increase and thus can buffer the effects of increasing CO2. The 

marine biogeochemical carbon cycle is controlled by biological processes that influence the 

distribution of carbon in the surface sediments of the Ocean and exchange processes between 

sediments and water column and by exchange processes with the tidal coastal areas (Brasse et 

al.1999; Kempe & Pegler, 1991; Thomas et al., 2007). After the death of organisms with biogenic 

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carbonate shells, part of this material arrives on the sea floor where it is decayed. Biogenic shells 

are an important reservoir of bound oxidized carbon. The periostracum layer of shells enhances 

the availability of electron donors for remineralizing epibiontes that may enhance carbonate 

dissolution (Knauth-Köhler et al. 1996). The geographic distribution of biogenic carbonates is the 

result of synergistic thermodynamic and kinetic interaction processes with aqueous solutions 

(Wollast 1994), controlling the degree of carbonate preservation in surface sediments. Higher 

solubility of CaCO3 in cold water regions leads to a low preservation potential in temperate and 

cold climate zones (Morse and Mackenzie, 1990; Wefer et al., 1987). Alexandersson (1979) was 

able to demonstrate the decay of carbonate shells in Skagerrak surface sediments. Most of the 

North Sea regions are living grounds for a large number of carbonate-forming species, mainly 

bivalve molluscs with an epibiontic (Mytilus edulis, Crassostrea gigas) or endobiontic life mode 

(Cerastoderma edule, Mya arenaria, Ensis ensis and Macoma baltica). In addition, eroded 

terrestrial carbonates form part of the carbonate pool in North Sea surface sediments that may 

interact with the pelagic dissolved carbonate system. Decreasing pH and increasing carbon 

dioxide partial pressure will increase the solubility and hence will affect the formation and 

dissolution rates of the different CaCO3 fractions. Although the influence of bacterial activity on 

carbonate precipitation has been demonstrated in tropical areas, less is known on the influence of 

biological (microbial) activity on the degradation processes in shallow water areas of the 

temperate climate zone (e.g. Aller 1982; Golubic & Schneider, 1979; Green and Aller 1998; 

Krumbein, 1979). Therefore, these processes are an as yet quantitatively unknown factor in the 

carbon cycle of coastal areas. The North Sea carbonate system is closely related via exchange to 

the processes taking place in the intertidal areas and is already under influence of human activity 

and global climate change (Brasse et al., 1999; Thomas et al., 2007). 

In the high-latitude oceans the carbonate concentration is the lowest in the world because of the 

very low water temperatures which shift the chemical equilibria of the carbonate system towards 

low carbonate concentration. The saturation state with respect to aragonite and calcite in these 

waters is, therefore, the lowest, although oversaturation is still prevailing. Changes in the 

carbonate system due to acidification will have the highest impact here. These waters will in the 

future turn out to be most corrosive to aragonite and calcite (e,g. Orr et al., 2005). Ocean 

acidification will first affect the surface ocean, but the signal will subsequently be transferred to 

the subsurface and deep oceans. This will largely occur in the polar oceans, where new deep and 

bottom waters are formed, mainly at and near the continental shelves. The high-latitude oceans 

are also the conduit for the transfer of anthropogenic CO2 from the surface layer to the deep 

oceans. Low carbonate concentration and high uptake of anthropogenic CO2 constitute the most 

destructive combination with respect to ocean acidification. Calcareous sediments in the polar 

regions represent the initial spatial and temporal buffering of global ocean acidification – further 

buffering will occur on centennial time scales on the abyssal sea floor. It has long been known 

that Antarctic Bottom Water plays a substantial role in the dissolution of calcareous sediments 

(Berger, 1970). Although the Southern Ocean is more known for its arenaceous sediments, also 

significant portions of carbonate can be found (Hulth et al., 1997). Foraminiferal assemblages are 

found in the surface sediments of the Weddell Sea (e.g. Anderson, 1975). Also pteropods shells 

are preserved in the surface sediment muds. Pteropods have high abundances in the Southern 

Ocean, both in the open ocean and on the continental shelves (e.g. Accornero et al., 2003; Hunt 

et al., 2007). Pteropods produce aragonite, a metastable form of CaCO3 that is more soluble than 

calcite, making aragonite the more important buffering agent of ocean acidification. Berner and 

Honjo (1981) estimated that, as an example, aragonite constitutes at least 12% of the global 

CaCO3 flux.



3.4.1 Michael E. Böttcher (IOW) has studied the biogeochemistry and stable isotope 

geochemistry of carbon, sulfur and metals in the water column, in modern and ancient marine 

sediments, and ground water systems at different locations, including the intertidal and the 

pelagial of the open North Sea, the Baltic Sea, and carbonate karst systems (e.g., Böttcher, 1999; 

Böttcher et al., 2000, 2007; Dellwig et al., 2007). Additionally, a number of studies were carried 

out on the geochemistry of the experimental formation and destruction kinetics in temperate 

carbonate systems (Böttcher, 1997, a,b, 1999), and the structural characterization of biogenic 

carbonates (e.g. Böttcher et al., 1997). Since 2006 he is heading the Marine Geochemistry group 

at IOW. 

Gerd Liebezeit is working since 1977 on different chemical aspects of coastal regions and landocean 

interactions, since 1991 with emphasis on the the Wadden Sea. Research topics include 

a.o. dating of biogenic carbonates (Behrends et al., 2003, 2005), geochemistry of intertidal 

deposits (Hertweck and Liebezeit, 1996; Hertweck and Liebezeit, 2002; Hertweck et al., 2006) 

and phosphate uptake by intertidal biogenic carbonates, dynamics of the nutrient cycles as well as 

characterization and transformation of organic compounds. 

3.4.2 Dirk de Beer (MPI-MM, Microsensor Group) studies how transport controls microbial 

processes in sediments, biofilms and microbial mats. Microbial processes studied include 

photosynthesis, aerobic respiration, denitrification, nitrification, sulphate reduction and sulphide 

oxidation. He has over 130 peer reviewed publications. His special expertise is use and 

development of micro-environmental analyses by microsensors, combined with geochemistry and 

community studies. Calcification studies were done on calcification in corals, foraminifera, 

cyanobacterial mats, Halimeda, tufas, and reef sediments. 

Marcel Kuypers (MPI-MM, Nutrient Group) is a marine chemist who studies microbial controls 

of the global element cycling using stable isotopes. He has studied mechanisms and 

biogeochemical implications of global organic carbon burial. He has made break-throughs in the 

research of the global nitrogen cycles, particularly on the importance of the recently discovered 

ANAMMOX process. He has over 30 peer reviewed publications in the last 5 years. He is 

responsible for the nanoSIMS, that is recently installed in our institute. 

3.4.3 Mario Hoppema (AWI) has been active in Antarctic research since 1992, where the 

biogeochemical cycle has been his main research interest (e.g. Hoppema, 2004; Hoppema and 

Anderson, 2007). Chemical data have been used to obtain biological parameters of the Weddell 

Sea (e.g. Hoppema et al., 2002; 2007). Within the framework of the EU project CarboOcean a 

carbon data synthesis is underway, scheduled to be finished in 2008. Hoppema is the leader for 

the Southern Ocean part of this synthesis 

Christoph Völker has been modelling the physics of the Southern Ocean (Olbers et al., 2004) 

and has developed a biogeochemical model, based on the MIT circulation model (Marshall et al., 

1997). He has studied the influence of phytoplankton community composition (Denman et al., 

2006) and phytoplankton physiology (Hohn et al., 2008) on elemental fluxes using global 

planktonic ecosystem models. He is currently involved in EU project CarboOcean dealing with 

physical and biological feedbacks of Southern Ocean carbon chemistry to climate change. 

Dieter Wolf-Gladrow has been research scientist at AWI since 1987 and since 1999 Professor of 

Theoretical Marine Ecology at the University of Bremen. He is an internationally recognized 

expert on all aspects of the marine carbonate system (e.g. Jansen et al., 2002) and wrote a much 

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used textbook on the CO2 system (Zeebe and Wolf-Gladrow, 2001). Ocean acidification already 

had drawn his interest at an early stage (Wolf-Gladrow et al., 1999). 


Subproject 3.4.1 Impact of biogenic carbonates on pH buffering in an acidifying coastal sea 

(North Sea) 

(M.E. Böttcher) 

We propose to investigate the influence of changing pH and carbon dioxide partial pressure, and 

alkalinity on carbonate dissolution (and production) in surface sediments of the Wadden Sea and 

the consequences for and relation to the carbonate system of the North Sea. Our hypothesis is that 

biogenic carbonates at and just below the surface of intertidal sediments play a role in modifying 

tidal waters that exchange with the shallow North Sea. This adds to carbonate fluxes caused by 

benthic organic matter degradation. The absolute and relative importances will change as the 

North Sea carbonate system will acidify in the future. 

One important aspect will be the reactivity (reaction rates and thermodynamic stability) of 

different biogenic carbonates (e.g., Mytilus, Crassostrea, Mya, Hydrobia, etc.) and the 

experimental quantification of the different dissolution rates in pure and seawater media under 

different controlled laboratory conditions, i.e. pH, partial pressure of carbon dioxide (pCO2-stat 

and free-drift), and carbonate undersaturation. The reactive surface of the carbonate will be 

characterized to extract specific dissolution rates from experimental data on the development of 

liberated major minor and trace cations and the dissolved carbonate species. Experiments will be 

carried out at different fixed CO2 partial pressures (as defined by the BIOACID consortium). This 

approach will include in parallel experiments geomicrobiological effects on microbial 

degradation of the organic matrix compared to purely abiotic reactions. 

Experimental laboratory-based approaches will be compared to field in-situ transformation 

experiments and will be followed by microscopic (e.g., SEM-EDX), inorganic geochemical (ICP- 

OES, photometry, microsensors) and stable isotope (C, O; irmMS) approaches. Experiments will 

also be carried out in collaboration with Dr. Hoppema (3.4.3) regarding benthic mesocosm 

experiments at AWI. Field work will additionally characterize the sources and surface textures of 

different carbonate fractions in surface sediments with geochemical methods and SEM-EDX. 

Thus we will evaluate the time-dependent corrosion of biogenic carbonates as a function of burial 

time. Application of microsensors to characterize the chemical gradients will be carried out in 

collaboration with Dr. D. de Beer (MPI-MM; 3.4.2). From the side of carbonate formation, the 

influence of a changed carbonate system on growth of Mytilus and Crassostrea is planned to be 

eventually assessed in a later phase of the project. 

Finally, pelagic measurements and experimental results will link the alkalinity and DIC exchange 

with the coastal North Sea and the possible ecosystem consequences via biogeochemical 

modelling. Parameters of the dissolved carbonate system (TA, CA, DIC, PCO2, pH) will be 

measured in collaboration with PD Dr. Bernd Schneider (IOW) while δ 13 C(DIC) will be 

determined via irmMS. These results will be provided for a collaborative integration in Theme 5 

(Pätsch et al.) and the inclusion in the modelling of the North Sea carbonate system. The 

approach provides the base for a quantitative understanding of the role of the carbonate system in 

the water column on preservation, destruction and temporal authigenesis in the intertidal surface 

sediments, and the role of benthic metabolisms.

Links to other subprojects 


There are links to subproject 1.2.5 regarding benthic carbonate-relevant processes and 2.1.3 and 

3.1.3 which will provide TP 3.4.1 with shell material grown under well-defined conditions for 

dissolution rate studies. Microsensor approaches in experimental dissolution studies will be used 

in collaboration with TP 3.4.2, and an experimental collaboration with TP 3.4.3 will make use of 

field aragonite in dissolution studies at IOW and benthic mesocosm experiments at AWI. 

A direct link exists to project 5.1.which focuses on the modelling of the carbonate system of the 

North Sea. TP 3.4.1 will provide field data for the carbonate system at the tidal-open North Sea 

boundary leading to a close link between experimental and modelling approaches. 

Schedule 


Measurements in the pelagic water 

column of the intertidal, in-situ 

experiments 

Carbonate system of the intertidal, 

transects and cycles 

Experimental dissolution of shells 

Evaluation of field data and calibration 

of future campaigns 

Mesocosm experiments with AWI 

(3.4.3) 

Growth experiments, Geomicrobiology 

and Modelling 

Writing of manuscripts presentation on 

conferences 



- Implementation of carbonate system data in model environment of theme 5 

(Pätsch et al.) 

month 9 and 24 

- Implementation of experimental setup month 12 

- Field cruises (transects) on the pelagic Intertidal month 27 

- Mesocosm experiments at low temperatures month 27 

- Reactivity of carbonates in the North Sea Intertidal month 33 

- PhD Thesis and publications month 36 

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Subproject 3.4.2 Benthic (de-)calcification driven by microbial processes 

(D. de Beer) 

Hypothesis: Whereas corals and coccolithophores are very sensitive to pH, microbially driven 

calcification is rather resilient against pH variations in the water column. This difference can be 

understood from the essential differences in calcification mechanism. Calcification by skeleton 

and shell building organisms is actively controlled by energy demanding active transport of H + 

and Ca +2 . A lower pH will make calcification more energy demanding, and thus make these 

calcifiers less competitive than non-calcifiers. Calcification in mats and sediments is a sideproces 

of photosynthesis, inducing a microenvironment controlled by the microbial activities and 

the transport resistance in the sediments. The transport barrier separates the benthos largely from 

seawater, and buffers the sediments against changes in the seawater chemistry. 

WP1 Experimental studies of calcification and decalcification 

We will investigate calcification in phototrophic communities by (1) budgeting the exchange of 

calcium between (de-)calcifiying sites and seawater and quantify the driving metabolic process 

rates, and by (2) studying incorporation of calcium carbonate in the calcite matrices, and (3) 

determine the effect of pH stress on the community structure and spatial distribution. 

1) We will determine the microenvironment in the benthic communities and the relevant fluxes 

by microsensors for pH, O2, Ca 2+ , CO2 and CO3 2- , and the rates of photosynthesis and respiration, 

by membrane inlet mass spectrometry (MIMS), photopigment fluorescence (PAM) and 

microsensor techniques. Calcification measurements in the field will be assessed using the eddy 

correlation technique (Berg et al., 2003), which is recently adapted to measure Ca 2+ fluxes. 

2) The incorporation of calcium and carbonate into the solid matrices will be studied by radio- 

and stable isotope techniques (by ß-imaging and nanoSIMS), to assess how closely calcification 

is associated with microbial cells (with sub-µm resolution). We will investigate, with Dr. A. 

Eisenhauer, trace element incorporation in response to environmental factors and calcification 

rates. 

3) We will determine the dynamics of phototrophic benthic communities in response to pH shifts 

by hyperspectral (HS) imaging, which allows quantitative assessment of the 2-D distribution of 

photopigments. By imaging their variable fluorescence we will assess whether they are part of an 

intact photosystem. HS-imaging is a non-destructive community analysis, and thus we can follow 

the short-term community dynamics in response to an environmental stress. 

Experiments will be done under a wide range of artificially imposed pH values, adjusted by CO2 

levels, in agreement with the strategy defined by the consortium. We will install mesocosms for 

incubations and measurements in the institute. Samples will be obtained from different habitats, 

from freshwater stromatolites (tufas), marine sediments and hypersaline mats. The tufas can be 

obtained from several creeks in the Harz (ca 200 km from the MPI), marine carbonate sediments 

from the Mediterranean and hypersaline mats are currently cultivated in the institute. The 

comparison of different salinity is interesting as the pK2 strongly decreases with salinity. Field 

measurements can well be done using a field station on Elba, from where we will also obtain 

carbonate sediments. The experimental work will be done by a PhD student. The student will also 

be active in teaching the microsensor course, with help of staff members from the microsensor 

group.


WP2 Biogeochemical modelling of calcification and decalcification 

The data will be used to construct and refine a transport-conversion model, that includes acidbase 

equilibria and calculates the pH and DIC profiles. An approach to build such a model was 

described by colleagues from the NIOO (Ierseke, Nl) and the Free University of Brussels (Dr. P. 

Meysman), with who we will collaborate. A second step will be to expand this model with 

calcium carbonate dissolution and precipitation reactions. Conceptually, how microbial and 

biogeochemical processes control calcification and decalcification are well known. Processes 

leading to acidification lead to dissolution and increased alkalinity can enhance precipitation. As 

for any heterogeneous (involving a water and solid phase) process, modelling of calcification 

needs to address the quantitative description of the microenvironment where the reaction 

(calcium carbonate precipitation or dissolution) occurs. A second step is to incorporate the local 

solid-liquid exchange, so between calcite matrices and porewater, depending on the local 

microenvironment. Both steps are highly challenging. The relevant microenvironmental 

parameters for calcium carbonate dissolution and precipitation include the calcium and carbonate 

concentrations, and thus the DIC and pH. Many biogeochemical processes influence this 

seemingly simple set of parameters, complicating the modeling. Moreover, the kinetics of 

calcium and carbonate exchange between the solid and water phase depends on a variety of 

poorly understood parameters, such as organic matrices associated with the precipitate, and the 

anion composition of the water phase. Thus meaningful modelling must be accompanied by 

measurements. 

A model of the carbonate system near photosynthesizing calcifying foraminifera was build 

several years ago (Wolf-Gladrow, Bijma & Zeebe, 1999), which calculated the local pH from 

respiration and photosynthesis in a spherical geometry. A comprehensive geochemical model 

should include a range of geochemical processes (Soetaert et al., 2007). We aim to construct such 

a more comprehensive biogeochemical model to calculate the local carbonate system and pH 

profiles. The problems to solve a system with many variables and an equal number of equations 

on processes with a wide variety of timescales are described in detail and solution methods are 

offered (Hofmann et al., 2008). Their approach is a series of local mass balances, and includes 

diffusional and/or advectional mass transfer. Such a model describes the gradients of the 

carbonate system in sediments and microbial mats, including the pH, but not yet the effects of the 

local microenvironment on the precipitation and dissolution of calcium carbonate. Parameters 

controlling these processes must be experimentally determined for each studied system. 

Obviously, calcification and decalcification again influence the local pH, and once known must 

be integrated into the model. The determination of these processes under different 

microenvironmental conditions is an important aim of our studies with microsensors, ß-imaging 

and nanoSIMS. The experiments can be done on a range of conditions, using the modelling a 

wide generalization of the phenomena observed will be achieved. Such a model will help 

defining the threshold values for net calcium dissolution versus precipitation in various benthic 

systems. 

Microsensor course 

We will offer a microsensor course for the participants of the Verbundprojekt. The course will be 

in two weeks. The first week making of microsensors for oxygen and various ions will be 

learned. The most practical teaching will be done by our expert team of technicians. The second 

week will be a training in various microsensor applications, under guidance of scientists. Here the 

participants are encouraged to bring their own samples, however, we have interesting sample 

material available. Hopefully, microsensors that are build in the first week may be used, whereas 

199

200 


microsensors build by our experts will be needed in addition. There will be daily seminars on the 

theory behind microsensor use and examples of complete studies, and also participants are 

invited to present their work. Each course will be for 8 participants maximal, we foresee the need 

for 2 courses 1 year separate. 

Collaborative microsensor experiments are planned with project 4.1.3 (Bischof ) 

Schedule 


Mesocosm experiments 

Tracerstudies incl. nanoSIMS 

ß-imaging 

modeling 

Field measurements 

Microsensor course 

Collaborative work 





- Implementation of experimental facility, and learning of basic experimental 

skills 

month 6 

- Experimental data set on CO2/pH sensitivity month 9 

- Field data set on diel calcium fluxes month 15 

- Model for benthic microprofiles of solutes and activities month 24 

- Expansion of model for calcium exchange month 36 

- Microsensor courses month 12 and 24 

- Experimental data sets on effect of salinity on CO2/pH sensitivity month 30 

- Thesis defense month 36 

3.4.3 Buffering ocean acidification: Dissolution of carbonate sediments in the Southern 

Ocean (M. Hoppema) 

We propose to examine the buffering capacity of Antarctic sediments, where our first focus is on 

the continental shelves. In particular, the role of pteropod aragonite in surface sediments in the 

buffering will be investigated. This shall be done in a two-pronged study, addressing issues that 

are related within the biogeochemical cycle of the ocean and the seafloor. The projection of the 

(future) decadal trend of acidification of Southern Ocean waters constitutes one thrust of our 

study. We shall determine the development of pH, pCO2 and the carbonate saturation index 

Omega based on both observations and modeling. Data both of ourselves and from the EU IP 

CarboOcean carbon synthesis will be used to obtain pH and pCO2 in waters from the Weddell 

Sea and environs, with emphasis on the shelves. In some regions, we may get a time series of


repeat occupations. This will allow us to calculate the degree of saturation for aragonite and 

calcite for the time series period. We will extend the observed time-series of anthropogenic 

carbon over the next 100 years, using a state-of-the-art global biogeochemical model that has 

been especially tuned to reproduce present-day observations of nutrients, biological activity and 

the carbonate system in the Southern Ocean, using our observations of acidification as additional 

initial constraints. We will run the model a) with a prescribed atmospheric pCO2 increase keeping 

the physical forcing (wind stress, heat flux) fixed, and b) also with changes in the physical 

forcing as obtained from coupled climate prediction runs, to separate the effect of changing 

climate from those of increasing atmospheric pCO2 on the CaCO3 saturation on the Antarctic 

shelf. This work will be done by a PhD student, strongly assisted by the PIs. As to the 

parameterization of calcification, pCO2 sensitivities and DOC cycling, experience and data will 

be shared with projects 1.3, 5.1 and 5.2. 

As to the second prong, we will address the rate of dissolution of carbonate on shelves in the 

Weddell Sea and Antarctic Ocean. The potential future acidification, determined from the other 

prong of our research, will serve as a realistic scenario, against which the dissolution of 

sedimentary carbonate will be assessed. Sediment samples of the Antarctic Ocean from the AWI 

core repository will be investigated with respect to composition. Of particular interest is the 

fraction of pteropod aragonite. The composition will be studied as a function of small-scale and 

large-scale environmental factors, like grain size, water depth, sedimentation rate, characteristics 

of the overlying water mass and current velocity. The maximum buffer capacity is obtained by 

the absolute content of carbonate in the sediment. Analyses will be done with Isotope Ratio Mass 

Spectrometry. As a further step, dissolution experiments with sediment samples and pteropod 

shells under various conditions (pH, pCO2, CaCO3 saturation state, water current) will be 

performed. Samples will be collected during a Polarstern cruise to the Southern Ocean using 

large box corer (GKG) or multiple corer (MUC). We will use Rhizon sampling of pore waters 

(Seeberg-Elverfeldt et al., 2005) and benthic flux chambers. Both microscopic and chemical 

methods will be applied, where part of the experiments will be performed in joint work with 

other sub-projects - a). Using micro-sensors of the MPI Bremen; Dr. de Beer, subproject 3.4.2, 

and b) With subproject 3.4.1 (Dr. Böttcher), aragonite dissolution experiments at IOW and 

benthic mesocosm work at AWI. This will render information on the dissolution rates. 

Additionally, with subproject 1.2.5 we envision cooperation with regard to pressure-adapted flux 

chambers. The pressure laboratories utilized for identification of turnover rates and fate of 

nutrients and carbon from aggregates in the benthic boundary layer permit to actively 

control/select erosion or deposition of sediments. This is achieved by pressure-adapted flux 

chambers where bottom stress together with water column turbulence mimics the phases of tidal 

cycles. Depending on the composition of the sediments and fluid introduced into the chamber, the 

buffering capacity of the sediment/pore water region can be obtained from fluid samples drawn in 

addition to the aggregate dynamics. Finally, within the Alfred Wegener Institute there is 

extensive experience with sediment work, while the core repository contains numerous cores 

from the Weddell Sea and Antarctic Ocean. Several sampling and measurement techniques are 

available within the geochemistry and geology departments. 

