VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

EMP066Nutritional physiology of Sarcinomyces petricola A95, amodel black fungus to study primary successions interrestrial ecosystemsC. Nai* 1,2 , A. Pannenbecker 1,2 , S. Noack 1 , W.J. Broughton 1 ,A.A. Gorbushina 1,21Department of Materials and Environment, Federal Institute for MaterialsResearch and Testing, Berlin, Germany2 Institute of Chemistry, Department of Biology, Chemistry and Pharmacy,Free University Berlin, GermanyPrimary successions in terrestrial ecosystems usually involve microbialgrowth on bare rock surfaces. In these extremely stressed environmentswhich include hot and cold deserts, complex microbial communities mustadapt to high solar irradiation, temperature extremes as well as low waterand nutrient availability. Subaerial biofilms (SABs) has been used todescribe these microbial communities and are dominated by associations offungi, algae, cyanobacteria and heterotrophic bacteria [1]. Althoughmicrobial members of SABs communities vary, the presence of melanizedascomycetous fungi is common on rock and material surfaces in all climaticzones. These fungi grow in restricted compact colonies and are thereforeoften referred to as microcolonial fungi (MCF, [2]). MCF persist on theinterface between a solid substrate and the atmosphere (e.g. on materialsurfaces, roof tiles, rocks) and actively alter the substrate by physical and/orchemical mechanisms. Despite being ubiquitous and important to rockweathering, soil formation and material deterioration and preservation,relatively little is known about their physiology.In our laboratory, we work on the black fungus Sarcinomyces petricola A95as a model organism to study life development and persistence on subaerialrock and material surfaces. We report here preliminary results on thephysiological characterization of S. petricola A95 for what concerns nutrientrequirements (e.g. C- and N-sources) and growth phenotypes in differentlaboratory media. Moreover, results on the symbiotic growth of S. petricolaA95 with the photosynthetic, diazotrophic cyanobacterium Nostocpunctiforme ATCC29133 are presented.[1] Gorbushina, A.A. and W.J. Broughton (2009): Microbiology of the Atmosphere-Rock Interface:How Biological Interactions and Physical Stresses Modulate a Sophisticated Microbial Ecosystem,Ann Rev Microbiol 63:431-50.[2] Staley et al (1982): Microcolonial Fungi: Common Inhabitants on Desert Rocks?, Science215:1093-5.EMP067Molecular biological methods for qualitative analysis ofcultured bacteria of workplacesS. Weiß*, U. Jäckel, J. SchäferFederal Institute for Occupational Safety and Health (BAuA), BiologicalAgents, Berlin, GermanyCurrent quantitative detection methods of workplace related bacterialexposure levels are based on cultivation dependent approaches. However,routinely a qualitative statement can not be made because of the timeconsuming but necessary isolation procedure. Here, genotypic identificationof bacteria could provide a possibility for the routine qualitative analysis.For this purpose an effective DNA extraction protocol is needed which iscapable for isolation of DNA from nearly all bacterial species. To establishsuch a DNA extraction method the efficiency of three different DNAisolation kits from different manufactures (Sigma Aldrich, Analytik Jenaand MP) was investigated, using fifteen different bacterial type species fromthe phyla Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria. TheGenElute TM Plant DNA extraction Kit, offered by Sigma Aldrich wasdeployed in two different procedures, once according to manufacturesinstructions and secondly combined with an additional mechanical celldisruption. To determine the DNA extraction efficiency of all investigatedDNA isolation kits a defined cell count (10 8 cells) of each bacterial specieswere deployed per assay. These cell counts were chosen to avoid anexceeding of columns binding capacity. To identify the DNA extractionmethod, which is most efficient according the amount of extracted DNA ofall employed bacterial strains a ranking procedure was applied. Preliminaryresults showed that the FastDNA ® SPIN Kit for Soil, offered by MP, was themost efficient DNA extraction method based on the chosen ranking criteria.Based on PCR amplification efficiencies using universal bacterial 16SrRNA-gene primers (27f/ 1492r) a second ranking will be done in futureinvestigations. Here the quality of isolated bacterial DNA for amplificationshould be ascertained.The results will be discussed at the