Eventually, we will estimate the spatio-temporal distribution of CaCO3 dissolution rates around 

Antarctica from the combination of the laboratory results with sediment distribution. This 

distribution will be implemented as bottom boundary condition for dissolved inorganic carbon 

and alkalinity in the biogeochemical model to investigate the propagation of the buffering signal 

into the ocean interior. Through the combination of observations, experiments, and simulations 

we will thus estimate the buffering effect of CaCO3 and its kinetics in Antarctic shelf sediments 

on a decadal to centennial time scale. 

201

Schedule 

202 



Reading, modelling of water column 

data 

Set up, preparation of experiments, 

calibrations 

Dissolution experiments on cores 

Polarstern cruise to Southern Ocean, on 

board experiments 

Analysis of results, computing 

Writing of manuscripts presentation on 

conferences 



- Future projection of acidification of Antarctic water column month 09 

- Implementation of experimental setup month 12 

- Data set of dissolution of sediments cores plus preliminary interpretation month 21 

- Polarstern cruise month 27 

- Data set of field data month 30 

- Thesis month 36 


Personnel 3.4.1costs 

3.4.1 

3.4.2 

3.4.3 

Subtotal 

Consumables 

3.4.1 

3.4.2 

3.4.3 

Subtotal 

Travel 

3.4.1 

3.4.2 

3.4.3 


Subtotal 

Investments 

3.4.1 Gas-mixing device for 

chemostat chamber 

pH-meter 

3.4.2 Microsensor 

equipment 

(micromanipulator, motor, 

data acquisition, cables) 

Subtotal 

Other costs 

3.4.1 Contract 

measurements, 

geomicrobiology and ship 

time for sampling the 

intertidal pelagial 

3.4.2 Microsensor course 

Subtotal 

TOTAL 

Budget Justification 

3.4.1 



Personnel costs: The experimental studies on the reactivity of biogenic carbonates and the 

carbonate system of the intertidal pelagial will be performed by a PhD student, paid according to 

German TVöD system. 

Consumables/p.a.: 

Filtration material, disposables € 

Pure gases and gas mixtures € 

p.a. chemicals, ultrapure acids, standards € 

Electrodes, cable € 

Glass ware, fittings, vales, plastic tubings € 

Pipettes € 

Nylon net (incubations) € 

Travel: BIOACID-meetings in year 1, 2 and 3 ( € each), 2 international scientific meetings 

in the last 2 years ( each), 2 cruises in years 2 and 3 ( each), regular visits with 

partners 

203

204 


Investment: Installation of a gas mixing (CO2-N2) device with fine-316ss scaling valves to 

ensure proper CO2-partial pressure equilibration. 

Other costs: For geomicrobiological work done by Prof. Dr. Wolfgang E. Krumbein (Biogeoma) 

and µ-CT measurements (Prof. Freiwald, Erlangen) we apply for financial support for analyses 

on the heterothrophic degradation of OM leading to CO2 production and in-situ destruction of 

carbonate shells and µ-CT measurements of initial and degraded carbonates. In addition, ship 

time for sampling the intertidal pelagial has to be paid to the University of Oldenburg with € 

per day. Prof. Krumbein will also be involved in the interpretation of geomicrobiological 

consequences of experimental results. 

3.4.2 

Personnel costs: The laboratory and field measurements will be performed by a PhD student, 

and the modelling by a PostDoc, both according to German tariff system (TVöD) 

Consumables: Disposables 

glass ware 

chemicals 

(incl. for microsensors) 

stable isotope tracers 

radiotracers 

Travel: Verbundmeeting in year 1, 2 and 3 ( ), 2 international scientific meetings in last 2 

years ( each), 2 field studies in year 2 and 3 ( each), regular visits with partners 

Investment: The student will need a dedicated equipment for microsensor measurements, 

consisting of a micromanipulator (Marzhäuser), a motorized stage (Faulhaber), a data-acquisition 

board (National Instruments), a variety of shielded cables and a laptop with special software 

(Labview, National Instruments), and special amplifiers for amperometric and potentiometric 

microsensors (custom build in MPI). 

Other costs: For the microsensor course we will have to supply additional microsensors, as 

many will be broken in the learning phase. As more sensors than normal will be broken, and the 

normal sensor production is interrupted, we will need to purchase sensors from a commercial 

company to maintain the research in the group (over 20 scientists depend on microsensors). One 

microsensor costs Euro, so we allow 2 sensors to be broken per participant, as conservative 

estimate. 

3.4.3 

Personal costs: Experimental work will be performed by a PhD student for three years, costs 

according to AWI table. Additional modelling and data work by PIs and AWI staff at no cost. 

Consumables: Laboratory measurements, glassware, disposables and chemicals. In the second 

year the Polarstern cruise brings more costs. 

Travel: BIOACID project meeting every year ( €). Two international conferences and 

workshops each year ( € each); in the second and third year more travelling to conferences 

for presenting results. Field studies, Polarstern cruise; meeting with project participants at 

laboratories.



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Project 3.5: Impact of present and past ocean acidification on metabolism, 

biomineralization and biodiversity of pelagic and neritic calcifiers 

(PI: Adrian Immenhauser) 

i. Objectives 

The main aim of this collaborative research project is a detailed assessment of the past and future 

performance (metabolism, biomineralization and biodiversity) of coastal/sessile and 

oceanic/planktonic calcifyers exposed to CO2-induced ocean acidification. For this purpose, we 

intend to combine observational (real-world) data from Cenozoic acidification events with such 

obtained from experimental (culturing) work. We will study the effects of decreasing seawater 

pH on groups that might potentially benefit (nannoplankton) and such that will most likely suffer 

(bivalves) from acidification (see discussion in Iglesias-Rodriguez et al., 2008). With the 

interdisciplinary approach applied here we will address the following six key questions: (1) How 

sensitive are coastal and planktonic calcifyers to changing seawater pH? (2) What are the pH 

threshold limits and are these limits universal or species dependent? (3) What is the impact of 

changing seawater pH on biodiversity? (4) What is the adaptational potential of coastal and open 

oceanic ecosystems in the time-scale of decennia to few centuries? (5) Are short-term (culturing) 

experiments with calcifying organisms suitable analogues for pH/CO2 conditions predicted for 

the end of the 21 st century? (6) To which degree are Cenozoic acidification events suitable 

analogues for predicted anthropogenic CO2 rise by 2100? 


The present knowledge of CO2-induced ocean acidification (OA) is mainly based on modelling 

and experimental (culturing) work providing valuable information on the response of specific, 

short-lived organism exposed to pH/CO2 conditions predicted for the end of the 21 st century (see 

discussion in Fabry et al., 2008; Caldeira and Wickett, 2005 and references therein). This 

approach, however, suffers from the inherent disadvantage related to the limited observational 

time window. This shortcoming seriously compromises a prediction of the longer-term effects on 

OA sensitive marine ecosystems and neglects the adaptational potential of natural systems. This 

becomes particularly important as recent work controversially suggests that some groups of 

calcifiers (mainly phytoplankton) might in fact benefit from increased CO2 partial pressure 

(Iglesias-Rodriguez et al., 2008). An attempt to solve some of these controversies is the 

investigation of past acidification events (Röhl et al., 2007; Pancost et al., 2007; and references 

therein). The degree to which these pre-anthropogenic OA events are suitable analogues for 

predicted scenarios in this century are, however, insufficiently understood (Zachos et al., 2001, 

Iglesias-Rodriguez et al., 2008). It thus comes as a surprise that previous work combining the 

strengths of past (observational) and future (experimental) effects of changing pH values on 

marine calcifyers is at best scarce. 

Perhaps the most prominent Cenozoic examples of transient climate perturbations are known 

from the Palaeocene and Eocene intervals. This period of Earth’s history is punctuated by at least 

four hyperthermals: the early Late Paleocene Event (58.4 Ma), the Paleocene-Eocene Thermal 

Maximum (PETM; 55 Ma), the Early Eocene ELMO Event (53.3 Ma), and the Early Eocene X- 

Event (52.5 Ma; e.g., Nicolo et al., 2007). Negative δ 13 C excursions of 1-5‰ imply a massive 

(ca. 2000 x 10 9 metric tons) input of 12 C enriched carbon into the ocean-atmosphere system 

probably in the form of methane. These events were marked by a general warming, at least 4x 

207

208 


current CO2 concentrations, a low seawater pH and a shallow CCD (e.g., Zachos et al., 2005). 

The rapid environmental changes are related to substantial changes in marine and continental 

biota. But what was the impact of these past events on oceanic calcifyers and what is the 

expected impact of anthropogenic CO2-induced OA by the end of the 21 th century? 

Nannoplankton (coccolithophores and some calcareous dinoflagellates) are among the most 

abundant plankton organisms in the world’s oceans, and due to their capability of carrying out 

both, photosynthesis and calcification, they have an impact on the organic and inorganic carbon 

pump. They have been identified as key organisms for studying the impact of ocean acidification 

on ocean ecosystems due to their debated response to elevated pCO2 concentrations (Iglesias- 

Rodriguez et al., 2008). Malformation and reduced calcite production per cell has been observed 

for coccolithophores in culture and mesocosm experiments (Riebesell et al., 2000; Engel et al., 

2006). However, so far a direct proof for a response of nannoplankton to ocean acidification in 

natural environments is lacking and short term disruptions of the carbonate system might not 

reflect the observed natural changes in an adequate manner. Calcification in coccolithophores 

may also be influenced by nutrient supply, temperature or salinity, and the effects of ocean 

acidification could have positive or negative feedback effects on the natural variability. During 

past ocean acidification events, coccolithophores as a group were capable to survive dramatic 

changes in the carbonate system (Raffi et al. 2005; Gibbs et al. 2006, Medlin et al. 2008). In 

comparison to other well studied plankton groups (e.g., foraminifera) research on nannoplankton 

ecology, morphometry and geochemistry was previously limited by time consuming counting, 

measuring and processing techniques. New technological developments are now available that 

allow to overcome these problems. 

Reef communities (corals, calcifying bivalves, macroalgae etc.) are one of the most OA sensitive 

ecosystems. Due to their diagenesis-sensitive aragonitic skeletons, however, corals are poor 

recorders of past acidification events. In contrast, sessile neritic bivalves (Mytilus, Artica etc.), 

and particularly calcitic ones, are well-established, high-resolution archives of past and present 

environmental parameters (Khim et al., 2000; Buick and Ivany, 2004; Immenhauser et al., 2005, 

Rexfort & Mutterlose, 2006). Previous work, however, was severely limited by the lack of 

stratigraphically continuous onshore sections providing data on pre-, syn- and post-OA events. 

Newly discovered Palaeocene/Eocene sections in Oman allow now for a detailed study of coastal 

section those are stratigraphically complete and hence allow for a time-resolved sampling of 

bivalve hardparts across the intervals of interest. 

Here, we propose to investigate OA sensitive sessile (bivalves) and nannoplanktonic 

(coccolithophores and calcareous dinoflagellates) calcifiers across these past punctuated 

acidification events and to compare results with cultured bivalves exposed to various pH, CO3 2 

and CO2(aq) levels. Following this approach it is intended to combine the strengths of controlled 

laboratory experiments with the biologically and geologically significant temporal element 

provided by the fossil shell material. Furthermore, the combination of data from sessile neritic 

organisms with such from open oceanic planktonic ones allows for a more holistic view of 

problems related to ocean acidification. 


3.5.1 (Immenhauser).- The Bochum research team has longstanding experience in the 

investigation of mollusc shell material as sensitive archives of global change (e.g., Immenhauser 

et al., 2002, 2005; Hippler et al., 2008 etc.) and the application of non-traditional isotope systems 

(δ 44 Ca, δ 26 Mg; e.g. Buhl et al., 2007). Previous collaborative research projects involve initiatives 

such as EUROCLIMATE as well as DFG and NOW financed projects involving cultured and


fossil shell material. The collaboration with Prof. Schmahl and Dr. Griesshaber at the LMU 

München further strengthens the research team by involving experts in shell material sciences 

(Griesshaber et al., 2007; Schmahl et al., 2008). 

3.5.2 (Mutterlose).- The applicant has a long going expertise in the study of calcareous 

nannofossils from both fossil and recent settings. Findings from a study of calcareous 

nannofossils across the PETM interval under discussion have been published recently 

(Mutterlose et al., 2007). These clearly show the following results: (1) Significant changes in the 

composition of the nannoplankton assemblages. (2) Occurrence of malformed taxa. (3) 

Extinction of various, deep dwelling taxa. These changes in the composition of the primary 

producers are thought to reflect an acidification of the ocean water during a period of extreme 

warmth and very high CO2 concentrations. The collaboration with Dr. P. Schulte from the 

University of Erlangen will add more specific expertise on sedimentological aspects. 

3.5.3 (Meier).- The proponents have longstanding experience in calcareous nannoplankton 

research. Their knowledge includes investigation of plankton samples, core top calibrations and 

reconstruction of palaeoecology and past carbonate fluxes. Recently they were trained to operate 

the automated coccolithophore recognition system SYRACO. The results from the analysis of 

coccolithophore morphometry and weight estimates within the ESF MERF project (Quaternary 

Marine Ecosystem Response to Fertilisation) show the influence of alkalinity and carbonate ion 

concentration on size and weight of coccoliths from the Mediterranean Sea. 


Subproject 3.5.1 Comparison of Cultured and Fossil Bivalve Geochemistry and Shell 

Ultrastucture 

(Immenhauser in collaboration with Schmahl, Griesshaber) 

Molluscs are sensitive organisms recording environmental change in their biomineralogy, shell 

geochemistry and shell morphology (Immenhauser et al., 2005). In addition, by forming highly 

time-resolved growth increments they represent one of the accurate archives of coastal neritic 

settings (e.g., Witbaard et al., 1994; Vander Putten et al., 2000). Culturing experiments of 

bivalves in tanks provide the opportunity to expose specimens to environmental conditions 

representative of an acidic ocean as predicted for the year 2100. Biomineralization (growth) rates 

and modes, metabolic activity, preproduction cycles and live span can be assessed under these 

circumstances. An obvious pitfall, however, is the short (months to at best years) observational 

window provided by such experimental setups. Because of this bias, important parameters such 

as adaptational patterns and the effects of longer term (decennia and more) exposure to stressed 

environments cannot be assessed adequately. In order to overcome these problems, we here 

propose a combined field and culturing approach using the arguably best ocean acidification 

analogue of more recent history of our planet: the thermal maximum at the Palaeocene-Eocene 

boundary (PETM). For this purpose, it is intended to link with project of T. Brey in the context of 

BioAcid providing cultured shell material exposed to variable and increasing levels of OA to us. 

This project proposed aims at investigating the shell ultrastucture and shell geochemistry of 

cultured bivalves kept under pre-OA conditions and to compare these findings with comparable 

data from fossil shell material obtained from neritic sections from Oman including the 

Paleocene/Eocene record of hypothermals and acidification events. 

The metabolic activity of bivalves influences the fractionation of both Mg and Ca isotopes 

incorporated in the bivalves shell (e.g., Immenhauser et al., 2005; Hippler et al., 2008). Under 

favourable environmental conditions, the shell isotopic fractionation is largely in concert with 

209

210 


seawater temperature and salinity fluctuations. The isotope fractionation–environment relation of 

shell calcite and aragonite is lost when the bivalve is exposed to a stressed environment. As a 

consequence, δ 26 Mg and δ 44 Ca isotope ratios are an indirect proxy for the environmental state 

(favourable versus stressed) in modern and fossil bivalves (Hippler et al., 2008). Furthermore, 

highly detailed investigations of the shell ultrastucture of bivalves (nm to µm) reveal specifically 

different patterns in the micro-scale arrangement of the crystallites that build bivalve shells under 

normal (aerobic) metabolism versus stressed (temperature, pH, salinity etc.) environments 

(Griesshaber et al., 2007). Detailed investigations of the shell ultrastucture using REM, FIB, 

TEM and EBSD, material properties and distribution of organic components (München and 

Bochum) are planned. With those two highly novel tools: geochemical analyses (δ 26 Mg, δ 44 Ca; 

geochemical laboratories RU Bochum) and shell ultrastucture (material sciences LMU 

München), the assessment of the performance and mineralization mode of fossil bivalves can be 

assessed and calibrated knowing the experimental conditions resulting from the culturing 

experiments by the twin project of T. Brey (Project 2.1.3). Additional collaboration with other 

groups involving bivalve shells will further strengthen this research 4.1.2 (Wahl). 

In overview, sub-project 3.5.1 intends to investigate the metabolic activity, shell geochemistry 

and shell ultra-structure of: 

(1) cultured mollusc shells (cultured in tanks under variable pH, CO3 2 and CO2(aq)) in the context 

of the twin-project by T. Brey; and 

(2) comparison of these geochemical and shell ultrastucture data from fossil bivalves representing 

pre-, syn- and post-OA during past Palaeocene and Eocene OA events. Here, the focus is on 

calcitic molluscs because of their stable shell mineralogy (as opposed to diagenetically instable 

coral aragonite), their expanded life time (10’s to 100’s of years), and their sessile, benthic 

habitat, making them valuable analogues for coastal calcifying ecosystems; and 

(3) to link results with data from twin-subprojects 3.5.2 and 3.5.3. 

Schedule 


Fine-tuning of analytical facilities, 

(München/Bochum) 

Fieldwork in Oman (collection of OArelevant 

shell material) 

CO 2 perturbation culturing experiments 

in Bremen (project T. Brey) 

Analytical work in Bochum (nonconventional 

isotopes) 

Analytical work in München (shellultrastucture) 

Possible second field period (collection 

of OA-relevant shell material) 








- All analytical facilities are fine-tuned to the project proposed month 4-6 

- Fieldwork phase I completed, sample material available month 2-3 

- Experimental (cultured) shells from Bremen available month 10-12 

- Possible fieldwork phase II completed month 15 

- Data set on shell ultrastucture and geochemistry available month 20 

- Comparison of results with those of partner projects month 24 


- Completion of project month 36 

Subproject 3.5.2 Biological response to short-termed ocean acidification events in the past: 

biodiversity and evolution patterns of marine primary producers (calcareous nannofossils) 

during the late Paleocene – early Eocene 

(Mutterlose in collaboration with Schulte) 


The applicants plan to study the calcareous nannofossils, stable isotope (δ 13 C, δ 18 O, δ 11 B) 

patterns and the sediment petrography of the Late Palaeocene (NP 5) to Early Eocene (NP 12) of 

an ODP Site in order to contribute to the understanding of the following questions: 

(1) Does the isotope record provide evidence for δ 13 C excursions additional to the PETM? 

(2) What is the sequence of nannofossil patterns (diversity, abundance, size evolution, extinction 

and origination patterns) for the Palaeocene/Eocene hyperthermals? 

(3) Do the excursion floras of the PETM reflect an acidification of the ocean waters? 

(4) Are the Palaeocene/Eocene hyperthermals characterized by similar turnovers like the PETM? 

(5) Were T°C, nutrients, CO2, pH or salinity controlling the composition of the assemblages? 

(6) What are the concomitant sedimentological and mineralogical changes? 

(7) What are the paleoceanographic and palaeoclimatic implications of these findings? 

(8) Are these biotic changes a potential scenario for the future of our oceans? 

For this purpose it is here proposed to investigate the isotopic composition (δ 13 C, δ 18 O, δ 11 B) of 

calcareous nannofossils, and the mineralogy of the Late Palaeocene/Early Eocene interval from a 

low latitudinal ODP Site. The pre- and post intervals of the hyperthermal events will be analysed 

at a sample spacing of 5 – 10 cm as defined by the isotope stratigraphy. The events itself will be 

studied with a spacing of at least 1 sample/5 cm, critical intervals at a resolution of 1 sample/1 

cm. 

211

212 


Oxygen and carbon isotope data of benthic foraminifera will be analysed to construct a highresolution 

isotope stratigraphy and establish a long term palaeo-temperature record. The isotope 

data will also be used for (1) gaining information on palaeo-environmental changes, (2) defining 

the perturbations of the carbon cycle during the investigated interval, and (3) understanding the 

chemical composition of potentially different water-masses through time. It is also attempted to 

use the boron isotope record (δ 11 B) of foraminifera tests as a proxy for potential pH changes 

About 400 samples will be analysed for calcareous nannofossils. In each settling slide 300-400 

specimens will be counted in order to record the biodiversity and the absolute abundance of all 

taxa encountered. The data obtained will include information about preservation and etching; 

ranges of index taxa; diversity and abundance patterns; absolute abundances of nannofossils per 

gram sediment; changes in the nannofossil assemblages; extinction/ origination of species; and 

onset of malformed taxa. The dataset will be analyzed with respect to single species carbonate 

content using the SYRACO system at Kiel University. 

Grain size analysis of the



- Low resolution study samples analysed for calcareous nannofossils month 12 

- Magnetic data and mineralogical analyses completed month 12 

- High resolution study samples analyzed month 26 

- Isotope studies completed month 24 


- Compilation of study month 36 

Subproject 3.5.3 Nannoplankton response to modern and past ocean acidification events 

(Meier in collaboration with Kinkel) 


Subproject 3.5.3 makes use of fossil nannoplankton in the oceanic sediment record providing the 

largest archive of any fossil organism group throughout the Mesozoic and Cenozoic (Young 

1998; Kinkel et al., 2000; Bown et al., 2004; Baumann et al., 2005). The amount of calcite stored 

in coccoliths or coccospheres can be estimated using light-optical measurements (Beaufort 2005). 

For this the automated recognition system SYRACO (Beaufort & Dollfus, 2004) will be installed 

to generate species specific data on morphometry and weight estimates of key taxa from different 

OA events in Earth’s history. SYRACO provides data for the calcite stored in each coccolith or 

coccosphere and can therefore be used to identify heterogeneity in calcite production between 

individual liths and cells. The system reliably recognises the dominant modern species and can be 

trained to recognise additional taxa. Due to the large amount of samples SYRACO can analyse 

(about 50 slides per day), an unprecedented spatial and temporal resolution will be reached, and 

statistically highly significant data will be produced. Parallel SEM investigations will allow 

control on possible different genotypes, as well as dissolution or diagenetic overprint in fossil 

assemblages. With this approach, the following hypotheses will be tested: Does single species 

calcification depend on various environmental factors as opposed to ocean acidification alone? 

and Does ocean acidification affect specific taxa only? By investigating single species response 

to OA as well as generating estimates on total calcareous nannoplankton carbonate production it 

is planed to identify possible common patterns of calcareous nannoplankton community reactions 

to OA events. Recently Iglesias-Rodriguez et al. (2008) have challenged the assumption that OA 

will lead to a reduction of coccolithophore calcification. However, their record only covers the 

past 200 years and lacks control on species specific carbonate production. Thus it is not clear, 

whether the observed increase in coccolithophore calcification in the past decades is a response to 

anthropogenic CO2 emissions or reflects natural variability, and what the species specific 

response is. 

Therefore, the project aims to investigate natural variability versus the impact of OA and possible 

climate feedback in three different scenarios: (1) Holocene (i.e. the last 10’000 years) during 

times of minimum variability of the carbonate system. (2) Termination of the penultimate 

glaciation (i.e. ~132’000 years B.P.), during which a doubling of the pCO2 concentration 

occurred and is coupled with a strong and fast climatic change. (3) Paleocene/Eocene Thermal 

Maximum (~55 million years ago), that is characterised by extreme OA events that led to 

extinction of some species, whilst so-called disaster taxa including some coccolithophores and 

213

214 


many calcareous dinoflagellates show relatively little response to the OA event. For all three 

scenarios, species specific carbonate weight estimates and morphometric data will be generated. 

Measurements will also be carried out on cells and liths obtained from culturing studies on 

coccolithophores and dinoflagellates proposed in BIOACID (subprojects 1.1.3 Müller, 1.1.4 

Reusch, 3.1.1 Schulz and 4.2.2 Rost) to compare our observed nannoplankton response to natural 

OA events to those created in the laboratory. Our results can also be of significance for those 

projects investigating how pelagic calcification should be modelled (1.3 Schneider, 5.2 Oschlies). 

All necessary samples for our project are already available (Holocene, Termination II) at our 

institute or will be provided by partners within BIOACID (PETM, cultures). 

Schedule 


Installation of SYRACO 

Training of PhD student and SYRACO 

Analysis of Holocene samples 

Analysis of Termination II samples 

Training of PhD student and SYRACO 

for PETM species 

Analysis of PETM samples 

SEM control of sample quality 

Continuous monitoring of culture 

species 

Image and data analysis, interpretation 

Manuscript preparation, presentation at 

conferences 



- SYRACO operational month 03 

- PhD student trained in nannoplankton taxonomy month 03 

- Holocene dataset compilation finished month 12 

- Termination II dataset compilation finished month 21 


- PETM dataset compilation finished month 30 

- Project synthesis month 36




3.5.1 

3.5.2 

3.5.3 

Subtotal 

Consumables 

3.5.1 

3.5.2 

3.5.3 


€, PhD 

€, HiWi 

€, PhD 

€, HiWi 

€, PhD 

€, HiWi 

€ 

(Analytical work 

Bochum & 

München) 

€ 

(Isotopes etc,) 

€ 

(SEM etc.) 

€ 


Bochum & 

München) 

€ 


€ 

(SEM etc.) 

€ 


Bochum & 

München) 

€ 


€ 

(SEM etc.) 

Subtotal € € € € 

Travel 

3.5.1 

3.5.2 

3.5.3 

€ 

(Fieldwork, 

meetings) 

€ 

(Kiel, meetings) 

€ (Training, 

meetings) 

€ 

(Fieldwork, 

meetings) 

€ 


€ 

(Training, 

meetings) 

€ (Meetings, 

workshops) 

€ 


€ 

(Training, 

meetings) 

Subtotal € € € € 

Investments 

3.5.1 n.a. n.a. n.a. n.a. 

3.5.2 

3.5.3 

€ 

(camera micros) 

€ 

(SYRACO) 

n.a. n.a. 

n.a. n.a. 

€ 

€ 

€ 

€ 

€ 

€ 

€ 

€ 

215

216 



Subtotal € n.a. n.a. € 

Other costs 

3.5.1 n.a. n.a. n.a. n.a. 

3.5.2 n.a. n.a. n.a. n.a. 

3.5.3 n.a. n.a. n.a. n.a. 

Subtotal n.a. n.a. n.a. n.a. 

TOTAL 


3.5.1 

Personnel costs: The research is ambitious and technically complex and is to be carried out by a 

capable PhD student paid according to the financial regulations of the Ruhr-University Bochum. 

Consumables: The combined geochemical and material properties research is cost intensive. 

Coast for a δ 26 Mg isotope analysis are in the order of € depending on the number of 

repetitions made. In general, each data point represents a total of five repeat analyses. The same 

accounts for calcium isotope analysis. Shell ultrastucture work as carried out in collaboration 

with München is both time and cost intensive. Hence most of the finances go into consumables 

for analytical work (reference gas, high-precision thin sections etc.). In order to support the PhD 

with time consuming work, a HiWi is hired for a moderate number of hours per week. 

Travel: The PhD should participate at workshops and attend conferences. Fieldwork in Oman 

and probably at other PETM onshore localities is performed. 

Investment: Not applicable. 

Other costs: Not applicable. 

3.5.2 

Personnel costs: The research will be performed by a PhD student paid according to the 

financial regulations of the Ruhr-University Bochum. 

Consumables: This money is requested for running the various analyses (stable isotopes, clay 

min. etc.) suggested in the proposal. This will also cover the running costs for glass war, 

adhesives etc. 

Travel: We scheduled a) once per year a 3 weeks visit of the Bochum PhD student to Kiel, to 

work on the automated microscope to gain data and discuss results; b) once per year a meeting of 

the BioAcid group in Kiel, c) two short visits per year to Erlangen, d) at least one international 

meeting per year to present findings. 

Investment: For measuring the sizes of calcareous nannofossils a new microscope camera is 

needed. 

Other costs: Not applicable.

3.5.3 


Personnel costs: The research will be carried out by a PhD student who will be paid according to 

the German tariff ( - € per year). A student worker is requested for assistance 

with operating the automated system, sample preparation and image analysis ( ,- € per year). 

Consumables: 

SEM analyses, about 150 hours at 

SEM consumables 

Glass slides, embedding material, filters 

Travel: Training in Aix en Provence (1 st year), regular meetings within partners (1 st -3 rd year), 

international conferences (2 nd & 3 rd year). 

Investment: The automated recognition system SYRACO is a prerequisite for generating single 

species carbonate production estimates. The system can analyse up to 50 samples per day (for 

comparison, traditional nannoplankton PhD studies would be based on a few hundred samples). 

Considering the large number of samples to be analysed as proposed conventional counting and 

imaging techniques are too slow. The system contains an automated microscope ( ), a 

high resolution b/w camera ( ), and a computer with monitor ( ). 

Other costs: Not applicable. 


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Eocene at ODP Site 1262: Implications for calcareous nannoplankton evolution. Mar Micropaleontol 64:215-248 

Baumann KH, Andruleit H, Boeckel B, Geisen M, Kinkel H (2005) The significance of extant coccolithophores as indicators of ocean water 

masses, surface water temperature, and paleoproductivity: a review. Pal Z 79:93-112 

Beaufort L (2005) Weight estimates of coccoliths using the optical properties (birefringence) of calcite. Micropaleontology 51:289-297 

Beaufort L, Dollfus D (2004) Automatic recognition of coccoliths by dynamical neural networks. Mar Micropaleontol 51:57–73 

Bown PR, Lees JA, Young JR (2004) Calcareous nannoplankton evolution and diversity through time. Coccolithophores - from molecular 

processes to global impact. H Thierstein and J R Young. Berlin, Springer: 481-508 

Buick DP, Ivany LC (2004) 100 years in the dark: Extreme longevity of Eocene bivalves from Antarctica. Geology 32:921-924 

Calderia K, Wickett ME (2005) Ocean model predictions of chemistry changes from carbon dioxide emissions to the atmosphere and ocean. J 

Geophys Res 110: doi:10.1029/2004JC002671 

Hippler D, Witbaard R, Richter D, Buhl D, Immenhauser A (2008) Inter- and intra-species variability of Mg isotopes in marine biogenic 

carbonates. Geochim Cosmochim Acta in press 

Engel A, Zondervan I, Aerts K, Beaufort L, Benthien A, Chou L, Delille B, Gattuso JP, Harlay J, Heemann C, Hoffmann L, Jacquet S, 

Nejstgaard J, Pizay MD, Rochell-Newall E, Schneider U, Terbrueggen A, Riebesell U (2005) Testing the direct effect of CO2 

concentration on a bloom of the coccolithophorid Emiliania huxleyi in mesocosm experiments. Limnol Oceanogr 50:493-507 

Fabry VJ, Seibel BA, Feely RA, Orr JC (2008) Impacts of ocean acidification on marine fauna and ecosystem processes. ICES Journal of 

Marine Science 65:414-432 

Gibbs S, Bralower TJ, Bown PR, Zachos JC, Bybell LM (2006) Shelf and open-ocean calcareous phytoplankton assemblages across the 

Paleocene-Eocene Thermal Maximum: Implications for global productivity gradients. Geology 34:233-236 

Gibbs SJ, Bown PR, Sessa JA, Bralower TJ, Wilson PA (2006) Nannoplankton extinction and origination across the Paleocene-Eocene 

Thermal Maximum. Science 314:1770-1773 

Griesshaber E, Schmahl WW, Neuser RD, Pettke T, Blüm M, Mutterlose J, Brand U. (2007) Crystallographic texture and microstructure of 

terebratulide brachiopod shell calcite: An optimized materials design with hierarchial architecture. Am Mineral 92:722-724 

Iglesias-Rodriguez MD, Halloran PR, Rickaby REM, Hall IR, Colmenero-Hidalgo E, Gittins JR, Green DRH, Tyrell T, Gibbs SJ, Dassow vP, 

Rehm E, Arbrust EV, Boessenkool KP (2008) Phytoplankton Calcificaiton in a High-CO2 world. Science 320:336-340 

Immenhauser A, Kenter JAM, Ganssen G, Bahamonde JR, van Vliet A, Saher MH (2002) Origin and significance of isotope shifts in 

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Immenhauser A, Nägler TF, Steuber T, Hippler D (2005) A critical assessment of mollusk 18O/16O, Mg/Ca, and 44Ca/40Ca ratios as proxies 

for Cretaceous seawater temperature seasonality. Palaeogeogr Palaeoclimatol Palaeoecol 215:221-237 

Khim BK., Woo KS, Je JG (2000) Stable isotope profile of bivalves shells: seasonal temperature variations, latitudinal temperature grdaients 

and biological carbon cycling along the east coast of Korea. Cont Shelf Res 20:843-861 

Kinkel H, Baumann KH, Cepek M (2000) Coccolithophores in the equatorial Atlantic Ocean: response to seasonal and Late Quaternary 

surface water variability. Mar Micropaleontol 39:87-112 

Medlin LK, Sáez AG, Young JR (2008) A molecular clock for coccolithophores and implications for selectivity of phytoplankton extinctions 

across the K/T boundary. Mar Micropaleontol 67:69-86 

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Mutterlose J, Linnert C, Norris R (2007) Calcareous nannofossils from the Paleocene-Eocene Thermal Maximum of the equatorial Atlantic 

(ODP Site 1260B): Evidence for tropical warming. Mar Micropalaeontol 65:13-31 

Nicolo MJ, Dickens GR, Hollis CJ, Zachos JC (2007) Multiple early Eocene hyperthermals: Their sedimentary expression on the New Zealand 

continental margin and in the deep sea. Geology 35:699-702 

Pancost RD, Steart DS, Handley L., Collinsion ME, Hooker JJ, Scott AC, Grassineau NV, Glasspool IJ (2007) Increased terrestrial methane 

cycling at the Palaeocene-Eocene thermal maximum. Nature 449:332-335 

Raffi I, Backman J, Pälike H (2005) Changes in calcareous nannofossil assemblages across the Paleocene/Eocene transition from the paleoequatorial 

Pacific Ocean. Palaeogeogr Palaeoclimatol Palaeoecol 226:93–126 

Rexfort A, Mutterlose J (2006) Oxygen isotope of Sepia officinalis – a key to understanding the ecology of belemnites? Earth Planet Sci Lett 

247:212-221 

Riebesell U, Zondervan I, Rost B, Tortell PD, Zeebe RE, Morel FM (2000) Reduced calcification of marine plankton in response to increased 

atmospheric CO2. Nature 407:364-367 

Röhl U, Westerhold T, Bralower TJ, Zachos JC (2007) On the duration of the Paleocene-Eocene thermal maximum (PETM). Geochem 

Geophys Geosyst 8:doi: 10.1029/2007GC001784 

Schmahl WW, Griesshaber E, Neuser RD, Merkel C, Deuschle J, Götz A, Kelm K, Mader W, Sehrbrock A (2008) Crystal morphology and 

hybrid fibre composit architecture of phosphatic and calcitic brachiopod shell materials- an overview. Mineral Mag in press 

Vander Putten E, Dehairs F, Keppens E, Bayens W (2000) High resolution distribution of tracce elements in the calcite shell layer of modern 

Mytilus edulis: Environmental and biological controls. Geochim Cosmochim Acta 64:997-1011 

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11.5: Theme 4: Species interactions and community structure in a 



Regime shifts are rapid reorganizations of ecosystems from one relatively stable state to another. 

Ecosystem regime shifts may not always be smooth, but can be abrupt resulting from complex, 

non-linear processes (Scheffer et al. 2001) that are driven by specific alterations in the interaction 

strength or mode between key species. For example, alternative stable states of coastal benthic 

communities can be triggered by pathogens or altered predator prey relationships of key trophic 

interactions (Sanford 1999). As such, relatively small changes in species composition may 

transform ecosystems from consumer to producer-dominated communities and vice versa 

(Scheibling 1986). To date, regime shifts have been often linked to climate forcing, e.g. 

fluctuations in the North Atlantic oscillation, El Niño events, or global climate change. For 

example, several warm water species have taken advantage of the warmer waters and expanded 

their ranges polewards (Hays et al. 2005), whereas other, cold-water adapted, species have 

retreated in the same direction (Perry et al. 2005). These range shifts had strong negative effects 

on the recruitment of higher trophic levels (e.g. fish and seabirds) that feed on key boreal species, 

such as cod feeding on the calanoid copepod Calanus finmarchicus (Beaugrand et al. 2003). 

Regime shifts and other structural re-organizations of marine communities are caused by diverse 

and imbalanced responses of different species or guilds to a changing environment. Even within 

species, susceptibility to stress can differ between genotypes and between different ontogenetic 

stages. This variability of stress effects at different organisational levels may lead to changes in 

trophic interactions or competitive hierarchies (Tortell et al. 2002), and result in a conspicuous 

shift in structural and functional properties of a community. 

Not only temperature, but also ocean acidification (OA) is a potential stress factor with a diverse 

set of individual reactions. As a result of the increasing pCO2, not only the stress related to 

acidification, but also the change in the availability of different nutrients (Si, N, P) in relation to 

carbon will change the competitive relationships between primary producers, especially in the 

light of the reduction in anthropogenic input of phosphorus and nitrogen in many coastal seas. 

Moreover, OA is accompanied by increases in water temperatures of ~2 to 5°C in many 

temperate regions. Therefore, studying the impacts of OA on community level biological 

interactions and cascades through the foodweb needs to consider potential synergisms between 

increases in temperature and decreases in ocean pH. Temperature increase will, among others, 

accelerate the metabolic rates of producers, consumers and remineralizers, whereas acidification 

may, among other effects, negatively affect the nutritional quality of primary consumers and 

detritus. Hence, the likelihood for consumer-prey mismatch situations may increase. In the 

marine benthos, for example, macroalgae and seagrasses, which often lack efficient carbon 

concentration mechanisms may benefit from increased pCO2 availability. In contrast, benthic 

filter feeders such as bivalves, bryozoans, corals or tube building polychaetes may suffer declines 

in their calcifying capacity (Shirayama & Thornton 2005), with concomitant decreases in 

population growth and persistence. Along rocky shores in particular, where marine filter feeders 

directly compete with marine plants for space, a climate change driven restructuring may shift 

communities from filter feeder dominated consumer communities to macroalgae dominated 

communities. 

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In short, ocean acidification may have important consequences for the biodiversity and 

functioning of marine ecosystems: even moderate stress impacts on individual species may be 

dramatically amplified (or buffered) by ensuing modulation of niches and interactions which 

ultimately will provoke a restructuring of communities. As a consequence, important ecosystem 

services (e.g. O2 production, CO2 sequestration, remineralization of nutrients, fisheries yields) 

may be jeopardized. Extending our understanding of stress impacts from the individual or species 

level to the level of interactions and community dynamics is crucial. 

The main gaps in our knowledge 

Our knowledge on the effects of OA has increased considerably in recent years, but there are still 

enormous gaps that need to be filled with regard to complex community responses. This 

knowledge is critical to assess the impacts of OA on all scales as well as to provide policy makers 

with the tools and information necessary to implement the necessary actions. One of the main 

gaps concerns the effect of OA on the interactions between species and the resulting community 

structure. Whereas many studies exist on the effects of low pH and/or high CO2 conditions on 

single organisms, not much is known about how these factors affect interactions between species 

and, through that, communities. Indeed, the predicted shifts in pCO2 and pH in many species will 

often only slightly impact performance and fitness of a given species. However, as shown for 

other stressors (Christensen et al. 2006, Wahl 2008) ensuing modifications of species interactions 

may substantially amplify or buffer the original stress (Wahl et al. 2004). How environmental 

stress spreads through a community via shifts in composition and interaction is the main focus of 

this theme. 

We define two central research questions dealing with interactions between organisms. (1) First, 

we ask how competitive interactions will change in a high CO2 world, and whether and how this 

may lead to alterations in the structure and functioning of communities. Species competing for 

similar resources may have completely different requirements to the physical environment. Most 

obviously, calcifying organisms may be more affected by a decrease in pH than non-calcifying 

organisms, and this should lead to a change in the competitive interactions between the two (Fig 

4.1d). However, we can also envisage more subtle effects where slight differences in for example 

carbon concentrating mechanisms may affect the competitive outcome between or even within 

species. This question is very important with regard to highly diverse bacterial communities and 

their key role as bioreactor for the remineralization of nutrients. Additionally, fitness shifts in 

macroorganisms and/or compositional changes in the colonizer pool will lead to altered epibiotic 

biofilms indirectly impacting the biotic and abiotic interactions of the host. Increasing CO2 levels 

will also increase the C:nutrient ratios available for primary production, and as these nutrient 

ratios can be considered an ecological niche (Loladze et al. 2004) we expect structural shifts in 

algal communities. We will investigate these competition mediated effects at different functional 

levels including macroalgae, benthic invertebrates, microalgae, and bacteria as we expect impacts 

to differ between these functional groups. Since further differences are expected between 

genotypes within species, microevolutionary aspects are also included.

C:N Ratio (molar) 

Respiration (µg O2 min -1 Animal -1 ) 

12 

11 

10 

9 

8 

7 

6 

5 

1.2 

1.0 

0.8 

0.6 

0.4 

0.2 

0.0 


200 ppm 

600 ppm 

900 ppm 

0 1 2 3 4 5 6 7 8 

Day of incubation 

nutrient replete High C:N High C:P 

Treatment of the food 

Rhodomonas salina (Ind ml -1 (d rate growth marina Oxyrrhis 

a 

0 

1.0 

b 

0.8 

0.6 

0.4 

nutrient replete 

High C:N 

High C:P 

0.2 

0.0 

1e+5 2e+5 3e+5 4e+5 5e+5 6e+5 

) 

c 

Crustose coralline algae 

Non-calcifying algae 

400 765 

CO2 treatment (ppm) 

Fig. 4.1: Examples of previous relevant work. a) nutrient stoichiometry of Rhodomonas salina under different CO 2 

concentrations (Gülzow 2008); b) Oxyrrhis marina growing on food with different nutrient stoichiometry (Hantzsche 

& Boersma submitted); c) Respiration of Acartia tonsa on different food sources (Boersma et al. unpubl); d) 

competitive interactions between calcifying algae and non-calcifying algae under different CO 2 conditions (Kuffner et 

al. 2007) 

(2) The second major aim of this theme is to elucidate the effects of OA on food web 

interactions. Obviously, loss of species or the replacement of one species (strain) with another 

one (diatoms versus dinoflagellates) as a result of OA will affect the flow of energy and matter 

through the food web, but more subtle processes may be just as relevant. High CO2 availability to 

primary producers may affect their quality as food for herbivores. Not only do we know that the 

production of secondary metabolites is highly dependent on the nutrient conditions (Velzeboer et 

al. 2001, Lippemeier et al. 2003), we also know that the concentration of essential components of 

the food such as fatty acids is dependent on the growing conditions of the algae (Müller-Navarra 

1995, Boersma 2000). Moreover, the stoichiometry of the macronutrients may change in primary 

producers as a result of high CO2 availability (Fig 4.1a and: Burkhardt et al. 1999, Riebesell et al. 

2000, Taraldsvik & Myklestad 2000, Urabe et al. 2003, Riebesell et al. 2007). As primary 

producers are not homeostatic with respect to their nutrient composition, they usually reflect the 

nutrient composition of their surrounding water, and hence are being expected to also show 

higher carbon-to-nutrient ratios. High carbon-to-nutrient algae are known to be inferior food 

quality for herbivorous organisms (Fig 4.1b and: Boersma 2000, Sterner & Elser 2002, Boersma 

& Elser 2006). These consumers have to handle more energy (carbon) in relation to nutrients 

needed for their assembly and acquisition machinery (phosphorus rich ribosomes and nitrogen 

rich proteins and mitochondria, respectively) (Klausmeier et al. 2004), which creates costs and 

leads to altered nutrient stoichiometry and reduced growth and reproduction of the consumers. 

These food quality effects have only recently been shown to travel up food chains to predatory 

d 

70 

60 

50 

40 

30 

20 

10 

0 

-1 ) 

Percentage cover 

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beetles in terrestrial systems (Kagata & Ohgushi 2007), to parasites feeding/living on nutrient 

limited hosts (Frost et al. 2008) and even to planktivorous fish (Malzahn et al. 2007a). The latter 

finding is very important, as it implies that through OA, and the resulting increase in the 

C:nutrient content of the algae (especially in the light of the re-oligotrophication of several 

coastal seas) the production of zooplankton should decline as a result of lower food quality. 

Moreover, the quality of this reduced stock of zooplankters for higher trophic levels should be 

low, thus potentially impacting production of economically important fish species. 

Changes in nutrient stoichiometry on all levels from water chemistry to secondary consumers are 

likely to impact the community structure and functioning of microbial communities as well. Not 

only as a result of direct stress effects, but also as a result of changes in excretory products of 

consumers as a result of their homeostasis which will affect the available carbon pool for bacteria 

(Fig 4.1c, different respiration rates under different food conditions). This is likely to favour new 

species assemblages, and hence, bacterial community functioning. 

Consequently, this theme comprises investigations of the horizontal (competitive interactions) 

and vertical (food chains) translation of abiotic stress into structural changes of the benthic and 

pelagic, microbial and macrobial communities considered (See Fig. 4.2 for an example in the 

benthic environment). 

Consumers 

Defence 

Food quality 

Compensatory 

growth 

Abiotic Stress 

acidification, warming 

Fucus 

Epibiosis 

Abundance & 

Depth 

distribution 

Recruitment 

Growth 

Competitors 

Fig. 4.2: Horizontal and vertical interactions with the macroalga Fucus as an example of a target species: Abiotic stress 

affects directly but often with different strength and/or sign the performance of interactors (target species Fucus, its 

consumers, competitors and epibionts). The ensuing shifts in biotic interactions may have still stronger effects on the 

survival and distribution of the target species than the direct stress itself. 

Objectives 

By means of manipulative experimentation from the individual up to the community level we 

will: 

1. Establish the role of differential sensitivities to OA at the community, species and 

subspecies (ontogenetic stages, genotypes) level 

2. Assess the synergistic effects of acidification and warming


3. Analyse changes in food quality and quantity of primary producers under various CO2 

concentrations 

4. Quantify the ensuing shifts in competitive abilities of benthic and pelagic model organisms 

5. Quantify the ensuing shifts in the performance and ecological efficiency of energy transfer 

between lower and higher trophic levels in pelagic and benthic systems 

6. Assess and model cascading effects in coastal marine systems from secondary consumers 

down to microbial communities via trophic, competitive and epibiotic interactions 

7. Compare the effect of natural and experimental stress gradients on community structure 

and dynamics 

8. Compare the relative impact of acidification/warming stress on different types of 

communities (benthic – pelagic, microbial – macrobial). 

Approach 

Different types of experimental designs will be applied to address changes in community 

structure, competitive interactions as well as nutritionally mediated responses in consumers over 

several trophic levels. The first type of setup will consist of using controlled laboratory systems 

that subject organisms to different levels of acidification (and warming) and investigate how 

sensitivities differ between genotypes, ontogenetic stages, and/or species and communities. 

Secondly, primary producers cultured under different conditions will be fed to consumers in 

sequential steps. This will allow quantification of the immediate impact of prey quality on the 

performance of secondary producers using advanced analytical methods to determine food 

quality parameters. The third type of experiments will consist of environmental samples from 

natural pH or CO2 gradients and larger scale in-door mesocosm setups within walk-in 

temperature controlled rooms where both temperature and several levels of acidification can be 

manipulated in a factorial design. This setting can be used for both pelagic and benthic 

community model systems, to study changes in community structure and interaction under 

different CO2/temperature scenarios. 