poster.EMP068Characterization of Steroid Degrading Bacteria from theBaltic Sea at Kiel GermanyG. Xiong*, T. Zhang, Y. Sang, E. MaserInstitute of Toxicology and Pharmacology, Christian-Albrechts-University,Kiel, GermanySteroid contamination of sea water is an ever growing problem and impactspopulation dynamics of all kinds of sea animals. We have long experiencewith the soil bacterium Comamonas testosteroni which is able to catabolizea variety of steroids and polycyclic aromatic hydrocarbons, and which mightbe used in the bioremediation of contaminated soil. For our studies we use3α-hydroxysteroid dehydrogenase/carbonyl reductase (3α-HSD/CR) as areporter enzyme, since it is the key enzyme in steroid degradation.Moreover, the expression of the corresponding gene, hsdA, is induced byenvironmental steroids. In previous investigations we have identified anddescribed several genes being involved in hsdA regulation. In this work weisolated several bacterial strains from the Baltic Sea at Kiel, Germany,which degrade steroids and which are able to use steroids as carbon source.Two of them, strain S19-1and H5, were characterized as being gramnegative. 16S-rRNA analysis showed that S19-1 belongs to Buttiauxellanoackiae and H5 is similar to Vibrio porteresiae. They could be best grownin SIN medium supplemented with 0.6 - 5.1 % NaCl and at 20°C. Both S19-1 and H5 can use testosterone, estradiol or cholesterol as a carbon source inminimum medium. In Comamonas testosteroni about 20 enzymes could beinduced by 0.5 mM testosterone. A new plasmid pKEGFP-2, suitable formetagenomic studies, and pGEM-EGFP were prepared for isolation ofsteroid inducible genes in S19-1 and H5. A 4.610 kb DNA fragment whichcontains the 3a-HSD/CR gene and its regulation elements from Comamonastestosteroni was cloned into plasmid pGEM-EGFP and pKEGFP-2. Theresult showed that testosterone induction could be detected by a microplatefluorescence reader after the plasmids were transformed into E coli HB101cells. Therefore, the system could be used to isolate steroid degradation andsteroid regulatory genes from S19-1 and H5. In addition, the exactcharacterization and systematic classification of these marine steroiddegrading bacterial strains is envisaged. The strains might be used for thebioremediation of steroid contaminations in sea water.EMP069Earthworms sustain Alpha- and Betaproteobacterial 4-Chloro-2-Methylphenoxyacetic Acid HerbicideDegraders in SoilY.-J. Liu 1,2 , S.-J. Liu 2 , H.L. Drake 1 , M.A. Horn* 11 Department of Ecological Microbiology, University of Bayreuth, Bayreuth,Germany2 Institute of Microbiology, Chinese Academy of Sciences, Beijing, China2-Methyl-4-chlorophenoxyacetic acid (MCPA) is a widely used herbicideand subject to aerobic microbial degradation. Earthworms represent thedominant macrofauna in many soils and enhance both growth and activity ofMCPA-degrading bacteria in soil. Thus, active MCPA degraders in soil anddrilosphere (i.e., burrow walls, gut content, and cast) of the earthwormAporrectodea caliginosa were assessed by 16S rRNA stable isotope probingin soil columns under experimental conditions designed to minimizelaboratory incubation biases. The presence of earthworms decreased the timetaken to degrade agriculturally relevant concentrations of [ 13 C]MCPA (20μg g dw -1 ). 16S rRNA analysis revealed 73 OTUs indicative of activeAcidobacteria, Actinobacteria, Bacteroidetes, Chloroflexi, Cyanobacteria,Firmicutes, Gemmatimonadetes, Planctomycetes, Proteobacteria, andVerrucomicrobia in soil and drilosphere. Seven OTUs indicative of Alpha-,Beta-, Gammaproteobacteria, and Firmicutes consumed MCPA.Alphaproteobacteria (Sphingomonadaceae and Bradyrhizobiaceae) weredominant consumers of MCPA in soil and drilosphere. In contrast, Beta-(Comamonadaceae) and Gammaproteobacteria (Xanthomonadaceae) wereMCPA consumers in burrow walls, indicating that this part of thedrilosphere is favorable for beta- and gammaproteobacterial MCPAdegraders. Approximately 20 and 350 μg [ 13 C]MCPA g dw -1 were consumedwithin 24 hours and 20 days, respectively, in oxic microcosms withdrillosphere material (i.e., bulk soil, burrow walls, and cast). Gut contentsdid not facilitate the degradation of MCPA. Sphingomonadaceae dominatedMCPA consumers in bulk soil and burrow wall microcosms, while BetaandGammaproteobacteria (Burkholderiacea, Comamonadaceae,Oxalobacteraceae, and Xanthomonadaceae) dominated MCPA consumersin cast microcosms, indicating that the latter taxa are prone to respond tospektrum | Tagungsband 2011