This theme involves two subprojects, both dealing with the gaps in our current knowledge 

described above. One project (4.1) investigates processes in the benthic system, whereas the other 

one (4.2) studies pelagic processes. Both projects comprise a number of complementary 

subprojects, which differ with regard to their focus on community type, habitat and/or interaction 

type. Within projects, the common habitats allow joint experiments, and combined sampling 

campaigns. Between the projects the joint questions allow us the development of common 

strategies and concepts. Moreover, investigating identical or similar questions over a range of 

differently sized organisms with different generation times and reproduction modes also enables 

us to test the robustness and generality of our findings as well as the dependency of our results on 

the differences between systems. Strong collaborations will exist with different projects in the 

other themes, more specifically with those projects investigating single species responses to OA 

in themes 2 and 3, and the microbiological projects in theme 1. 

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224 


Project 4.1: OA impacts on interactions in and structure of benthic communities 

(M. Wahl) 

i. Objectives 

To assess the impact of ocean acidification (for some experiments in conjunction with predicted 

warming) on 

• performance of genotypes and populations 

• competitive interactions within and between species 

• associations such as epibiosis 

• structure and function of communities 

• microbial as compared to macrobial communities 


Ocean acidification (in conjunction with warming) effects in benthic habitats can be found at 

very different levels of organisation. Thus, the expected abiotic shifts may impact the activity of 

important enzymes, calcification processes and the general performance or fitness of individuals 

(Saborowski et al. 2004, Fabry et al. 2008). Such organismic responses to climate change are 

conventionally categorized as either ecological or evolutionary. The former includes phenotypic 

plasticity and dispersal, while the latter entails genetic change (Reusch & Wood 2007). 

Evolutionary theory suggests that with increasing unpredictability of the environment, as 

expected under global change, selection should favour genotypes that possess a broader tolerance 

towards stress and other environmental challenges. Ontogenetic and genetic differences in the 

susceptibility to acidification and warming may affect population dynamics and intra-specific 

competition. Within populations, susceptibility to stress changes in the course of ontogeny (Fabry 

et al. 2008) and often highest mortality occurs at early life stages (Gosselin & Qian 1997). 

The direct effects of abiotic stress such as predicted during global change will usually not be fatal 

at the individual or species level, but when they modify interactions or associations (such as 

epibiosis) the impact of these stress factors may be substantially larger (Wahl 2008). When cooccurring 

and competing species are differentially impacted by this abiotic stress we may expect 

shifts in the outcome of competitive interactions and, ultimately, in the structure and functioning 

of the community (Jaschinski & Sommer 2008). Indeed, the modulation of interactions by shifts 

in environmental conditions can be considered as ecological levers which may alter the purely 

physiological impact of the stressor(s) in 2 directions: (i) at the level of the individual the direct 

physiological effects may be masked, dampened or enhanced by changes in biological 

interactions such as predation or competition and (ii) the impacts at the species level – even when 

non-lethal – by interaction modulation will be projected onto the community level and may lead 

to “surprising” (Christensen et al. 2006) re-organisations of the structural and functional 

properties of a community. While population or community level effects have received little 

attention, one particularly understudied component of benthic systems in this regard are 

microbial communities (Inagaki et al. 2006). 

We will compare the interaction-mediated responses of benthic populations and communities in a 

variety of complementary systems: seagrass and macroalgae beds, macrobenthic and seafloor 

microbial communities. Where possible, regional and systemic comparisons will be carried out 

(e.g. tropical – temperate, deep sea – shore habitats).


Most of past research on hypercapnia effects has focussed either on tropical reefs or on 

planktonic organisms (Fabry et al. 2008). However, some examples for interaction mediated 

effects of environmental stress in benthic systems have been reported. Calcifying organisms, such 

as coralline algae, may suffer a reduction in competitiveness in the course of environmental 

acidification (Kuffner et al. 2007). Consequently, their non-calcifying local competitors may 

indirectly benefit from OA even if they are only slightly less negatively impacted by this 

environmental stress than the corallines. Competition between algae with and without carbon 

concentration mechanisms is expected to shift when pCO2 rises (Zimmerman et al. 1997). A 

higher C:N:P ratio as a possible consequence of CO2 enrichment may decrease food quality and 

consumer fitness (e.g. Cruz-Rivera & Hay 2000). Shifts in predation pressure are expected. Also, 

two (or more) stressors may interact and modulate their respective effects on a given response. 

Thus, growth and shell properties play important roles for the competitiveness and the 

susceptibility to predation of an important bioengineer such as mussels (e.g. Gutierrez et al. 

2003). The interaction of acidification, warming and other stressors (e.g. eutrophication, 

desalination) affect these traits in various and often unexpected ways with far-reaching 

consequences for this species’ persistence (Kossak 2006, Wahl et al. work in progress). 

Hence, it is essential to expand the OA impact studies beyond the species level to community 

structure, comprising both “horizontal” interactions (competition among basal species, epibioses) 

as well as the “vertical” aspects, i.e. food quality and trophic interactions. The interaction with 

the second project in this theme will be very strong. 

Principal foci will be shifts in nutritional quality/prey palatability and predation-mediated 

environmental stress (4.1.1, 4.1.2), competitional shifts through species specific differences in 

stress susceptibility (4.1.1, 4.1.3.), restructuring of populations by differences in stress sensitivity 

between ontogenetic stages and genotypes (4.1.2, 4.1.3.), interaction-modulation through altered 

epibioses (4.1.1, 4.1.2, 4.1.3.), restructuring of microbial and macrobial communities through 

interaction-mediated stress effects (4.1.2, 4.1.4) 


4.1.1 

Ragnhild Asmus has long-standing experience in benthic ecology of soft bottom areas especially 

temperate and tropical seagrass beds (Asmus & Asmus 2000). She also is experienced in field 

experiments and community ecology with a focus on the ecology of benthic primary producers in 

different soft bottom communities. She has large expertise in network analysis of food webs and 

stable isotope techniques (Baird et al. 2004, 2007). Christian Wiencke has long standing 

expertise in seaweed ecology and physiology and in research on anthropogenic effects (e.g. 

increased UV radiation) on seaweed performance (Wiencke et al. 2006, Roleda et al. 2007, 

Steinhoff et al. 2008). Lars Gutow studies plant-grazer interactions at both the ecological and the 

physiological level with emphasis on the implications of nutritional quality and quantity on 

meso-herbivore fitness (Gutow et al. 2006, Gutow et al. 2007). 

4.1.2 

Martin Wahl has extensive experience in benthic community ecology with a strong emphasis on 

complex biotic interactions (defences, epibiosis, consumption, competition), their modulation by 

abiotic factors, and the impact of environmental stress. His experimental approach comprises in 

situ SCUBA experiments, climate simulation mesocosm experiments and analytical lab 

techniques (Wahl et al. 2004, Sugden et al. 2008, Wahl 2008). Thorsten Reusch has broad 

225

226 


experience in ecological genetics of aquatic and marine organisms, including the development 

and analysis of genetic marker data, the set-up of selection experiments and their analysis. 

4.1.3 

Kai Bischof is studying the eco- and stressphysiology of benthic macroalgae from polar, 

temperate and tropical ecosystems (Bischof et al. 2006a). Interactive stress effects on macroalgal 

photosynthesis, growth, reproduction and competition, as well as physiological acclimation 

mechanisms (e.g. antioxidative strategies (Bischof et al. 2006b)) are the prime focus of his 

research. By his workgroup tropical reef algae are successfully cultivated at the ZMT aquaculture 

facility. 

4.1.4 

Alban Ramette has a broad expertise in high-throughput molecular characterization of microbial 

communities in environmental samples and in the further statistical evaluation of multivariate 

data (Ramette & Tiedje 2007). Antje Boetius has extensive experience with biogeochemical and 

microbiological investigations of marine sediments, both in the field including in situ 

measurements and in experimental approaches. In addition, previous molecular work has been 

successfully accomplished at the proposed natural CO2 reservoir site (Inagaki et al. 2006). 


Subproject 4.1.1 Effects of ocean acidification on trophic interactions in coastal seaweed 

and seagrass ecosystems 

R. Asmus, C. Wiencke, L. Gutow, H. Asmus, D. Hanelt, R. Saborowski, I. Bartsch 

This subproject provides many opportunities for close cooperation with other BIOACIDprojects. 

The reference numbers of cooperating subprojects are given in brackets. 

The combined effects of ocean acidification and increased seawater temperatures on marine 

macrophytes from the North Atlantic will be investigated in multi-factorial experiments under 

controlled laboratory conditions [3.2.4, 4.1.3]. In the facilities of the AWI at Sylt and 

Bremerhaven selected seaweed and seagrass species will be raised under constant pCO2-levels 

representing pre-industrial (280 ppm), current (380 ppm), and predicted future (700 ppm) pCO2conditions 

and constant temperatures of 10 and 20°C. An additional experimental temperature 

of 25°C will be applied to the seagrass species. The selected species will include the North Sea 

seagrass species Zostera marina and Z. noltii and representatives of the three major taxonomic 

seaweed groups (i.e. Chlorophyta, Rhodophyta, and Phaeophyceae). 

For each species photosynthetic activity, growth, and elemental composition (C, N, P) will be 

measured for each pH/temperature combination [4.1.2, 4.1.3, 4.2.2]. Photosynthetic activity will 

be measured by pulse amplitude modulated (PAM) fluorometry and O2-electrodes. Since both 

methods do not measure carbon fixation directly, net photosynthetic rates will be inferred from 

exemplary accompanying 14 C-isotope measurements. Brown algae produce polyphenolic 

compounds such as phlorotannins via the secondary metabolic pathway. Besides their structural 

and UV-protective function phlorotannins act as feeding deterrents for many herbivores. Due to 

this implication on algal palatability the phlorotannin content will be quantified in brown algae 

by the Folin-Ciocalteu method.


In no-choice feeding experiments seaweeds grown under different pH/temperature treatments 

will be offered to selected meso-herbivores (peracarids and gastropods) collected in natural 

seaweed beds of Sylt and Helgoland. These experiments will focus on the treatment effects on 

macrophyte nutritional quality. In order to avoid pH/temperature effects on the grazer 

physiology the experiments will be conducted in untreated (but filtered) North Sea water at a 

constant temperature of 15°C. Each individual grazer will be fed with one seaweed species from 

one treatment only. In short-term experiments we will study the effects of altered food 

chemistry on consumption rates, assimilation efficiency, and respiration of the grazers [4.2.1]. 

The effects of altered food quality on grazer fitness will be studied in long-term experiments. 

Fitness will be determined in terms of growth, reproduction, and survival of the animals. In 

population models, these life history parameters will be used to calculate fitness parameters 

such as population growth rates. 

We will study how the combined effects of temperature and acidification affect the activity of 

highly important enzymes like cellulase, laminarinase, endo- and exopeptidase, phosphatase, 

and lipase [1.2.1]. Digestion and assimilation of the diets by the grazers will be determined by 

the activities and the characteristics of selected digestive enzymes in the digestive tract of the 

animals, and by the chemical composition of the faecal pellets in comparison to the food (C, N, 

P). Comparison between pH effects on the activities of endogenous digestive enzymes and of 

those produced by heterotrophic bacteria will be performed in collaboration with project [1.2.1]. 

The consequences for the decomposition of faecal pellets and nutrient recycling will be studied 

by the activity of digestive enzymes within faecal pellets and by the release of organic and 

inorganic degradation products (carbohydrates, proteins, phosphate) [1.2.1]. 

In natural seagrass beds, the removal of seagrass epiphytes by grazers has strong effects on 

seagrass performance. Seagrasses that were grown together with their epiphytes under different 

pH/temperature regimes will be offered to selected meso-herbivores in feeding experiments in 

order to investigate how ocean acidification in conjunction with increasing temperatures will 

affect the balance between epiphyte growth and removal by grazers and the possible 

implications on seagrass performance (measured as biomass increment, e.g. leaf area index) 

[1.1.5, 4.1.2, 4.1.4]. These experiments will be conducted in untreated (but filtered) North Sea 

water at a constant temperature of 15°C. 

Ecosystem consequences of ocean acidification will be addressed in CO2-incubated benthic 

chambers (5 to 500 dm 3 volume) in Wadden Sea seagrass beds at Sylt [4.1.2, 5.1]. In situ and 

benthic mesocosm enclosures (ca. 3 m 3 ) of partial systems of a seagrass bed will be exposed to 

different CO2 concentrations (from weeks to ½ year). System effects will be measured as 

changes in biomass, abundance, oxygen and nutrients (N, P, Si) to compute growth, respiration, 

productivity, and egestion of the community and particular community compartments [1.1.5, 

4.1.4]. The results will be subject to a comprehensive network analysis (ENA) of altered trophic 

and metabolic pathways in seaweed and seagrass systems in order to identify the sensitivity of 

trophic pathways within these systems to acidification of coastal waters. 

227

Schedule 

4.1.1 

Incubation of macrophytes at different 

pH/temperature combinations 

Laboratory experiments on macrophytes 

Laboratory experiments on herbivores 

Laboratory experiments on fecal pellet decomposition 

Field experiments on community effects in benthic 

chambers 

Analysis of experimental data 

Network analysis of data from benthic chambers 

Presentation of data and results, manuscript 

preparation 

228 





- Incubated seaweed and seagrass species will be available for laboratory 

experiments 

month 6 

- First experimental data on pH/temperature effects on macrophytes month 15 

- First results from benthic chamber experiments month 21 

- Experimental data on consumption and assimilation efficiency of grazers month 24 

- Experimental data on pH/temperature effects on fecal pellet decomposition month 24 

- Data set on pH/temperature effects on macrophytes completed month 30 

- Experimental data on the effects of altered macrophyte stoichiometry 

on grazer fitness 

month 33 

- Final results from benthic chamber experiments month 33 

- Evaluation of data sets month 35 

Subproject 4.1.2 Acidification stress: Early life stage ecology in times of global change. 

M. Wahl & T. Reusch 

We will ask to which extent under different pCO2/temperature settings survival and performance 

of early life stages of some key species are affected (year 1), how sensitivity varies among 

genotypes (year 2), and how intra- and interspecific competitiveness is determined by differences 

in sensitivity among genotypes and species, resp. (year 3). We expect early life stages to react 

more sensitively to stress as compared to adults [in cooperation with projects 2.1.1, 2.1.3, 2.3.1]. 

Many of these aspects will be run in close cooperation with other projects (reference number in 

brackets). In benthic hard bottom communities of the Western Baltic, barnacles (Balanus 

improvisus, Bi) and mussels (Mytilus edulis, Me) are the ecologically most important suspension 

feeders. The bladder wrack Fucus vesiculosus (Fv) is an important primary producer and 

ecosystem engineer. The small calcareous polychaete Spirorbis spirorbis (Ss) is a suspension 

feeder living preferentially as epibiont on macroalgae. All 4 species control a substantial amount 

of the flow of matter between the open water and the benthos. They compete directly for 

resources (food and/or attachment substratum) and indirectly by attracting or distracting shared 

consumers. 

All experiments will be run in micro- and mesocosms allowing the control of all relevant 

variables. Survival and performance (year 1) will be studied on freshly settled juveniles obtained


from in vitro reproduction (Bi, Fv, Ss) or field sampling (Me in June – August). Acidification 

will be achieved by bubbling with CO2 enriched air at defined and automated concentrations. 

Acidification treatment levels as agreed project-wide (i.e. 380 – 560 – 980ppm) will be combined 

with 2 temperature levels (ambient – predicted ∆5°C). Species performance (photosynthesis as 

assessed by in situ PAM, respiration rates, growth, calcification) [2.1.3, 3.1.3, 3.1.4, 3.5.1, 4.1.3] 

will be assessed in the absence of biotic stressors (fouling, grazing, competition). Although we 

estimate the immediate stress impact to be small relative to its long-term and indirect effects we 

will strive to develop a reliable quantification of the stress level experienced (potential stress 

indicators: heat shock proteins, ion-regulation proteins, hypoxia-induced factor) [3.2.3]. 

Increased variance with stress strength will serve as an indication for inter-genotype variation in 

sensitivity and allow selecting suitable species for the 2 nd question. In the second year we will run 

simplified pilot studies on the role of genetic diversity for stress resistance [1.1.4, 2.2.1] which 

will be substantially deepened in a possible second 3-year period of BIOACID (i. e. using 

genetic marker loci, either gene-linked or anonymous microsatellites, both of which are 

currently being developed in Me and Fv). In a first step we will assess differences in stress 

susceptibility between genotypes (equivalent to individuals in these sexually reproducing species) 

using the performance parameters defined above. For this experiment, only species will be used 

for which reproduction, embryogenesis and recruitment can be handled in the lab, i.e. Fv, Bi a/o 

Ss. In a second and optional step [1.1.4], we assess potential evolutionary mediation of survival 

and plasticity evolution using controlled crossings that serve as base to assemble genetically 

diverse versus depauperate larval pools. This can be realized by either testing sibships alone or in 

various combinations (Gamfeldt et al. 2005). Ideally, sibships from mother-father pairs which 

differ in stress-sensitivity will be used. By obtaining half-sibs of identical female individuals 

fertilized by different fathers, a heritability analysis of tolerance to acidification will provide 

data on the predicted evolutionary response of the targeted key species to 

acidification/warming. High heritability estimates in conjunction with strong selection 

differential would indicate that persistence of benthic key species is possible due to 

microevolution (Falconer & Mackay 1996). At the community level, we will finally (year 3) 

investigate whether competitive hierarchies [3.2.2, 4.1.1] among selected sessile species (Bi, 

Me, Fs) change under various acidification/warming scenarios. Competitiveness of a species 

may be modulated by different levels of epibiosis resulting from weakened antifouling 

defences or altered fouling pressure [4.1.1, 4.1.4, 4.2.1, 4.2.2]. To this purpose we will 

assemble 2- or 3-species communities of Bi, Me a/o Fv at comparable initial density and 

genetic diversity, and follow their structural changes over time under different pCO2/T 

settings. [In a second 3-yr phase of BIOACID we will aim to disentangle the effects if species 

identity and genetic diversity on competitiveness.]. 

Schedule 


Survival and performance 

Genetic variability 

Competition 

Analysis & Publication 


229


230 



- Experimental data on CO2/pH sensitivity of juveniles month 10 

- Experimental data genetic variability of CO2/pH sensitivity month 21 

- Data set on changing competitiveness month 30 

- Evaluation of combined data sets, sensitivities & uncertainties month 36 

Subproject 4.1.3: Competitive success of calcifying and non-calcifying macroalgae under 

shifting pH regimes in tropical vs. temperate regions 

K. Bischof, A. Kunzmann, M. Nugues, T. Rixen, M. Teichberg 

We will examine the physiological response of calcifying and non-calcifying macroalgae to 

multiple abiotic stress and predict their respective competitive success on tropical reefs and 

temperate rocky shore habitats in a scenario of ocean acidification. 

The main questions we will address are: 

1) To what extent is photosynthesis and growth of calcifying macroalgae affected by shifts in pH, 

temperature and (PAR & UV) radiation in comparison to non-calcifying macroalgae, and 

how are these responses modified under uni- vs. multifactorial stress? To what extent is 

calcification affected in tropical vs. temperate algal species in the respective treatments? 

2) What are the underlying physiological acclimation mechanisms involved in stress responses? 

3) How will the competitive strength of calcifying and non-calcifying macroalgae be altered by 

future ocean acidification? 

4) How will local stress factors such as eutrophication interfere with the competitive success of 

calcifying, and non-calcifying, particularly ephemeral opportunistic macroalgae under ocean 

acidification? 

We will examine the physiological response of commonly found calcifying and non-calcifying 

macroalgae from the tropics and Helgoland (e.g. Padina, Halimeda, Corallina, Lobophora, 

Hypoglossum, Entermorpha, Acrosiphonia), applying uni- and multifactorial stress experiments 

in the laboratory and mesocosms at ZMT, University of Bremen, AWI Sylt and AWI Helgoland. 

(coordinated with 4.1.1-4.1.4). In the second year of the project, field and mesocosm studies will 

be conducted at a tropical field site (e.g. Similan Islands, Thailand). In the experiments we will 

expose the algae to different CO2 regimes including the pre-industrial level (280 ppm), present 

day conditions (380 ppm) and 560, 700 and 1000 ppm. In short-term experiments temperature, 

UV-radiation and nutrient levels will be changed according to the degree of expected variation of 

the respective factor within the respective habitat. In mesocosm ponds run at different pH and 

temperature conditions and field set-ups we will offer sterile settlement tiles as well as coralline 

algae as substrate and monitor algal recruitment, primary succession and epibiosis under varying 

abiotic conditions and examine the interspecific competition of calcifying and non-calcifying 

macroalgae. In all experiments we will measure photosynthesis (by oxygen evolution and 

chlorophyll fluorescence), growth and uptake of dissolved inorganic C and N on an


individual/species specific and community base. In situ measurements of calcification will be 

conducted in cooperation with Project 3.4.2. using microsensors. We will measure oxidised and 

total glutathione content (GSH/GSSG) as an indicator of oxidative stress, and thus a measure of 

general stress exposure, easily to compare between species. Studies on coral/alga interactions will 

be conducted in cooperation with the projects 3.2.2 and 3.2.3, and by Dr. Maggy Nugues through 

institutional funding by ZMT. We will closely cooperate with projects on trophic interactions 

within seagrass and seaweed dominated communities under shifting pH scenarios [4.1.1], and 

with 4.1.4. on bacterial communities on these organisms. An exchange of samples to study 

changes in ultrastructure of the CaCO3 matrix of the different calcifying macroalgal species under 

examination is envisaged with project 3.2.4. 

Schedule 


Development of exp. infrastructure 

Cultivation of macroalgae under 

varying pH/temperature 

Stress experiments in the laboratory 

Cruise preparation 

Field experiments (1 st year 

Helgoland/Sylt, 2 nd year Tropics) 

Sample processing 



Manuscript preparation, conferences 



- CO2 incubation chamber at ZMT will be available to the project month 3 

- First laboratory data on pH/temperature effects on temperate and 

tropical macroalgae 

month 6 

- Results from field & mesocosm experiments in temperate regions 

(physiology vs. competition) 

month 12 

- Results from field & mesocosm experiments in tropical regions 

(physiology vs. competition) 

month 24 

- Last laboratory studies & experimental work completed 

- Data implementation: 

month 30 

1.) patterns in physiology and competition: temperate vs. tropical habitats month 32 

2.) common patterns within the other projects of cluster 4 month 34 

- Evaluation of data sets, publication, modeling approaches month 35 

231

232 


Subproject 4.1.4: Effects of ocean acidification on microbial community structure, 

composition and activity in natural and experimental systems 

A. Ramette & A. Boetius 

Longterm CO2 effects on microbial diversity, biomass and function in the seabed 

Transects of seabed through which CO2-rich porewaters seep from a natural CO2 reservoir 

(sediment-hosted CO2 lake of the southern Okinawa Trough hydrothermal system (Inagaki et al. 

2006)) will be used as natural laboratory for the study of long-term consequences of varied CO2 

and pH levels on microbial community structure. Long term adaptation and consequent 

structuring of natural benthic communities cannot be studied experimentally. A series of 

molecular techniques will be applied to determine shifts in community structure (T-RFLP, 

ARISA), community composition (sequencing of 16S rRNA genes), abundance (fluorescent in 

situ hybridization, quantitative PCR), biomass and activity (sulfate reduction, respiration, 

heterotrophy, CO2 fixation, methane production and consumption). In addition, biomass and 

diversity of meio-, macrofauna and megafauna will be conjointly assessed to address thresholds 

of high CO2 and low pH effects on benthic ecosystems. Adequate samples were obtained recently 

from an expedition to Okinawa Trough (SO196, March 2008). This work will also contribute to 

Theme 3 projects on buffering effects of different sediments in calcification/decalcification 

processes and to Theme 1 with the possible detection of epsilonproteobacteria and 

chemolithotrophic bacteria in the naturally enriched habitats, which are the target of the 

experiments in Theme 1.1.1. 

CO2 and pH thresholds in experimental model systems 

Short-term shifts in microbial community diversity, composition and abundance will be 

experimentally investigated as a function of pH, temperature and CO2 variation using flowthrough 

reactors as sedimentary mesocosms. These experiments will also assess changes in 

ecosystem services mediated by microbes such as remineralization, sulfate reduction, methane 

production or consumption. Bioreactors will be filled with standard heterotrophic sediments from 

coastal areas (e.g. long-term research station, Sylt, Germany; Wadden Sea sediments of different 

grain sizes, also see 4.1.1), for which gradients of pH, temperature and CO2 in the porewater will 

be established. Using this approach, different factor combinations will be tested on microbial 

communities, with for instance: a) different levels of CO2 concentration applied to the same 

sediment samples, or conversely, b) the application of one CO2 concentration to different 

sediment types (silicate vs. carbonate), c) Varying levels of heterotrophic activity under constant 

CO2 concentrations, and d) combinations of increasing temperature and decreasing pH. In the 

different experimental setups, abundance, composition and activity of the communities in each 

bioreactor will be determined at different time intervals to assess resistance (extent of the shift 

measured shortly after the end of the experiments), and resilience (observation of shifts after 

long-term incubations). Those experiments will be done in collaboration with 4.1.1, 4.2.1, 4.2.2., 

3.1.2 and 3.4.2 allowing the comparison of different systems.


Effects of acidification on microbial communities associated with multicellular organisms 

A subset of experimental analyses will deal with microbial communities associated with coldwater 

coral surfaces. Using Lophelia pertusa maintained in aquaria we will test how the 

alteration of environmental parameters (pH, CO2 concentration, temperature) in the water column 

may directly affect microbial communities that colonize coral branches. Control treatments 

consisting of dead coral skeletons will be used to assess whether changes are substrate (i.e. 

coral)-specific or not. Those experiments will be done in collaboration with Armin Form (Theme 

3). In close collaboration with projects 4.1.1, 4.1.2 and 4.1.3, shifts in microbial communities 

colonizing the surface of seagrass and macroalgae will be determined in parallel to the respective 

shifting pH and temperature experiments. This integrated strategy will enable a direct observation 

of the coupling between microbial and macrobial shifts in complex ecosystems, and the intensity 

of the respective shifts. 

Schedule 


DNA preparation, fingerprinting, 

cell counts 

Function measurements 

Data analysis, manuscript 

preparation 

Column setup, CO2 effects, 

community analyses 

Sediment effect 


preparation 

DNA preparation, community 

analyses 


preparation 


Longterm CO2 effects on microbial diversity, biomass and function in 

the seabed 

CO2 and pH thresholds in experimental model systems 

Effects of acidification on microbial communities associated with 

multicellular organisms 

233


234 


- Data on diversity and functional shifts in natural CO2 reservoirs month 10 

- Set up of flow-through columns month 12 

- Data on CO2 effects on communities month 16 

- Data on response of different sediment types month 19 

- Data on response of coral-associated microbes month 28 




4.1.1 

4.1.2 

4.1.3 

4.1.4 

Subtotal 

Consumables 

4.1.1 

4.1.2 

4.1.3 

4.1.4 

Subtotal 

Travel 

4.1.1 

4.1.2 

4.1.3 

4.1.4 

Subtotal 

Investments 

4.1.1 

4.1.2 

4.1.3 

4.1.4 

Subtotal 

Other costs 

4.1.1 

4.1.2 

4.1.3 

4.1.4 

Subtotal 

Total 



4.1.1 


Personnel costs: Experimental work on various aspects of this sub-project (primary 

producers, enzymatic nutrient recycling, community processes) will be 

conducted contemporaneously by master and diploma students at the 

different facilities of the AWI at Bremerhaven, Helgoland and Sylt and at the 

University of Hamburg. Funding is requested for 1 PostDoc ( ) 

for 36 months who will coordinate and supervise the students’ work and 

assist in thesis preparation. This scientist will be responsible for data 

management and synthesis of the results from different project 

compartments. Experimental work of the PostDoc will encompass herbivore 

feeding experiments and field investigations. 

4.1.2 

Personnel costs: 1 PhD student for 36 months. 

Consumables: Equipment for experiments, reagents, fees for external measurements 

Travel: Visit to other projects within BioAcid, 1 symposium in last year 

Investment: upgrading of existing culturing infrastructure for appropriate heating, 

cooling, pCO2 and irradiation treatment plus, pCO2 measurement techniques 

(e.g. CO2 and pH microsensor systems) 

4.1.3 

Personnel costs: 1 PhD student for 36 months. In the first and second year another 

EUR/year are requested for student workers, assisting in culture work, in 

field and laboratory experiments and routine analytic procedures 

Consumables: Lab chemicals (HPLC grade) for analysis of photosynthetic pigments and 

cellular oxidative state (GSH/GSSG), pre cast gels and primary & secondary 

antibodies for SDS-PAGE and Western Blotting for protein analysis 

Travel: for annual project workshop, conferences and field research. In the second 

year, field research has to be conducted at a tropical field site, which requires 

funding for airplane tickets, freight of scientific equipment, lab fees, local 

field & diving support, and accommodation. In the third year, we will seek to 

present our results in national and international conferences. 

Investment: Oxygen electrodes, CO2 and pH microsensor systems to measure 

photosynthesis & calcification on small spatial scales, 1x Imaging-PAM for 

measuring whole plant photosynthesis with an image analysis system, 

Cryostats and pumps for aquaculture and mesocosm systems. 

4.1.4 

Personnel costs: 1 PhD student for 36 months. 

Consumables: Molecular biology reagents (DNA extraction, purification, quantification, 

PCR enzymes, etc.), including capillary electrophoresis to separate PCR 

amplicons (ARISA, T-RFLP). Consumables for DNA sequencing (done at 

the MPI) are also included. Microbial activity measurements (sulfate 

reduction, respiration, heterotrophy, CO2 fixation, methane production and 

235

236 


consumption) are also included in the estimated costs, as well as 

consumables for microscopic counts (Acridine Orange, Fluorescent In Situ 

Hybridization). 

Travel: One international conference and one BIOACID meeting per year for the 

student 

Investment: Microsensor construction (pH, CO2) will be done at the MPI during the first 

year (collaboration with Dirk de Beer; Theme 3). Flow-through columns and 

related equipment (pump, tubes, etc.) will be purchased in year 1. 

Project 4.2: OA effects on food webs and competitive interactions in pelagic 

ecosystems (M. Boersma) 

i. Objectives 

We will investigate the role of OA shaping competitive interactions and food web dynamics in 

pelagic ecosystems, working on microalgae, their grazers, and associated bacterial communities. 

More specifically our objectives are: 

• To assess the sensitivity of different microalgal species and strains to OA 

• To provide a process-based understanding for the observed responses to OA by applying 

a combination of different techniques and molecular tools 

• To establish the influence of OA on mixed assemblages in competition experiments at 

the intra-population level as well as at the inter species level 

• To assess the consequences of OA on population community structure of key species as 

well as on the nutrient stoichiometry of primary producers and primary consumers 

• To establish the way grazers deal with excess carbon when feeding on food with 

imbalanced C:N:P ratios. We will investigate the reaction of bacterial communities on 

different carbon sources originating from these grazers. 


Species-specific differences in CO2/pH-sensitivity may directly impact phytoplankton succession 

and distribution (Hansen 2002, Rost et al. 2003), as well as competitive interactions between 

organisms. Not only direct sensitivities to CO2/pH as a stress factor influences succession and 

competitive interactions, but also changes in the availability of essential nutrients may affect the 

outcome of competitive interactions for these nutrients (Tilman 1982). Loladze and co-workers 

(2004) took this one step further and showed that even the ratio of different nutrients can be 

regarded as the limiting nutrient, and thus the outcome of competition may even depend on the 

supply ratio of these nutrients. This is particularly relevant as the ratio of the available carbon to 

other nutrients is bound to change as a result of OA, especially in coastal areas where a 

concurrent trend of decreasing nitrogen and phosphorus inputs can be found (Wiltshire et al. 

2008). This results in increasing C:N and C:P ratios of dissolved substances available for 

phytoplankton growth.


These changing growing conditions are particularly relevant for dinoflagellates. Dinoflagellates 

are a diverse and abundant group of protists with complex interactions in the food web. They can 

form ‘red-tides’, and some species/strains are known to produce various toxins with large 

consequences for ecosystems, human health and economy. Given the increasing number and 

intensity of dinoflagellate blooms over the last decades (Hallegraeff 2003), it is remarkable that 

relatively little is known about the underlying causes. It has been suggested that CO2/pH affects 

photosynthesis, growth, and toxin production of phytoplankton (Lundholm et al. 2004, Hansen et 

al. 2007), moreover nutrient (nitrogen) limitation can also affect toxin production of some species 

as well (Parkhill & Cembella 1999), and it could be hypothesized that an increase in the C:N ratio 

of the available resources should lead to a reduction of overall toxicity. Moreover, these different 

environmental conditions could lead to strain or species replacements, with potentially large 

secondary effects on competitors and consumers. 

Not only do we expect species or strain replacements as a result of changing conditions, even in 

genetically identical lineages other aspects affecting the quality as food for higher trophic levels 

might change. Increasing CO2 concentrations will affect the nutrient stoichiometry of primary 

producers. Provided that the levels of nutrients such as Si, N and P do not change, an increase in 

C-uptake by autotrophs should cause a shift in the elemental composition of the primary 

producers, and hence influence their quality as food for their consumers. They will have more C 

relative to the nutrients, at the same time the total biomass of these producers will probably be 

higher as a result of the increased C-availability. In most known cases an increase in the 

C:nutrient ratio of primary producers causes a reduction in their quality as food for the next 

trophic level (Boersma 2000, Malzahn et al. 2007a). Hence, the situation might arise that 

consumers are faced with more food, which at the same time is of inferior quality. The aim of this 

project will be to investigate the effects of these changes in food quality for consumers, whereas 

we expect direct pH effects to be of less importance (Kurihara et al. 2004, Mayor et al. 2007). 

An excess amount of carbon relative to nutrients in algae should lead to an increased excretion of 

carbon by secondary producers. Depending on how this excess C is excreted (DIC/CO2 or DOC) 

this could have major consequences for the flow of energy and matter (Darchambeau et al. 2003). 

The nature of these excretory products has been under considerable debate (Sterner et al. 1998, 

Plath & Boersma 2001, Darchambeau et al. 2003, Jensen & Hessen 2007), but still has not been 

answered. DOC is potentially taken up rapidly by bacteria, which might benefit from increased 

C:nutrient levels in algae, but virtually nothing is known here (Sterner et al. 1998, Riebesell et al. 

2007). Since bacteria are homeostatic and “more like animals than plants” with respect to their 

stoichiometry (Makino et al. 2003), changes in the elemental composition of excretory products 

should cause shifts in bacterial community composition. To date, the main focus of the studies on 

the response of bacterial communities has been direct, i.e. the direct effects of acidification on 

diversity, and there seem to be no large effects (Allgaier et al. 2008). We have no idea on the 

indirect effects, and the consequence of these population shifts due to “imbalanced nutrients” for 

functional diversity and biogeochemical cycles are still unknown 

Hence, potentially, the biological production of the world-seas will increase in the first trophic 

level as a result of higher CO2 concentrations, but the second trophic level might not be able to 

benefit from this increased biomass. 


4.2.1 Maarten Boersma has considerable experience with experimental plankton ecology at 

many different scales, ranging from laboratory experiments on individual organisms to 

large-scale laboratory and field enclosures. Moreover, he is an expert on direct 

237

238 


manipulations of components of the food in algal-grazer interactions (Boersma 2000, 

Boersma & Elser 2006, Boersma et al. in press). Arne Malzahn is a fisheries biologist 

with expertise in experimental and field approaches in larval fish and plankton ecology. 

His main research interests are ecological stochiometry and trophic transfer. He was the 

first to show that limitations on the primary producer level can be transfered through 

primary consumers to secondary consumers (Aberle & Malzahn 2007, Malzahn et al. 

2007a, Malzahn et al. 2007b). Gunnar Gerdts is an expert on bacterial community 

analysis of different habitats and pro-eukaryotic consortia. He has managed to investigate 

bi-trophic systems of microalgae amended with 13 C-carbonate and associated bacterial 

community feeding on 13 C containing algal exudates using stable isotope probing, which is 

the first time that this approach has been applied to marine pelagic systems. 

4.2.2. Björn Rost has studied the carbonate chemistry effects on marine phytoplankton over the 

last decade, focusing on physiological key processes such as photosynthesis, carbon 

acquisition and calcification (Rost et al. 2003, Rost & Riebesell 2004). He is an expert on 

membrane-inlet mass spectrometry (MIMS), which allows monitoring gas exchange 

processes in real-time under different environmental conditions, a technique that is central 

for the planned experiments (Rost et al. 2007). Uwe John’s research emphasis is on the 

molecular regulation of toxin synthesis and growth regulation of marine protists, 

combining ecological and physiological experiments with functional genomics to reveal 

the mechanisms behind algal bloom formation (Cembella & John 2006, Tillmann et al. 

2007, John et al. 2008). He has established several assays for the estimation of the 

molecular response of microalgae to environmental stressors using standard protein assays 

and microarrays/quantitative PCR (qPCR), respectively. 


Subproject 4.2.1: OA effects on pelagic community structure and food chains 

M. Boersma, A. Malzahn, & G. Gerdts 

One of the main questions targeted by this project is as to the effects of high CO2 induced 

differences in algal quality on herbivores and bacteria. Furthermore, food with a high C:nutrient 

should lead to an increase in the excretion of carbon by consumers, but to date there is no 

information on these processes. The high CO2 world will be realized by culturing algae under 

elevated CO2-concentrations, relevant to the different IPCC scenarios, and agreed in framework 

of BIOACID. These algae will be fed to grazers. It is important to note that we will culture the 

algae separately, harvest them and then feed them to the grazers at ambient pCO2. This is 

important, as only in this way, we can guarantee that we are investigating food effects only, and 

not direct effects of CO2/pH on the grazers, which will be dealt with in 2.1.2 (in cooperation with 

this sub project) 

We will carry out laboratory experiments with different algae-grazer systems, as it might well be 

that different grazers have different mechanisms to void excess C, using at least partly identical 

algal species/strains as the ones used in subproject 4.2.2. Therefore, in these experiments we will 

assess the reaction of the grazers on the food sources of different quality in terms of nutrient 

stoichiometry by analysing growth and reproduction of the consumer, coordinating techniques 

with 4.1.1 and 4.1.2. At the same time we will measure respiration rates and DOC production to 

better understand the relevance for the ecosystem. The two most important mechanisms being


respiration (CO2-production), and release as dissolved organic carbon. Whether one pathway or 

the other is chosen has large consequences for the system as DOC is available for heterotrophic 

bacterial production, CO2 for autotrophic bacteria only. 

We will then proceed to investigate the bacterial community as well as the bacterial growth rates 

and activities related to these different conditions. We hypothesise that bacterial activity will be 

much higher in those cases that excess carbon is voided as DOC, and that bacterial diversity will 

be different, for example by favouring those groups able to fix nitrogen. To investigate the effects 

of different C:N:P ratios on bacterial populations as well as the influence of excreted DOC from 

grazers, a state of the art methodological stable isotope probing approach will be applied 

(Manefield et al. 2002, Radajewski et al. 2003, Sapp et al. 2008). Selected microalgae will be 

incubated with different 13 CO2 levels under defined N:P ratios and fed to grazers. Microalgal 

exudates and grazer excretes (filtrates) will be incubated with natural marine bacterial 

communities. Bacterial RNA or DNA will be extracted and “heavy” ( 13 C-containing) RNA or 

DNA will be separated from “light” ( 12 C-containing) ribo/nucleic acid molecules by isopycnic 

ultracentrifugation and gradient fractionation. Collected fractions will be analyzed for 13 C/ 12 C 

isotope ratios by mass spectrometry. RNA will be transcribed to DNA by RT-PCR (Reverse 

Transcriptase PCR) and analyses of the bacterial community will be performed by PCR with 

(group) specific primers, followed by DGGE (Denaturing Gradient Gel Electrophoresis), ARISA 

(Automated ribosomal intergenic spacer analysis) or DHPLC (Denaturing high performance 

liquid chromatography) and 16S-rDNA sequencing of major bands. 

The outcome of the experiments are of quantitative and qualitative nature: 

a) Identification of those bacterial populations which benefit from enhanced carbon levels 

in the coastal marine system or which are negatively affected. 

b) Estimation of bacterial community compositions in conjunction with different C:N:P 

ratios of bacterial nutrients 

All molecular analyses will be performed in close cooperation with subproject 4.1.4, as well as 

with the subprojects in theme 1 [1.2.1-1.2.4]. In contrast to the subprojects in Theme 1, we will 

primarily focus on the analysis of shifts in bacterial community composition due to changes in 

stoichiometry of algal exudates and excretes of grazers. In Theme 1, the stability of algal 

exudates to hydrolysis in dependence of the water pH will be examined, which is a functional 

response. We will examine if certain C:N:P ratios of "bacterial food" caused by elevated pCO2 in 

the ecosystem will result in specific community compositions, which can be defined as a 

structural response. Molecular techniques will be standardized and fingerprint-data will be shared 

using advanced database software. 

In a next step, we will use laboratory microcosms to investigate system effects. An increase in 

bacterial activity should lead to increased micrograzers which are a good food source for 

zooplankters, thus fuelling the microbial loop. The great advantage of a stronger microbial loop 

for e.g. copepods will be that much of the excess carbon has already been respired by the 

intermediate trophic levels, which should cause the micro grazers to be a better quality food. For 

this we will use set-ups that are in the size of 10-50 l, stocked with different algae-grazer 

combinations and inocula of natural seawater to enter a microbial community component to the 

system. 

239

Schedule 

240 



Set-up of culturing facility, 

instrument calibration 

Collection and rearing of organisms 

Algal-Grazer-Bacteria experiments 

Cell counts, respirometry & 

chemical analyses 

DNA/RNA extraction and 

preparation (SIP) 

Molecular fingerprints & 

sequencing 

Microcosm experiments 



Manuscript preparation, presentation 

of results at conferences 




- Experimental data set on CO2/pH sensitivity of grazer-algal interactions month 15 

- Data set on bacterial diversity and functional shifts month 24 

- Data set on microcosm experiments month 30 


Subproject 4.2.2: Competitive interactions in planktonic microalgae under OA-stress 

B. Rost & U. John 

We aim to examine the responses of different dinoflagellate species and strains to multiple 

abiotic and biotic stressors. Applying a combination of different techniques and molecular tools 

will provide a process-based understanding for the observed responses. In addition to 

experiments with mono-clonal incubations, competition experiments will be conducted under 

respective conditions to investigate the influence of stressors on the intra-specific level (i.e. 

population) and species interaction. The project will strongly improve our predictive capabilities 

on how different dinoflagellate species as well as mixed assemblages will respond to ocean 

acidification. Findings about competitiveness and species/strain interaction of dinoflagellates 

under the ocean acidification scenarios will be used for general comparison within theme 4 

[4.1.1, 4.1.2, 4.1.3, 4.1.4, 4.2.1]. 

Representative species of marine dinoflagellates and corresponding strains will be grown under 

different pCO2 levels, representing last glacial maximum (180 ppm), present-day (380 ppm) and 

those predicted for the future (750-1000 ppm). Since other environmental factors, such as light


and nutrient availability, are also changing and possibly modifying the CO2/pH-related responses 

interactive effects are investigated in a matrix approach. In these acclimations various parameters 

like growth rates, photosynthesis, elemental ratios (C:N:P), photosynthetic pigments or toxin 

production/toxicity (Tillmann & John 2002) are determined. These experiments will be 

performed with mono-clonal cultures of species/strains. Data and samples will be provided to 

1.2.2 (for comparative analysis of C:N:P stoichiometry), to 1.2.1 (for analysis of extracellular 

organic material), to 4.2.1 (for feeding experiments) and to 3.5.3 (for morphological examination 

of calcifying dinoflagellates). Subsequently, competition experiments will be conducted under 

different pCO2 scenarios using strains and later species assemblages that responded differently to 

the respective treatments. Because strains and some species cannot be differentiated 

morphologically in such mixed assemblages, molecular markers such as AFLP and 

microsatellites (John et al. 2004, Alpermann et al. 2006) will be used to qualitatively and 

quantitatively distinguish species/strains and hence assess the success of certain 

pheno/genotypes. 

To get a process-based understanding of the responses to changes in CO2/pH different bio-assays 

(in vivo) have been developed at the AWI. The membrane-inlet mass spectrometry (MIMS) 

allows monitoring gas exchange processes in real-time under different environmental conditions 

(experience exchange and data discussion with 3.1.1). One application allows distinguishing 

between CO2 and HCO3 - as carbon sources and determines the fluxes as function of 

concentration. In other applications, the use of stable isotopes enables to measure activities of 

carbonic anhydrase (CA) or photosynthetic electron fluxes (ETR). The advantage of our MIMS 

approach is that several processes can be observed and quantified simultaneously, even with 

sensitive species such as dinoflagellates (e.g. Rost et al. 2006). These methods unravel the carbon 

and energy fluxes, a prerequisite to understand the CO2/pH-dependence in photosynthesis and 

other down-stream processes. To further advance our process-understanding the expression of 

proteins and genes of key enzymes (e.g. RubisCO, PKS) will be quantified using standard protein 

assays and microarrays/quantitative PCR (qPCR), respectively. Those methods including Oligonucleotide 

DNA microarrays will provide, in combination with detailed physiology, a 

comprehensive mechanistic overview of the cascade from gene expression to physiological 

responses. 

Schedule 


Experimental set up 

Phenotypic characterisation/ MIMS 

Genotypic characterisation 

Interspecific competition 

Intraspecific competition 

Data evaluation and publication 


241


242 



- Data on species/strain-specific sensitivity towards stressors month 24 

- Competition experiments concluded month 30 

- Evaluation of influence of species/strain success due to stress tolerance month 36 



4.2.1 

4.2.2 

Subtotal 

Consumables 

4.2.1 

4.2.2 

Subtotal 

Travel 

4.2.1 

4.2.2 

Subtotal 

Investments 

4.2.1 

4.2.2 

Subtotal 

Other costs 

4.2.1 

4.2.2 

Subtotal 

Total 


4.2.1 


Personnel costs: 2 PhD students, one candidate will perform the grazing experiments and 

investigate the effects of different conditions on growth, reproduction and 

excretory processes of grazers, the other one will investigate the responses of 

the bacterial community on the differing conditions as a result of these 

different grazing conditions. 

Consumables: Molecular biology reagents (DNA extraction, purification, quantification, 

PCR enzymes, etc.), including electrophoresis reagents and consumables to 

separate PCR amplifons (DGGE, ARISA, DHPLC). Consumables for DNA 

sequencing are included. Costs for isotope ratio measurements and 

consumables for determination of C,N & P and DOC.


Travel: Covers the attendance of the annual project workshops and one conference 

per year for the Ph.D. students. 

4.2.2 

Personnel costs: 1 PhD student for 36 months. The candidate will perform experiments on 

physiological performance and competitive success of different 

dinoflagellate species and strains as outlined in the proposal. 

Consumables: Lab chemicals for culturing and in vivo assays (e.g. MIMS). Lab chemicals 

(HPLC grade) for analysis of secondary metabolites, pre cast gels and 

primary & secondary antibodies for SDS-PAGE and Western Blotting for 

protein analysis. Kits for RNA and DNA extraction, microarrays and the 

hybridisation chemistry. Consumables for PCR and qPCR. 

Travel: Covers the attendance of the annual project workshops and one conference 

per year for the Ph.D. student. 


Aberle N, Malzahn AM (2007) Interspecific and nutrient-dependent variations in stable isotope fractionation: experimental studies simulating 

pelagic multitrophic systems. Oecologia 154:291-303 

Allgaier M, Riebesell U, Vogt M, Thyrhaug R, Grossart HP (2008) Coupling of heterotrophic bacteria to phytoplankton bloom development at 

different pCO2 levels: a mesocosm study. Biogeosciences Discuss 5:317-359 

Alpermann TJ, John UE, Medlin LK, Edwards KJ, Hayes PK, Evans KM (2006) Six new microsatellite markers for the toxic marine 

dinoflagellate Alexandrium tamarense. Mol Ecol Notes 6:1057-1059 

Asmus H, Asmus R (eds) (2000) ECSA-Workshop on Intertidal Seagrass Beds and Algal Mats: Organisms and Fluxes at the Ecosystem Level, 

Vol 54 (2/3) 

Baird D, Asmus H, Asmus R (2004) Energy flow of a boreal intertidal ecosystem, the Sylt-Romo Bight. Mar Ecol Prog Ser 279:45-61 

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11.6: Theme 5: Integrated assessment – sensitivities and uncertainties 



German Advisory Council on Global Change (WBGU, Berlin 2006) states: “Because of the 

importance of the consequences of ocean acidification, research in this area should be intensified 

considerably. As long as there is no general scientific consensus about the tolerable limit for the 

effects of acidification, a margin of safety according to the precautionary principle should be 

observed. The suggestion of the WBGU to prevent a pH decrease of more than 0.2 is oriented 

toward the goal of avoiding an aragonite undersaturation in the ocean surface layer. If it is found 

that other intolerable damages already occur before reaching aragonite undersaturation, then the 

guard rail will have to be adjusted accordingly.” 

As stated in the report, the tolerable window for ocean acidification defined by WBGU presently 

relies on an extremely small data base. In fact, rather than using the limited data on observed 

biological consequences of ocean acidification, the WBGU reaches its recommendation on the 

basis of projected changes in water chemistry (aragonite saturation state). While this is an 

appropriate approach in view of the scarcity of biological information, there is a clear need to 

establish a reliable data base on tolerance levels for ocean acidification in key groups of oceanacidification 

sensitive marine organisms in order to reach a more informed recommendation. 

Theme 5 of BIOACID will take the challenge of integrating the information gained under 

Themes 1 to 4 in order to identify the potential thresholds associated with ocean acidification. 

Uncertainties, probabilities and risks to the marine environment have to be assessed as well as 

their feedback to climate systems. During the first 3-year phase of BIOACID, our main aim is to 

develop and establish the tools that will allow us fulfil the BIOACID synthesis needs. For the 

three subprojects proposed here, the synthesis tools to be established within BIOACID range 

from meta-analysis techniques, over regional and global numerical ecosystem models to 

economic methods of integrated assessment. These tools will help to better understand ongoing 

changes in chemical and biological state of the North Sea from alkalinity fluxes originating from 

the Wadden Sea over a synthesis model that integrates OA sensitivities at organism level into a 

North Sea ecosystem model (5.1) to an economical impact assessment. (5.3). Newly developed 

assessment tools will also be used to improve parameterisations of calcium carbonate production 

in global biogeochemical climate models (5.2). By investigating the combined effects of 

variations in temperature and ocean acidity, such parameterisations will allow to put better 

constraints on possible threshold levels on ocean acidification in a warming world. 

Objectives 

• Synthesize information obtained in Themes 1 to 4 to achieve an integrated understanding of 

biological responses to ocean change, integrating effects of ocean acidification and warming 

• Develop a framework for integrating ocean acidification sensitivities at the organism level into 

ecosystem modelling 

• Identify critical threshold levels (‘tipping points’) of ocean acidification for irreversible 

ecosystem changes providing sound information for adaptation and mitigation measures 

• Define dangerous ocean acidification in terms of the goods and ecosystem services lost 

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5.1 Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle and the primary 

production in the North Sea. 

The central objective is to relate the Wadden Sea’s AT flux to the pelagic and benthic 

environment, with emphasis on biogeochemical cycling of carbon in conjunction with other 

nutrients, such as nitrogen. The impact of the AT flux from the Wadden Sea on the general North 

Sea carbon cycling. and on the long-term trend with respect to the overall pH signal within the 

North Sea will be investigated. A future scenario model simulation will be performed and 

analysed with respect to critical ecological and biogeochemical regime shifts, as we impose an 

atmospheric partial pressure (pCO2) of 1000 µatm. 

5.2. Evaluating and optimising parameterisations of pelagic calcium carbonate production in 

global biogeochemical ocean models. 

The aim is to critically review existing parameterisations of calcification currently used in largescale 

biogeochemical climate models and to compile published experimental findings on 

temperature- and pCO2-sensitivities of calcium carbonate production, its export and dissolution. 

The ability of current parameterisations of calcification in biogeochemical models to reproduce 

observed alkalinity fields will be quantitatively assessed and a Bayesian meta-analysis will be 

applied to BIOACID experimental findings in order to condense distributed information into 

plausible model parameterisations. 

5.3. Viability-method for the impact assessment of ocean acidification under uncertainty - 

Development of the method and exemplary application to the impact of acidification on the North 

Sea cod fishery. 

This project will further develop the ecological-economic viability-method towards a general 

approach for integrated assessment of human actions influencing ocean acidification and the 

consequences for human well-being that takes uncertainties about future development into 

account. 

The usefulness of the viability-method will be demonstrated by applying it exemplarily to the 

assessment of acidification effects on the North Sea cod fishery.


Project 5.1: Impact of Alkalinity fluxes from the Wadden Sea on the carbon cycle 

and the primary production in the North Sea 

PI: Johannes Pätsch (1), Co-PIs Helmuth Thomas (2), Markus Schartau (3) 

(1) Institute of Oceanography, Hamburg; (2) Dalhousie University, Halifax, NS, Canada; (3) 

GKSS Forschungszentrum Geesthacht 

i. Objectives 

Recent findings suggest that the Wadden Sea at the southeastern North Sea acts as a major source 

of total Alkalinity (AT) (Thomas et al., subm.). This will affect the carbon cycling within the 

North Sea and is expected to have an impact on the overall primary production in pelagic and 

benthic systems. During summer the AT release lowers the marine partial pressure of carbon 

dioxide (pCO2) and the CO2 release to the atmosphere, in particular in the southern bight of the 

North Sea. This AT flux buffers the pH decline, induced by anthropogenic CO2, as already 

observed in the area of the North Sea (Thomas et al., 2007). Yet, the extent to which the AT flux 

from the tidal flat areas of the Wadden Sea can buffer pH and its response to the biological 

ecosystem has to be assessed. 

The central objective is to relate the Wadden Sea’s AT flux to the pelagic and benthic 

environment, with emphasis on biogeochemical cycling of carbon in conjunction with other 

nutrients, such as nitrogen. With our modelling activity within BIOACID we specifically propose 

the following: 

• We will use an existing ecosystem of the North Sea (ECOHAM) to extrapolate 

information’s gathered from local observations to the entire North Sea. 

• We will quantify the impact of the temporally resolved AT flux from the Wadden Sea on 

the general North Sea carbon cycling. 

• We will investigate to which extent this AT flux affects the long-term trend with respect 

to the overall pH signal within the North Sea. 

• The response of primary producers to pH changes will be addressed by extrapolating 

results from mesocosm and chemostat experiments in an attempt to crudely discriminate 

non-calcifying from calcifying primary producers. 

• A future scenario model simulation will be performed and analysed with respect to 

critical ecological and biogeochemical regime shifts, as we impose an atmospheric partial 

pressure (pCO2) of 1000 µatm. 

The proposed modelling activities integrate very well with the BIOACID-investigations of Prof. 

Dr. Böttcher and Prof. Dr. Liebezeit on biogenic carbonates in the Wadden Sea. In addition, we 

will consider characteristic parameter values of phytoplankton productivity that are derived from 

data measured under different growth conditions by Dr. Engel and her group. This will provide 

an excellent basis for assessing our modelling approach of “excess production” (Pätsch & Kühn, 

2008) and “carbon overconsumption” (Schartau et al., 2007), which refers to the description of 

carbon fixation under nutrient limited conditions (Fogg, 1983). 

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Continental shelves play a key role in the global cycling of biogeochemically essential elements 

(Christensen, 1994; Jickells, 1998). From observations in the East China Sea, Tsunogai et al. 

(1999) concluded that the global shelves act as a sink for atmospheric carbon dioxide and as a 

source of carbon for the ocean (Borges et al., 2005). The North Sea, as part of the Northwest- 

European shelf, has been characterized as a sink for atmospheric CO2 (Thomas et al., 2004). 

These findings were confirmed by simulations, using the ecosystem model ECOHAM (Pätsch & 

Kühn, 2008) during a Diploma study (Prowe, 2006). The dependency of the carbon fluxes in the 

North Sea on the Alkalinity especially on the Alkalinity produced in the Wadden Sea is under 

debate (Thomas et al., in prep). For the southern North Sea a modelling effort was successful 

focusing on the effect of phytoplankton blooms on the carbonate system (Gypens et al., 2004). 

On large scales, the multi-functional plankton modelling approach needs to be gradually and 

slowly substantiated rather than precipitated; from this perspective, we will coordinate our 

modelling activity during the proposed project in collaboration with our neighbor Institute of 

Hydrology and Fisheries (Diekmann, et al., submitted), Institute for Coastal Research at GKSS, 

and the Alfred Wegener Institute (Dr. Engel and her group) within BIOACID. 


Johannes Pätsch has worked since two decades in the field of numerical modelling. In the last 

years he has applied (Pätsch & Radach, 1997) and developed (Pätsch, et al. 2002) 

biogeochemical models for different marine environments mainly shelf seas. He has gathered 

experience in plankton dynamics and their mathematical expression in complex ecosystem 

models during the European MAST project ERSEM (1990-1997). He has also applied statistical 

methods for large data sets (Radach & Pätsch, 1997). The main focus of his work is on the 

interplay between biogeochemical fluxes of carbon and nitrogen in various marine environments 

(Pätsch et al., 2002; Pätsch & Kühn, 2008). 

Helmuth Thomas has worked since many years in the field of carbon dynamics in marine 

biogeochemical environments. Using field data sets, which resolve the North Sea system in time 

and space, he established the first annual carbon budget of this shelf sea (Thomas et al., 2004, 

2005). Furthermore he characterized the North Sea as a shelf sea which is highly vulnerable to 

acidification processes (Thomas et al., 2007). 

Markus Schartau has gathered experience in optimisation of ecosystem models based on 

sophisticated data-assimilation methods (Schartau et al., 2001; Schartau and Oschlies, 2003). He 

has participated in international projects that address model assessment and validation, but also 

the modelling of functional plankton groups (e.g. Friedrichs et al, 2006; Hood et al., 2006). 

During the last years his work focused on the development of biological models with variable 

stoichiometry, with emphasis on carbon overconsumption in conjunction with abiotic organic 

particle formation (Schartau et al., 2007). He is involved as principle investigator in the two 

collaborating projects EPOCA (European Project on Ocean Acidification) and SOPRAN (Surface 

Ocean Processes in the Anthropocene).



• 5.1.a: We will simulate the biogeochemistry of the North Sea with the ecosystem model 

ECOHAM (Pätsch & Kühn, 2008) for recent years with prescribed Alkalinity fields (reference 

run). The model includes a bulk formulation for calcifying phytoplankton. In order to improve 

the already existing bulk parametrization of calcification this mechanism and the pCO2 

sensitivities will be compared cautiously with M. Hoppema’s approach in project 3.4.3 

(Buffering ocean acidification: Dissolution of carbonate sediments in the Southern Ocean). 

• 5.1.b: In cooperation with the Institute of Hydrobiology and Fishery (Uni-Hamburg) and the 

BIOACID-community we will perform cross-validation experiments with our already existing 

plankton module, while separating between the three phytoplankton groups: calcifiers, silicifiers 

and flagellates. The model-system with this plankton module will also be tested against the 

reference run. 

• 5.1.c: Results from experiments of A. Engel; project 1.2.1 (Production and decomposition of 

exudates) will allow us to refine model parameterisations with respect to the production of 

dissolved organic matter and its dependence on pH, temperature, and nutrient availability. Data 

from M. and G. Nausch; project 1.2.3 (DOM availability and phosphorus utilization) will also 

be regarded. All implementations and parametrizations, which concern the cycling of DOM, will 

be aligned with the developments in the project 1.3 of B. Schneider (Modelling biogeochemical 

feedbacks of the organic carbon pump). 

• 5.1.d: In cooperation with the group of M. Böttcher; project 3.4.1 (Impact of biogenic 

carbonates on pH buffering in an acidifying coastal sea (North Sea)) we will estimate the 

recent temporally resolved flux of AT from the Wadden Sea into the open North Sea during low 

tides. During high tides when the water is flushing the Wadden Sea we will provide the simulated 

pH, as well as DIC and Alkalinity concentration to this group. The described boundary data will 

be used to run our model with prognostic Alkalinity dynamics. For further model confirmation, 

the GKSS will provide data from “ships of opportunity” (fluorescence, pH, temperature, salinity). 

This data set can be updated in the near future (during BIOACID funding period) with pCO2 

Ferry Box measurements, in collaboration with Prof. Friedhelm Schroeder (GKSS). At the 

beginning of BIOACID, the sensors for pCO2 will not be in operational use and will also need to 

be calibrated. Until now the carbon budget of the Wadden Sea is not fully understood. Therefore 

the investigations by R. Asmus, project 4.1.1 (Effects of ocean acidification on trophic 

interactions in coastal seaweed and seagrass ecosystems) are of great importance for our 

studies. 

• 5.1.e: In a first sensitivity study we will vary the amount of AT flux from the Wadden Sea, in 

order to study the pH variation of the North Sea.. The impact of these pH variations will be 

investigated with respect to 1) carbon fluxes in the North Sea and 2) primary production in 

conjunction with the production of dissolved organic matter. 

• 5.1.f: In a second sensitivity study we will run the model with an atmospheric pCO2 of 1000 

atm. The impact of the corresponding pH changes on different primary producers will be 

examined. We hypothesise a significant response in primary production that translates into 

modifications in the overall carbon flux in the North Sea and across the boundaries of the North 

Sea. The latter results can be identified as the exchange between the North Atlantic (open ocean) 

and the Northwest European Shelf. We will compare the dynamics of this explicitly resolved 

fluxes with those parameterised in global modelling approaches (project 5.2: Evaluating and 

optimising parameterisations of pelagic calcium carbonate production in global 

biogeochemical ocean models). 

• 5.1.g: The results will be documented by an article within an international journal. 

251

Work Schedule 

252 



5.1.a 

5.1.b 

5.1.c 

5.1.d 

5.1.e 

5.1.f 

5.1.g 



- Establishing and evaluation of the reference run 3 months 

- Implementation of the new plankton module 18 months 

- Exchange of data with other institutions 27 months 

- Sensitivity study 1 30 months 

- Sensitivity study 2 33 months




PhD (1/2 job) 

Three months Post-Doc 

(Markus Schartau, GKSS) 

Subtotal 

Consumables 

1 Personal Computer for the 

PhD student 

Travel 

Uni Hamburg: 

EGU meetings 

Uni Hamburg: 

BIOACID meetings 

Uni Hamburg: 

Cooperation Halifax 

GKSS: EGU meetings 

GKSS: 

BIOACID meetings 

GKSS: 

Cooperation Halifax 

Subtotal 

Investment 

Other costs 

Total Uni Hamburg 

Total GKSS 

TOTAL 



Personnel costs: The University of Hamburg will employ a PhD student on a half time position. 

This student will be supervised by Johannes Pätsch and Markus Schartau. Markus Schartau will 

give substantial advice and support in data assimilation and parameter optimisation techniques in 

conjunction with the proposed model development. Since his contribution is not part of the 

Helmholtz Association (HGF) research programme (PACES), one post-doc monthly salary will 

be allocated to the GKSS per year. 

Consumables: 1 PC: The employee needs access to our computer center. Simulation results will 

be evaluated with the PC. 

Travel: Results from the proposed BioAcid model simulations will be presented at EGU 

meetings. Different aspects of the here proposed topics are likely to be addressed at the annual 

253

254 


meetings, allowing for presentation that are given not only by the PhD student but also by the 

advisors. The PhD student is expected to gain international experience in the field of 

biogeochemical modelling. The EGU meetings are a prominent place to learn this item. 

For cooperation with Dalhousie University we intend to have at least one meeting per year, either 

in Halifax, or in Hamburg. Travel money for GKSS partner will be allocated in order to join at 

least one abroad meeting in Halifax, Canada. 


Anderson TR (2005) Plankton functional type modelling: running before we can walk? Journal of Plankton Research 27(11): 1073-1081 

Borges AV, Delille B, Frankignoulle M (2005) Budgeting sinks and sources of CO2 in the coastal ocean: Diversity of ecosystems counts. 

Geophys Res Let 32 L14601 doi:10.1029/2005GL023053 

Christensen JP (1994) Carbon export from continental shelves, denitrification and atmospheric carbon dioxide. Continental Shelf Research 14: 

547-576 

Fogg GE (1993) The ecological consequence of extracellular products of phytoplankton photosynthesis. Bot Mar XXVI: 3-14 

Gypens N, Lancelot C, Borges AV (2004) Carbon dynamics and CO2 air-sea exchanges in the eutrophied coastal waters of the southern bight 

of the North Sea: a modelling study. Biogeoscience 1: 147-157 

Hood RR, Laws EA, Moore, JK, Armstrong RA, Bates N, Brown C, Carlson C, Chai F, Doney SC, Ducklow H, Falkowski P, Feely RA, 

Friedrichs M, Landry M, Nelson D, Richardson T, Salihoglu B, Schartau M, Wiggert J (2006) Functional Group Modelling: Progress, 

challenges and prospects. Deep Sea Research II 52: 459-512 

Jickells TD (1998) Nutrient biogeochemistry of the coastal zone. Science 281: 217-222 

Friedrichs MAM, Dusenberry J, Anderson L, Armstrong R, Chai F, Christian J, Doney SC, Dunne J, Fujii M, Hood R, McGillicuddy D, 

Moore K, Schartau M, Spitz YH, Wiggert J (2006). Assessment of skill and portability in regional marine biogeochemical models: The 

role of multiple phytoplankton groups. Journal of Geophysical Research available: http://www.ccpo.odu.edu/RTBproject/Publications 

Pätsch J, Kühn W (2008) Nitrogen and carbon cycling in the North Sea and exchange with the North Atlantic - a model study, Part I. Nitrogen 

budget and fluxes. Continental Shelf Research 28: 767-787 

Pätsch J, Kühn W, Radach G, Santana Casiano JM, Gonzalez Davila M, Neuer S, Freudenthal T, Llinas O (2002) Interannual variability of 

carbon fluxes at the North Atlantic station ESTOC. Deep-Sea Res II 49(1-3): 253-288 

Pätsch J, Radach G (1997) Long-term simulation of the eutrophication of the North Sea: temporal development of nutrients, chlorophyll and 

primary production in comparison to observations. Journal Sea Research 38: 275-310 

Prowe F (2006) Simulating and budgeting the carbon fluxes in the North Sea (2001/2002). Diploma Thesis University of Oldenburg: 1-128 

Radach G, Pätsch J (1997) Climatological annual cycles of nutrients and chlorophyll in the North Sea. Journal of Sea Research 38: 231-248 

Schartau M, Engel A, Schröter J, Thoms S, Völker C, Wolf-Gladrow D (2007) Modelling carbon overconsumption and the formation of 

extracellular particulate organic carbon. Biogeosciences 4: 433-453; open access: www.biogeosciences.net/4/433/2007/bg-4-433- 

2007.html 

Schartau M, Oschlies A (2003) Simultaneous data-based optimisation of a 1D-ecosystem model at three locations in the North Atlantic Ocean: 

Part 1) Method and parameter estimates. Journal of Marine Research 61(6): 765-793 

Schartau M, Oschlies A, Willebrand J (2001) Parameter estimates of a zero-dimensional ecosystem model applying the adjoint method. Deep 

Sea Research II 48: 1769-1800 

Thomas H, Prowe AEF, van Heuven S, Bozec Y, de Baar HJW, Schiettecatte L-S, Suykens K, Kone M, Borges AV, Lima ID, Doney SC 

(2007) Rapid decline of the CO2 buffering capacity in the North Sea and implications for the North Atlantic Ocean. Global Biogeochem. 

Cycles 21 doi:10.1029/2006GB002825 

Thomas H, Bozec Y, de Baar HJW, Elkalay K, Frankignoulle M, Schiettecatte L-S, Kattner G, Borges AV (2005) The carbon budget of the 

North Sea. Biogeoscience 2: 87-96 

Thomas H, BozecY, Elkalay K, deBaar HJW (2004) Enhanced open ocean storage of CO2 from shelf sea pumping. Science, 304, 5673: 1005- 

1008 

Thomas H, Schiettecatte L-S, Suykens K, Kone YJM, Bozec Y, de Baar HJW, Borges AV (in prep.) Anaerobic oxidation of organic matter - a 

major sink for atmospheric CO2 in the coastal ocean 

Tsunogai S, Watanabe S, Sato T (1999) Is there a 'continental shelf pump' for the absorption of atmospheric CO2? Tellus 51B: 701-712


Project 5.2: Evaluating and optimising parameterisations of pelagic calcium 

carbonate production in global biogeochemical ocean models 

PI: A. Oschlies; Co-PIs: I. Kriest, W. Koeve; IFM-GEOMAR Kiel 

i. Objectives 

We propose 

- to critically review existing parameterisations of calcification currently used in largescale 

biogeochemical climate models, 

- to compile published experimental findings on temperature- and pCO2-sensitivities of 

calcium carbonate production, its export to the interior of the oceans and its dissolution, 

and to put these findings in perspective with the model parameterisations, 

- to quantitatively assess the ability of current parameterisations of calcification in 

biogeochemical models to reproduce observed alkalinity fields, and to suggest improved 

parameterisations that can reliably predict the response of pelagic calcium carbonate 

production to variations in both temperature and ocean carbonate chemistry, 

- to initiate a Bayesian meta-analysis of BIOACID experimental findings 


Two aspects of global change, warming and acidification of the ocean, will most likely affect 

future global calcification rates and thereby influence the ability of the ocean to sequester 

anthropogenic CO2. Current biogeochemical climate models use various, often pragmatic, 

formulations of the production of particulate inorganic carbon. Parameterisations of the 

sensitivity of calcification to acidification include nonlinear dependencies on the CaCO3 

saturation state (Gehlen et al., 2007; Ridgewell et al., 2007), or linear relationships with pCO2 

(Heinze, 2004). While these parameterisations all predict a decrease of calcification in response 

to acidification, the models also include responses to temperature changes. These are accounted 

for either explicitly or implicitly via (a) temperature dependent photosynthesis, in conjunction 

with a linear coupling of autotrophic calcification (e.g., Archer et al., 1998), (b) direct 

temperature dependence of the calcification rate (e.g., Maier-Reimer, 1993; Archer et al., 2000, 

Aumont and Bopp, 2006), or (c) nonlinear effects between the temperature dependent growth and 

loss rates of phytoplankton functional types, including coccolithophores (Gregg and Casey, 2007; 

Aumont and Bopp, 2006). The combined effects of warming and acidification are uncertain and, 

in current climate models, have been shown to result either in a simulated decrease (Gehlen et al., 

2007) or increase (Schmittner et al., 2008) of calcification over the next centuries. 


Prof. Andreas Oschlies is a biogeochemical modeller with extensive experience in simulating 

the interaction of physical transport and biotic processing of biogeochemical tracers at basin and 

global scales. Data assimilation methods have been used to optimise the difficult-to-constrain 

parameters of marine ecosystem models (Schartau and Oschlies, 2003a,b; Oschlies and Schartau, 

255

256 


2005), and mechanistic models have been developed to improve descriptions of nutrient uptake 

and particle export (Kriest and Oschlies, 2007, 2008). 

Dr. Iris Kriest is a biogeochemical modeller with particular interest in large scale particle flux 

and particle dynamics (Kriest and Evans, 1999; 2000; Kriest, 2002; Kriest and Oschlies, 2008). 

Recently she implemented, in close collaboration with Dr. S. Khatiwala, the tracer-transportmatrix 

approach of Khatiwala et al. (2005) and Khatiwala (2007) at the IFM-GEOMAR 

Biogeochemical Modelling group. Currently she applies this approach to optimise 

parameterisations of organic matter export and remineralisation profiles by evaluating simulated 

nutrient and oxygen fields against observed data, a work that will directly feed into this project. 

Dr. Wolfgang Koeve is a biological oceanographer with particular experience in the analysis and 

evaluation of biogeochemical datasets from local to global scales (Koeve and Ducklow, 2001; 

Koeve et al. 2002; Koeve, 2006). Focus of his recent work was on the coupling of nitrogen and 

carbon cycles (Koeve, 2004; 2005), as well as on synthesis of organic and inorganic particle flux 

estimates (Koeve, 2002). 


Task 5.2.a 

Parameterisations of pelagic calcium carbonate production currently used in biogeochemical 

climate models will be analysed with respect to their sensitivity to temperature and carbonate 

chemistry. These parameterisations are not always well documented in the relevant literature, and 

a careful investigation of technical reports and computer codes will in many cases be required. A 

first attempt to collect this information has shown that this is not a trivial task and that the results 

of such a review of existing parameterisation would be of interest to many modellers that have 

already been using such models for some time. 

Task 5.2.b 

A meta-analysis of hydrographic data and published experimental findings will be used in a first 

step to evaluate existing parameterisations. In collaboration with WP9 of the European EPOCA 

project (led by PI A. Oschlies), which will provide a synthesis of a number of mesocosm 

experiments, we will compile the relevant information with the aim to develop improved 

parameterisations of pelagic CaCO3 production and its sensitivity to environmental conditions. 

This will, in particular, require careful consideration of the different experimental protocols, 

which may help to reconcile apparently contradictory experimental findings (e.g., Riebesell et al., 

2000; Langer et al., 2006; Iglesias-Rodriguez et al., 2008). 

Task 5.2.c 

To obtain an objective assessment of the different parameterisations of calcification and its 

sensitivity to variations in temperature and carbonate chemistry, we will combine the various 

parameterisations with a new computational framework which allows efficient simulations of 

biogeochemical tracers over hundreds to thousands of years, needed to model the impact of 

changed carbonate production, export and dissolution parameterisations on the alkalinity field. 

Export formulations to be investigated range from globally uniform remineralisation length 

scales (Maier-Reimer, 1993) to complex ones invoking processes such as a link between organic 

carbon to CaCO3 flux via the ballast particle flux hypothesis (Armstrong et al., 2001; Heinze, 

2004), aggregation, or calcite dissolution in copepod guts or in marine aggregates (Aumont and 

Bopp, 2006; Gehlen et al., 2007).


The agreement between simulated and observed alkalinity fields will then be quantified by a 

“misfit function” that measures how well the individual parameterisations can reproduce current 

regional variations in CaCO3 production, export and dissolution as a function of regional 

variations in temperature and carbonate chemistry. Impacts of model errors in the circulation and 

organic matter fluxes will be accounted for by a complementary analysis of the model-data 

misfits of hydrographic and biogeochemical tracer distributions. All simulations will be carried 

out under present environmental conditions, but it is expected that the emerging optimised 

parameterisations will improve future climate scenario simulations performed elsewhere. 

Task 5.2.d 

While tasks5.2.a and 5.2.b out of necessity base their synthesis and modelling work on pre- 

BIOACID experimental and modelling efforts, task 5.2.d intends to use novel (novel with respect 

to oceanographic applications) Bayesian meta-analysis methods to analyse the experimental 

findings obtained within themes 1 to 4 of the BIOACID project. Twelve European PhD students 

are currently trained in these techniques as part of the METAOCEANS Marie-Curie action. They 

all work on the application of various meta-analysis techniques to different aspects of marine 

ecosystems and are expected to finish their PhDs in 2010. We hope that BIOACID can take 

advantage of the METAOCAENS project and offer a PostDoc position to one of theses welltrained 

young scientists. 

A start as early as 1.5 years into the project should allow for a timely analysis of the experimental 

findings already during the first BIOACID phase. This will allow the rapid identification of 

remaining uncertainties as well as of the potential for uncertainty reduction. As such, it is 

expected that this initiative will allow for an immediate feedback into experimental strategies, 

which should be of benefit to the entire BIOACID consortium. This will also establish able 

personnel and potent analysis methods in preparation for a second BIOACID phase that only will 

allow the completion of a sound synthesis of the BIOACID experimental findings. 

Interaction with other projects and subprojects 

Concerning tasks 5.2.a to 5.2.c, this project will develop strong links to projects from theme 3 

(Calcification), in particular to projects 3.1 (Cellular mechanisms of calcification; in particular 

3.1.1 Coccolithophores) and 3.2 (Calcification under pH stress; in particular 3.2.1: Sub-polar 

pteropods). Research carried out in project 3.4 (Microenvironmentally controlled 

(de)calcification mechanisms; in particular 3.4.3) and 5.1 (Alkalintiy fluxes in the North Sea) will 

help to clarify the longer term needs for (a) a sediment model compartment and (b) shelf to open 

ocean interactions in the global modelling perspective. Bayesian meta-analysis (Task 5.2.d) shall 

be applied to experimental work planned in Theme 1 (1.2 Turnover of organic matter). 

Descriptions of acidification impacts on the production and export of organic and inorganic 

carbon needed in 5.2 will rely on experimental evidence gathered particularly in subprojects 1.2.1 

(Production and decomposition of exudates), 1.2.2 (Dissolved organic nitrogen release and 

uptake unders stress), and 1.2.5 (Effect of decreasing calcareous/lithogenic ballast on aggregates 

in the benthic boundary layer). The global modelling work of Project 5.2, which uses a stationary 

present-day climate to evaluate model results against observations, has close links to the 

prognostic climate model runs performed in Project 1.3 (Organic carbon pump feedbacks) and 

will further benefit from long-term (geological) perspectives as provided by 3.5.2 and 3.5.3 (Past 

ocean acidification events). 

257

Schedule 

258 



Review of IPCC-type model 

parameterizations of marine calcium 

carbonate cycling 

Review of published experimental 

findings (POC-net production, PIC 

production, DOM, TEP, C:N, N2fix) 

Set-up of prototype 3D model, first 

exploratory model experiments 

Preparation of datasets for model-data 

comparison (from GLODAP, 

CARBOOCEANS, and PANGAEA 

sources). 

Development of data analysis tools 

Model experiments applying a variety 

of formulations for CaCO3 production, 

export and dissolution 

In-depth analysis of model results 

through comparison with observations 

(GLODAP etc.) and subsequent 

parameter optimization 

Compilation of datasets from 

BIOACID experiments from the first 

1.5 years of project into one database 




- Draft documentation (manuscript) on parameterization of marine 

calcium carbonate cycling in IPCC type models 

- Review (manuscript draft) on experimental work 

- Prototype 3D model with carbonate cycle processes implemented, 

major data analysis tools and datasets prepared 

- Implementation of at least three distinct sets of carbonate models 

- Three manuscripts submitted 



5.2.a–c 

5.2.d 

Subtotal 

Consumables 

5.2.a–c 

5.2.d 

Subtotal 

Travel 

5.2.a-c 



month 9 

month 18 

month 21 

month 24 

month 36

5.2.d 

Subtotal 

Investments 

Subtotal 

Other costs 

Subtotal 

Total 


5.2.a-c 



Personnel costs: 1 full time researcher, Dr. W. Koeve, shall be employed to carry out the 

planned review work (see objective 1+2), do the model experiments as well as model-data 

analysis (objective 3) and publishing in peer reviewed journals. Both the review work as well as 

the model-data analysis require a high level of scientific expertise provided by the candidate, who 

has f.e. served as guest editor of a Deep-Sea Research II volume on carbon cycle synthesis, 

convenor at scientific conferences, and reviewer for various funding agencies and scientific 

journals 

Consumables: Euro/yr for publication fees in open access journals (e.g. Biogeosciences) or 

journals with fee-based open access option (e.g. Limnology Oceanography). 

Travel: Euro/yr are requested for annual 2-week visits of Dr. Samar Khatiwala (Columbia 

University, New York) to IFM-GEOMAR. Euro/yr are requested for travel to international 

and national conferences and workshops AGU Autum-2009, AGU Ocean Sciences- 2010, EGU- 

2011, in order to present scientific results of the project. 

Investment: no 

Other costs: no 

5.2.d 

Personnel costs: 1 NN PostDoc for 1.5 years (second half of project). The PostDoc will be 

responsible to carry out the Bayesian meta-analysis of BIOACID experimental findings. 

Currently the EU Marie Curie action METAOCEANS trains PhD students in meta-analysis 

techniques. METAOCEANS students will finish their PhDs in 2010, we are hence expecting a 

number of well trained PostDocs available in summer 2010, being potential candidates for this 

BIOACID task. 

Consumables: Euro for third year. Publication fees. 

Travel: Euro for third year only. Presentation of scientific results at national and international 

meetings; potentially at the AGU autumn meeting in 2001 (San Francisco). 



259


260 


Archer D, Kheshki H, Maier-Reimer E (1998) Dynamics of fossil fuel CO2 neutralization by marine CaCO3. Global Biogeochem Cycles 12(2): 

259-276 

Archer D, Winguth A, Lea D, Mahowald N (2000) What caused the glacial/interglacial atmospheric pCO2 cycles? Rev Geophysics 38(2): 159- 

189 

Armstrong RA, Lee C, Hedges JI, Honjo S, Wakeham SG (2001) A new, mechanistic model for organic carbon fluxes in the ocean based on 

the quantitative association of POC with ballast minerals. Deep-Sea Res II 49: 219-236 

Aumont O, Bopp L (2006) Globalizing results from ocean in situ fertilization studies. Global Biogeochem Cycles 20: GB2017, 

doi:10.1029/2005GB002591 

Gehlen M, Gangto R, Schneider B, Bopp L, Aumont O, Ethe C (2007) The fate of pelagic CaCO3 production in the high CO2 ocean: a model 

study. Biogeosciences 4: 505-519 

Gregg W, Casey N (2007) Modeling coccolithophores in the global oceans. Deep-Sea Res II 54: 447-477 

Heinze C (2004) Simulating oceanic CaCO3 export production in the greenhouse. Geophys Research Lett 31: L16308, 

doi:10.1029/2004GL020613 


P, Rehm E, Armbrust EV, Boessenkool KP (2008) Phytoplankton calcification in a high-CO2 world. Science 320: 336-340 

Khatiwala SA (2007) A computational framework for simulation of biogeochemical tracers in the ocean. Global Biogeochem Cycles 21: 

GB3001, doi:10.1029/2007GB002923 

Khatiwala S, Visbeck M, Cane MA (2005) Accelerated simulation of passive tracers in ocean circulation models. Ocean Modelling 9: 51-69 

Koeve W (2002) Upper ocean carbon fluxes in the Atlantic Ocean - the importance of the POC:PIC ratio. Global Biogeochem Cycles 16: 

GB1056, doi:10.1029/2001GB001836 

Koeve W (2004) Spring bloom carbon to nitrogen ratio of net community production in the temperate N. Atlantic. Deep-Sea Res I 51: 1579- 

1600 

Koeve W (2005) The significance of the TEP-pump to deep ocean carbon fluxes. Mar Ecol Prog Ser 291: 53-64 

Koeve W (2006) Stoichiometry of the biological pump in the North Atlantic - constraints from climatological data. Global Biogeochem Cycles 

20: GB3018, doi:10.1029/2004GB002407 

Koeve W, Ducklow H (2001) JGOFS synthesis and modelling: the North Atlantic ocean. Deep-Sea Res Part II 48: 2141-2154 

Koeve W, Pollehne F, Oschlies A, Zeitzschel B (2002). Storm induced convective export of organic matter during spring in the northeast 

Atlantic. Deep-Sea Res I 49: 1431-1444 

Kriest I (2002) Different parameterizations of marine snow in a 1D-model and their influence on representation of marine snow, nitrogen 

budget and sedimentation Representing phytoplankton aggregates in biogeochemical models. Deep-Sea Res I 49: 2133-2162 

Kriest I, Evans GT (1999) Representing phytoplankton aggregates in biogeochemical models. Deep-Sea Res I 46:1841-1859 

Kriest I, Evans GT (2000) A vertically resolved model for phytoplankton aggregation. P Indian Acad Sci – Earth 109: 553-469 

Kriest I, Oschlies A (2007) Modelling the effect of cell-size-dependent nutrient uptake and exudation on phytoplankton size spectra. Deep-Sea 

Res I 54:1593-1618 

Kriest I, Oschlies A (2008) On the treatment of particulate organic matter sinking in large-scale models of marine biogeochemical cycles. 

Biogeosciences 5: 55-72 

Langer G, Geisen M, Baumann K-H, Kläs J, Riebesell U, Thoms S, Young JR (2006) Species-specific responses of calcifying algae to 

changing seawater carbonate chemistry. Geochemistry Geophysics Geosystems 7: Q09006, doi:10.1029/2005GC001227 

Maier-Reimer E (1993) Geochemical cycles in an ocean general circulation model. Preindustrial tracer distributions. Global Biogeochem 

Cycles 7(3): 645-677 

Oschlies A, Schartau M (2005) Basin-scale performance of a locally optimised marine ecosystem model. Journal of Marine Research 63: 335- 

358 

Ridgwell A, Zondervan I, Hargreaves JC, Bijma J, Lenton TM (2007) Assessing the potential long-term increase of oceanic fossil fuel CO2 

uptake due to CO2-calcification feedback. Biogeosciences 4: 481-492 



Schartau M, Oschlies A (2003) Simultaneous data-based optimization of a 1D-ecosystem model at three locations in the North Atlantic Ocean: 

Part 1. Method and parameter estimates. Journal of Marine Research 61: 765-793 

Schartau M, Oschlies A (2003) Simultaneous data-based optimization of a 1D-ecosystem model at three locations in the North Atlantic Ocean. 

Part 2: Standing stocks and nitrogen fluxes. Journal of Marine Research 61: 795-821 

Schmittner A, Oschlies A, Damon H, Galbraith E (2008) Global Biogeochem Cycles 22: GB1013, doi:10.1029/2007GB002953


Project 5.3: Viability-method for the impact assessment of ocean acidification 

under uncertainty 

PI: M. Quaas 

Development of the method and exemplary application to the impact of acidification on the 

North Sea cod fishery 

i. Objectives 

The objectives of the project 5.3 are 

1. To further develop the ecological-economic viability-method towards a general approach 

for integrated assessment of human actions influencing ocean acidification and the 

consequences for human well-being that takes uncertainties about future development 

into account. 

2. To demonstrate the usefulness of the viability-method by applying it exemplarily to 

assess effects of acidification on the North Sea cod fishery. 


The tolerable-windows approach (Bruckner et al. 1999, Tóth et al. 2002) employed by the 

German Advisory Council on Global Change (WBGU) in the Special Report “The Future Oceans 

– Warming up, Rising High, Turning Sour“ has been conceptualized as a general framework that 

allows an impact assessment for complex systems such as the ocean-climate system. While 

uncertainties about the future development of such systems are of high relevance, the tolerablewindows 

approach does not explicitly address these uncertainties (a recent exemption is Kleinen 

et al. 2007). By contrast, the ecological-economic viability approach (Baumgärtner and Quaas 

2007), based on the concept of stocks (Faber et al. 2005) and on ecological population viability 

analysis (Beissinger and McCullough 2002) allows to assess the sustainability of human actions 

under conditions of uncertainty in a general and unified way. Despite many studies on cod 

fisheries (e.g. Döring and Egelkraut 2008, Röckmann et al. 2008), the effects of acidification 

have not yet been incorporated into a comprehensive ecological-economic modelling analysis. 


The viability-method that shall be developed in the proposed project will build upon the results of 

two ongoing projects funded by the BMBF within the key area Economic sciences for 

sustainability: The proponent Martin Quaas is involved as PI in the project Sustainable use of 

ecosystem services under uncertainty and partner in a sub-project of The Concept of Stocks as 

Decision Support for Sustainability Policy. Within the first project, a general conception of 

ecological-economic viability as a measure of sustainability under uncertainty has been 

developed by the proponent and a coauthor (Baumgärtner and Quaas 2007) and to some extent 

already been applied using methods of ecological-economic modeling (Quaas et al. 2007, Quaas 

and Baumgärtner 2008).The proponent Martin Quaas is leading the Junior Research Group on 

Sustainable Fisheries within Kiel's Cluster of Excellence Future Ocean, where currently 

ecological-economic models of fisheries are developed. 

261

262 



Building on the ecological-economic concept of viability (Baumgärtner and Quaas 2007), a 

“viability-method” shall be developed that allows assessing the consequences human actions both 

leading to ocean acidification and dealing with the consequences of acidification. The 

applicability of the method shall be proven by applying it to the case of the North Sea cod 

fishery. 

For this sake, an ecological-economic model shall be developed that incorporates the effect of 

acidification, building on both available ecological knowledge and the results obtained in 

BIOACID projects 2.3.1 and 2.3.2 (Effects on top predators) on the susceptibility of cod larvae to 

acidification. By means of the viability-method, the impacts of different human actions shall be 

assessed (including measures to mitigate acidification or adapting the management of the North 

Sea cod fishery), taking into account the uncertainties about the future development of 

acidification and the exact impact on cod recruitment. 

Schedule 


Development of the viability-method 

and adaptation to the issue of ocean 

acidification 

Assessment of the effects of 

acidification on the North Sea cod 

fishery using the viability-method 





- Viability-method adapted for the issue of ocean acidification month 30 

- Ecological-economic model of North Sea cod fishery including acidification month 30 

- Two manuscripts submitted month 36 



Subtotal 

Consumables 

Subtotal 

Travel 

Subtotal 

Investments 

Subtotal 

Other costs 

Subtotal 

Total 



Personnel costs: 1 NN (e.g. Sandra Derissen) Postdoc for 1 1/4 years. The PostDoc shall conduct 

the main part of the research in collaboration with Martin Quaas. As the BMBF-funded project 

Sustainable use of ecosystem services under uncertainty will end in the second half of 2010, it is 

very likely that one of the four current PhD students of that project can be hired as Postdoc, who 

already has substantial experience in the methods involved. 

Consumables: Euro in the last year, mainly to cover publication fees. 

Travel: Euro in the last year to present scientific results at national and international conferences, 

e.g. the biannual conferences of the European Society for Ecological Economics or the World 

Congress of Environmental and Resource Economists. 




Baumgärtner S. Quaas MF (revise and resubmit) Ecological-economic viability as a criterion of strong sustainability under uncertainty. 

Ecological Economics 

Beissinger S, McCullough D (eds.) (2002. Population Viability Analysis. Chigaco: University of Chicago Press 

Bruckner T, Petschel-Held G, Tóth FL, Füssel HM, Helm C, Leimbach M Schellnhuber HJ (1999) Climate-change decision support and the 

tolerable windows approach, Environmental Modeling and Assessment 4: 217–234 

Döring R, Egelkraut TM (2008) Investing in natural capital as management strategy in fisheries: The case of the Baltic Sea cod fishery. 

Ecological Economics 64: 634–642 

Faber M, Frank K, Klauer K, Manstetten R, Schiller J, Wissel C (2005) On the foundation of a general theory of stocks. Ecological Economics 

55(2):155-175 

Kleinen, T Petschel-Held G, Bruckner T(2007) The probabilistic tolerable windows approach, submitted to Climatic Change 

Quaas M F. Baumgärtner S (2008) Natural vs. financial insurance in the management of public-good ecosystems. Ecological Economics 

65:397-406 

Quaas MF and Requate T (in preparation). Sushi or fish fingers? How preferences for sustainability affect the sustainability of fisheries 

Quaas M ., Baumgärtner S, Becker C, Frank K, Müller B (2007). Uncertainty and sustainability in the management of rangelands. Ecological 

Economics 62:251-266 

Röckmann C, Schneider UA, St.John MA, Tol RSJ (2008). Rebuilding the Eastern Baltic cod stock under environmental change - a 

preliminary approach using stock, environmental, and management constraints, forthcoming in Natural Resource Modeling 

Skonhoft A, Quaas MF, Requate T, Ruckes K (in preparation). A bioeconomic modelling analysis of how to optimally fish an age-structured 

population 

Tóth FL, Bruckner T, Füssel HM, Leimbach M, Petschel-Held G, Schellnhuber HJ (2002). Exploring options for global climate policy: a new 

analytical framework. Environment 44 (5):22–34 

263

12. Appendices 

264 


- Project structure 

- Contact data of principal investigators

Theme 

Project and data 

management, 

training and 

infrastructure 

development 

Primary 

production, 

microbial 

processes and 

Theme 

Leader 

Ulf 

Riebesell 

Maren 

Voß 


Code Project (Cluster) Project PI Code Subproject 

0 

1 

Project coordination Ulf Riebesell 0.1 

Data management Ulf Riebesell 0.2 

Infrastructure 

development 

Training and transfer 

of knowhow 

Hans Pörtner 0.3 pH stat mesocosms 

Michael 

Meyerhöfer 

265 

Development of chemical 

optical sensor technology 

for the determination of 

pCO2 in fluids of marine 

organisms and the marine 

environment 

Subproject - 

PI 

Franz-J. 

Sartoris 

Athanas 

Apostolidis / 

Christian 

Huber 

Code Links to Subprojects 

0.3.1 

0.3.2 

0.4 All 

3.4.2 / 1.1.5 / 2.1.2 / 

2.1.3 / 2.3.1 / 2.3.2 / 

3.1.3 / 3.1.4 / 3.2.2

266 

Theme 

biogeochemical 

feedbacks 

Theme 

Leader 



Acclimation versus 

adaptation in 

autotrophs 

Thorsten 

Reusch 

1.1 

Impact of changing climate 

on a 

chemolithoautotrophic 

epsilonproteobacterium 

from a pelagic redoxcline 

The interplay between 

carbon-and iron-availability 

and its impact on 

photosynthesis of primary 

producers in the ocean 

Long-term response of 

phytoplankton on climate 

change: a 

multidimensional approach 

Rapid evolution of key 

phytoplankton species to a 

high pCO2 ocean 

Interactive effects of CO2 

concentration and 

temperature on 

microphytobenthic 

Subproject - 

PI 

Günter Jost / 

Klaus 

Jürgens 

Michael 

Hippler / Julie 

LaRoche 


1.1.1 

1.1.2 

Marius Müller 1.1.3 

Thorsten 

Reusch / Ulf 

Riebesell 

Ulf Karsten / 

Thomas 

Hübener 

1.1.4 

1.1.5 

1.1.2 / 1.1.3 / 1.1.4 / 

1.1.5 / 1.2.4 / 4.1.4 

1.1.3 / 1.1.1 / 1.1.4 / 

1.2.2 / 1.2.4 / 1.3 / 

3.3.1 

1.1.2 /1.1.4 / 1.3 / 

3.1.1 /3.1.4 / 3.5.2 / 

3.5.3/ 

1.1.2 / 1.1.3 / 3.1.1 / 

3.5.2/ 4.1.2 / 4.2.1 

0.3.2/ 1.1.1 / 3.4.1 / 

3.4.2 / 4.1.1

Theme 

Theme 

Leader 



Turnover of organic 

matter 

Modelling 

biogeochemical 

feedbacks of the 

organic carbon pump 

Anja Engel 1.2 

Birgit 

Schneider 

267 

biodiversity and 

ecosystem function 

Production and 

decomposition of exudates 

Dissolved organic nitrogen 

release and uptake under 

stress 

DOM availability and 

phosphorus utilization 

Microbial response to 

DOM release and 

aggregation 

Effect of decreasing 

calcareous / lithogenic 

ballast on aggregates in 

the benthic boundary layer 

1.3 no subproject 

Subproject - 

PI 


Anja Engel 1.2.1 4.1.1/ 4.2.1 / 4.2.2 / 5.1 

/ 5.2 

Maren Voß 1.2.2 4.2.1 / 4.2.2 / 5.2 

Monika 

Nausch / 

Günther 

Nausch 

Hans – Peter 

Grossart 

Laurenz 

Thomsen / 

Giselher Gust 

Birgit 

Schneider 

1.2.3 

1.2.4 

1.2.5 

1.3 

4.1.1 / 4.1.4 / 4.2.1. / 

4.2.2. 

1.1.1 / 1.1.2 / 1.3 / 

3.4.2 / 4.1.4 / 4.2.1 / 

5.1 / 5.3 

1.2 / 1.3 / 3.1.1 / 3.4.1. 

/ 3.4.3 / 5.2 

1.1.2/ 1.1.3/ 1.2.1/ 

1.2.2/ 1.2.3/ 1.2.4/ 

1.2.5/ 3.1.1/ 3.2.1/ 

3.4.3/ 3.5.2 / 3.5.3/ 5.1/ 

5.2/ 5.3

268 

Theme 

Performance 

characters: 

reproduction, 

growth and 

behaviours in 

animal species 

Theme 

Leader 

Hans 

Pörtner 



2 

Effects on grazers and 

filtrators 

Long-term 

physiological effects 

on different life stages 

of benthic 

Thomas Brey 2.1 

Felix Mark 2.2 

Ocean Acidification and 

Reproduction: Is the 

beginning of Life in 

Danger? 

The response of 

zooplankton organisms to 

elevated CO2 

concentrations 

Calcifying 

macroorganisms in 

acidifying & warming 

shallow waters 

Hyas araneus: Sensitivity, 

adaptive capacities and 

evolutionary 

consequences in 

Subproject - 

PI 

Angela 

Köhler 

Barbara 

Niehoff 


2.1.1 

2.1.2 

Gisela Lannig 2.1.3 

Daniela 

Storch 

2.2.1 

2.2.1/2.3.1 /3.1.3/ 3.1.4 

/ 3.2.2 / 4.1.2 

0.3.1/ 0.3.2 /2.1.3 / 

2.2.1 / 3.1.3 / 3.1.4/ 

4.2.1 

0.3.1 / 0.3.2 / 2.1.2 / 

2.2.1/ 2.2.2 / 2.3.1 / 

2.3.2 / 3.1.3 / 3.1.4 

/3.2.4 / 3.3.1 / 3.3.2/ 

3.4.1 / 3.5.1 /4.1.2 

0.3.1 / 2.1.2 / 2.1.3 / 

2.2.2 / 2.3.2 / 3.1.3 / 

4.1.2

Theme 

Theme 

Leader 



crustaceans populations from different 

Effects on top 

predators (fishes, 

cephalopods) 

Catriona 

Clemmesen 

269 

2.3 

latitudes 

Cancer pagurus: Chronic 

and acute responses – 

Adaptation versus 

Tolerance 

Effects of changes in 

ocean pH on the 

development, growth, 

metabolism and 

otolith/statolith formation 

and composition of fish 

and cephalopod early life 

stages: a comparative 

approach 

Mechanisms setting and 

compensating for animal 

sensitivity to ocean 

acidification: functional 

capacities, thermal 

interactions and 

mechanism-based 

modelling 

Subproject - 

PI 

Christopher 

Bridges 

Uwe 

Piatkowski 

Magnus 

Lucassen 


2.2.2 2.1.3 / 2.2.1 / 3.1.3 

2.3.1 

2.3.2 

0.3.1 / 0.3.2 / 2.1.1 / 

2.1.3 / 2.3.2 / 3.1.3 / 

3.1.4/ 3.3.1 / 4.1.2/ / 

5.3 

0.3.1 / 0.3.2. / 2.1.3. / 

2.2.1. / 2.2.2. / 2.3.1. / 

3.1.3. / 3.1.4

270 

Theme 

Calcification: 

Sensitivities 

across phyla 

and ecosystems 

Theme 

Leader 

Dirk de 

Beer 



3 

Cellular mechanisms 

of calcification 

Frank 

Melzner 

3.1 

Inorganic carbon 

acquisition for calcification 

and photosynthesis in 

marine coccolithophores: 

towards a unifying theory 

Transepithelial calcification 

processes in the 

hermatypic cold-water 

coral Lophelia pertusa 

(Scleractinia) 

Calcification & ion 

homeostasis in the phylum 

mollusca in response to 

ocean acidification 

Sea urchin membrane 

transport mechanisms for 

calcification and pH 

regulation 

Subproject - 

PI 

Kai Schulz 3.1.1 

Armin Form 3.1.2 

Frank 

Melzner 

Markus 

Bleich 


3.1.3 

3.1.4 

1.1.3 / 1.1.4/ 1.2.5 / 1.3 

/ 3.4.2 / 3.5.2 / 3.5.3 / 

4.2.2 / 5.2 

1.2.5 / 3.2.2 / 3.2.4 / 

3.3.1 / 3.3.2 / 3.4.2 / 

4.1.4 

2.1.1 / 2.1.3 / 2.2.1 / 

2.2.2 / 2.3.1 / 2.3.2 / 

3.2.1 / 3.4.2 / 4.1.2 

1.1.3. / 2.1 / 2.3 / 3.2 / 

3.3 /3.4.2 / 4.1.2.

Theme 

Theme 

Leader 



Calcification under 

pH-stress: Impacts on 

ecosystem and 

organismal levels 

Ultra-structural 

changes and trace 

element / isotope 

partitioning in 

calcifying organisms 

(foraminifera, corals) 

Ralph 

Tollrian 

Jelle Bijma 3.3 

271 

3.2 Impact of ocean 

acidification and warming 

on sub-polar shelled 

pteropods 

Impact of ocean 

acidification on 

reproduction, recruitment 

and growth of scleractinian 

corals 

Coral calcification in 

marginal reefs 


acidification on coralline 

red algae 


acidification on the 

calcification mechanisms 

in marine calcifying 

organisms and on ultra 

structural changes of 

biogenic calcite 

Subproject - 

PI 


Ulf Riebesell 3.2.1 1.2.5 / 1.3 / 3.1.3 / 

Ralph Tollrian 3.2.2 

Claudio 

Richter 

Jan Fietzke 3.2.4 

Jelle Bijma 3.3.1 

3.2.4 / 3.3.1-3.4.3/ 

3.5.1 / 5.2 

2.1.1 / 3.1.2 / 3.2.3 / 

3.2.4 / 3.3.2 / 3.4.2 / 

3.5.2 / 4.1.2 / 4.1.3 

3.2.3 3.1.2/3.2.2/3.2.4/3.3.2/ 

3.4.2/ 4.1.3 

2.1.3 / 3.1.2 / 3.2.1 

/3.2.2 / 3.2.3 / 3.4.2 / 

4.1.3 / /4.1.1 

2.1.3 / 2.3.1 / 3.1.2 / 

3.1.3 / 3.1.4 / 3.2.1 / 

3.2.4

272 

Theme 

Theme 

Leader 



Microenvironmentally 

controlled (de- 

)calcification 

mechanisms 

Impact of present and 

past ocean 

acidification on 

metabolism, 

biomineralization and 

biodiversity of pelagic 

and neritic calcifiers 

Michael 

Böttcher 

Adrian 

Immenhauser 

3.4 

3.5 

The effect of decreasing 

pH, salinity and 

temperature on the trace 

element and isotope 

partitioning between 

marine calcifying 

organisms and seawater 

Impact of biogenic 

carbonates on pH 

buffering in an acidifying 

coastal sea (North Sea) 

Benthic (de-)calcification 

driven by microbial 

processes 

Buffering ocean 

acidification: Dissolution of 

carbonate sediments in 

the Southern Ocean 

Comparison of Cultured 

and Fossil Bivalve 

Geochemistry and Shell 

Ultrastructure 

Subproject - 

Anton 

PI 

Eisenhauer 

Michael 

Böttcher 


3.3.2 0.4 / 2.1.3 / 2.3.1 / 

3.4.1 

Dirk de Beer 3.4.2 

Mario 

Hoppema 

Adrian 

Immenhauser 

3.4.3 

3.1.2 / 3.1.3 / 3.1.4 

3.2.1 / 3.2.4 

1.2.5 / 2.1.3 / 3.4.2 / 

3.4.3 / 5.1 

0.3.2 / 1.1.5 / 3.1 / 

3.2.3 / 3.2.4 / 3.3.2 

/4.1.3 

1.2.5 / 1.3 / 3.4.1 / 

3.4.2 / 5.1 / 5.2 

3.5.1 2.1.3 / 3.5.3 / 4.1.2

Species 

Theme 

interactions and 

community 

structure: will 

ocean 

acidification 

cause regime 

shifts? 

Theme 

Leader 

Maarten 

Boersma 



4 

OA impacts on 

interactions in and 

structure of benthic 

Martin Wahl 4.1 

273 

Biological response to 

short-termed ocean 

acidification events in the 

past: biodiversity and 

evolution patterns of 

marine primary producers 

(calcareous nannofossils) 

during the late Paleocene 

– early Eocene 

Nannoplankton response 

to modern and past ocean 

acidification events 

Effects of ocean 

acidification on trophic 

interactions in coastal 

Subproject - 

Jörg 

PI 

Mutterlose 

Sebastian 

Meier 

Ragnhild 

Asmus 


3.5.2 

3.5.3 

4.1.1 

1.1.3 / 1.1.4/ 1.3 / 3.1.1 

/ 3.2.2 / 5.2 

1.1.3 / 1.1.4 / 1.3 / 

3.1.1 / 4.2.2 / 5.2 

1.1.5 / 1.2.1 / 3.2.4 / 

4.1.2 / 4.1.3 / 4.1.4 / 

4.2.1 / 4.2.2 / 5.1

274 

Theme 

Theme 

Leader 



communitiesbenthos seaweed and seagrass 

OA effects on food 

webs and competitive 

interactions in pelagic 

ecosystems 

Maarten 

Boersma 

4.2 

ecosystems 

Acidification stress: Early 

life stage ecology in times 

of global change 

Competitive success of 

calcifying and non- 

calcifying macroalgae 

under shifting pH regimes 

in tropical vs. temperate 

regions 

Effects of ocean 

acidification on microbial 

community structure, 

composition and activity in 

natural and experimental 

systems 

OA effects on pelagic 

community structure and 

food chains 

Subproject - 

PI 

Martin Wahl 4.1.2 

Kai Bischof 4.1.3 

Alban 

Ramette 

Maarten 

Boersma 


4.1.4 

4.2.1 

1.1.4 / 2.1.1 / 2.1.3 / 

2.2.1 / 2.3.1 / 3.1.3 / 

3.1.4 / 3.2.2 / 3.2.3 / 

3.5.1 / 4.1.1 / 4.1.3 

4.1.4 / 4.2.1 / 4.2.2 

3.2.2. / 3.2.3. / 3.2.4. / 

3.4.2. / 4.1.1 / 4.1.2 / 

4.1.4 / 4.2.2 

1.1.1 / 3.1.2 / 3.4.2 / 

4.1.1 / 4.1.2 / 4.1.3 / 

4.2.1 / 4.2.2 

1.2.1 / 1.2.2 / 1.2.3 / 

1.2.4. 2.1.2. / 4.1.1 / 

4.1.2 / 4.1.4 / 4.2.2

Theme 

Integrated 

assessment: 

Sensitivities and 

uncertainties 

Theme 

Leader 

Andreas 

Oschlies 



5 

Impact of Alkalinity 

fluxes from the 

Wadden Sea on the 

carbon cycle and the 

primary production in 

the North Sea 

Evaluating and 

optimising 

parameterisations of 

pelagic calcium 

carbonate production 

in global 

biogeochemical ocean 

models 

Johannes 

Pätsch 

Andreas 

Oschlies 

275 

Competitive interactions in 

planktonic microalgae 

under OA-stress 



Subproject - 

PI 

Björn Rost 4.2.2 


1.2.1 / 1.2.2 / 1.2.3 / 

3.1.1 / 3.5.3 / 4.1.1 / 

4.1.2 / 4.1.3 / 4.1.4 / 

4.2.1 

1.2.1 / 1.2.2 / 1.2.3 / 

1.3 / 3.4.1 / 3.4.3 / 

4.1.1/ 5.2 

1.2.1 / 1.2.2 / 1.2.5 / 

1.3 / 3.1.1 / 3.2.1 / 3.4 

(3.4.3) / 3.5.2 / 3.5.3 / 

5.1

276 

Theme 

Theme 

Leader 



Viability-method for 

the impact 

assessment of ocean 

acidification under 

uncertainty 

Subproject - 

PI 


Martin Quaas 5.3. no subproject 2.3.1 / 2.3.2

Appendix: Principal Investigators 


Project Name Institute Address Email Phone 

0.1 Riebesell Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel uriebesell@ifm-geomar.de 0431 600-4444 

0.2 Diepenbroek MARUM - Institute for Marine Environmental Sciences , University of Bremen Leobener Strasse, POP 330 440, 28359 Bremen mdiepenbroek@pangaea.de 0421 218-65590 

0.3 Pörtner Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Hans.Poertner@awi.de 0471 4831-1307 

Buck Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Bela.H.Buck@awi.de 0471 4831-1868 

0.3.1 

Fisch 

Krieten 

Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Ralf.Fisch@awi.de 

Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Guido.Krieten@awi.de 

0471 4831-1639 

0471 4831-1418 

Sartoris Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Franz-Josef.Sartoris@awi.de 0471 4831-1312 

0.3.2 

Apostolidis Presens Precision Sensing GmbH Josef-Engert-Str. 11, 93053 Regensburg athanas.apostolidis@presens.de 0941 942 72 150 / 114 

Huber Presens Precision Sensing GmbH Josef-Engert-Str. 11, 93053 Regensburg christian.huber@presens.de 0941 942 72 150 / 114 

0.4 Meyerhöfer Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel mmeyerhoefer@ifm-geomar.de 0431 600-4214 

1.1 Reusch University of Münster Hüfferstr. 1, 48149 Münster treusch@uni-muenster.de 0251 - 83 - 2 1095 

1.1.1 

Jost Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock guenter.jost@io-warnemuende 0381 5197-270 

Jürgens Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock klaus.juergens@io-warnemuende.de 0381 5197-250 

1.1.2 

Hippler University of Münster Hindenburgplatz 55, 48143 Münster mhippler@uni-muenster.de 0251 - 83 2 4790 

LaRoche Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel jlaroche@ifm-geomar.de 0431 600-4212 

1.1.3 Müller Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel mnmueller@ifm-geomar.de 0431 600-4526 

1.1.4 

Reusch University of Münster 

Riebesell Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel 

Hüfferstr. 1, 48149 Münster 

Düsternbrooker Weg 20, 24105 Kiel 

treusch@uni-muenster.de 

uriebesell@ifm-geomar.de 

0251 - 83 - 2 1095 

0431 600-4444 

1.1.5 

Karsten University of Rostock Albert-Einstein-Str. 3, 18059 Rostock 

Hübener University of Rostock Wismarsche Str. 8, 18051 Rostock 

ulf.karsten@uni-rostock.de 

thomas.huebener@uni-rostock.de 

0381 498 6090 

0381 498 6210 

1.2 Engel Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven anja.engel@awi.de 0471 4831-1055 

1.2.1 Engel Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven anja.engel@awi.de 0471 4831-1055 

1.2.2 Voß Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock maren.voss@io-warnemuende.de 0381 5197 209 

1.2.3 

Nausch, M. Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock monika.nausch@io-warnemuende.de 

Nausch, G. Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock guenther.nausch@io-warnemuende.de 

0381 5197 227 

0381 5197 332 

1.2.4 Grossart Leibniz-Institute of Freshwater Ecology and Inland Fisheries (IGB), Berlin Alte Fischerhütte 2, OT Neuglobsow,16775 Stechlin hgrossart@igb-berlin.de 033082 699 91 

1.2.5 

Thomsen Jacobs University Bremen gGmbH Campus Ring 1, 28759 Bremen l.thomsen@jacobs-university.de 0421 200-3254 

Gust Hamburg University of Technology (TUHH) Schwarzenbergstraße 95 C, 21073 Hamburg gust@tu-harburg.de 040 42878 6000 

1.3 Schneider University of Kiel Ludewig-Meyn-Straße 10, 24118 Kiel bschneider@gpi.uni-kiel.de 0431 880-3254 

2.1 Brey Alfred Wegener Institute for Polar and Marine Research Am Alten Hafen 26, 27568 Bremerhaven Thomas.Brey@awi.de 0471 4831-1300 

2.1.1 

Köhler Alfred Wegener Institute for Polar and Marine Research 

Bickmeyer Alfred Wegener Institute for Polar and Marine Research 

Am Handelshafen 12, 27570 Bremerhaven 

Kurpromenade 201, 27498 Helgoland 

Angela.Koehler@awi.de 

Ulf.Bickmeyer@awi.de 

0471 4831-1407 

04725 819-3224 

2.1.2 Niehoff Alfred Wegener Institute for Polar and Marine Research Am Alten Hafen 26, 27568 Bremerhaven Barbara.Niehoff@awi.de 0471 4831-1371 

2.1.3 Lannig Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Gisela.Lannig@awi.de 0471 4831-2015 

2.2 Mark Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven fmark@awi.de 0471 4831-1015 

2.2.1 Storch Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Daniela.Storch@awi.de 0471 4831-1934 

2.2.2 Bridges University of Düsseldorf Universitätsstraße 1, 40225 Düsseldorf bridges@uni-duesseldorf.de 0211 81-14991 

277

Appendix: Principle Investigators (continued) 

278 


Project Name Institute Address Email Phone 

2.3 Clemmesen Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel cclemmesen@ifm-geomar.de 0431 600 4558 

2.3.1 Piatkowski Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel upiatkowski@ifm-geomar.de 0431 600-4571 

2.3.2 Lucassen Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Magnus.Lucassen@awi.de 0471 4831-1340 

3.1 Melzner Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel fmelzner@ifm-geomar.de 0431 600-4274 

3.1.1 Schulz Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel kschulz@ifm-geomar.de 0431 600-4510 

3.1.2 Form Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel aform@ifm-geomar.de 0431 600-1987 

3.1.3 Melzner Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel fmelzner@ifm-geomar.de 0431 600-4274 

3.1.4 Bleich University of Kiel Hermann-Rodewald-Straße 5, 24118 Kiel m.bleich@physiologie.uni-kiel.de 0431 880-2961 

3.2 Tollrian Ruhr-University Bochum Universitätsstraße 150; 44801 Bochum tollrian@rub.de 0234 32-14114 

3.2.1 Riebesell Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel uriebesell@ifm-geomar.de 0431 600-4444 

3.2.2 Tollrian Ruhr-University Bochum Universitätsstr.150, 44780 Bochum tollrian@rub.de 0234 32-24998 

3.2.3 Richter Alfred Wegener Institute for Polar and Marine Research Columbusstrasse, 27568 Bremerhaven crichter@awi.de 0471 4831-1304 

3.2.4 Fietzke Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Wischhofstraße 1-3, 24148 Kiel jfietzke@ifm-geomar.de 0431 600-2106 

3.3 Bijma Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Jelle.Bijma@awi.de 0471 4831-1831 

3.3.1 Bijma Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Jelle.Bijma@awi.de 0471 4831-1831 

3.3.2 Eisenhauer Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Wischhofstraße 1-3, 24148 Kiel aeisenhauer@ifm-geomar.de 0431 600-2282 

3.4 Böttcher Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock michael.boettcher@io-warnemuende.de 0381 5197-402 

3.4.1 Böttcher Leibniz Institute for Baltic Sea Research (IOW), Warnemünde Seestr. 15, 18119 Rostock michael.boettcher@io-warnemuende.de 0381 5197-402 

3.4.2 de Beer Max Planck Institute for Marine Microbiology Celsiusstr. 1, 28359 Bremen dbeer@mpi-bremen.de 0421 2028 - 802 

3.4.3 Hoppema Alfred Wegener Institute for Polar and Marine Research Bussestrasse 24, 27570 Bremerhaven Mario.Hoppema@awi.de 0471 4831-1884 

3.5 Immenhauser Ruhr-University Bochum Universitätsstraße 150, 44801 Bochum Adrian.Immenhauser@rub.de 0234 32-28250 

3.5.1 Immenhauser Ruhr-University Bochum Universitätsstraße 150, 44801 Bochum Adrian.Immenhauser@rub.de 0234 32-28250 

3.5.2 Mutterlose Ruhr-University Bochum Universitätsstraße 150, 44801 Bochum Joerg.Mutterlose@rub.de 0234 32-23249 

3.5.3 

Meier 

Kinkel 

University of Kiel 

University of Kiel 

Ludewig-Meyn-Straße 14, 24118 Kiel 

Ludewig-Meyn-Straße 12, 24118 Kiel 

sm@gpi.uni-kiel.de 

hki@gpi.uni-kiel.de 

0431 880-2936 

431 880-2878 

4.1 Wahl Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel mwahl@ifm-geomar.de 0431 600-4577 

4.1.1 R. Asmus Alfred Wegener Institute for Polar and Marine Research Hafenstraße 43, 25992 List Ragnhild.Asmus@awi.de 04651 956-4308 

4.1.2 Wahl Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel mwahl@ifm-geomar.de 0431 600-4577 

4.1.3 Bischof University of Bremen Leobener Str., NW 2, 28359 Bremen kbischof@uni-bremen.de 0421 218-2859 

4.1.4 

Ramette 

Boetius 

Max Planck Institute for Marine Microbiology 

Max Planck Institute for Marine Microbiology 

Celsiusstr. 1, 28359 Bremen aramette@mpi-bremen.de 0421 2028 - 863 

Celsiusstr. 1, 28359 Bremen aboetius@mpi-bremen.de 0 421 2028 - 860 

4.2 Boersma Alfred Wegener Institute for Polar and Marine Research Kurpromenade, 27498 Helgoland Maarten.Boersma@awi.de 04725 819-3350 

4.2.1 Boersma Alfred Wegener Institute for Polar and Marine Research Kurpromenade, 27498 Helgoland Maarten.Boersma@awi.de 04725 819-3350 

4.2.2 Rost Alfred Wegener Institute for Polar and Marine Research Am Handelshafen 12, 27570 Bremerhaven Bjoern.Rost@awi.de 0471 4831-1809 

5.1 Pätsch Institute of Oceanography, University of Hamburg Bundesstr. 53, 20146 Hamburg paetsch@ifm.uni-hamburg.de 040 - 42838 6628 

5.2 Oschlies Leibniz Institute of Marine Sciences, IFM-GEOMAR, Kiel Düsternbrooker Weg 20, 24105 Kiel aoschlies@ifm-geomar.de 0431 600-1936 

5.3 Quaas University of Kiel Wilhelm-Seelig-Platz 1, 24118 Kiel quaas@economics.uni-kiel.de 0431 880 -3616

BIOACID Programme - Natural Environment Research Council

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