School of Engineering and Science - Jacobs University

CHAPTER III
filtering with a combination of 3 µm GFF + 0.2 µm membrane filter. The water was
stored sterile and dark in the thermo-constant room together with the mesocosms.
Particle-freeness was proved via flow-cytometry (FACSCalibur, Becton & Dickinson)
before each experiment started. Four exact dilutions of 10, 25, 50 and 100% of
undiluted seawater from each mesocosm were prepared. For the incubation three 2.3 L
polycarbonate bottles were gently filled with water from each dilution. The Funnel-
Transfer-Technique appropriate for ciliates (Löder et al., 2010) was used for filling
purposes as these organisms are very sensitive to destruction by vigorous filling and
mixing procedures (Landry, 1993).
To prevent nutrient limitation biases during the phytoplankton bloom sterile filtered
nutrient solutions (F/2 medium, Guillard & Ryther (1962)) were added to the dilution
series (8 x 10 -4 mNO 3 , 1.3 x 10 -5 mPO 4 and 2.4 x 10 -5 mSiO 2 , Experiment 1+2 no SiO 2 ).
One control bottle per mesocosm was incubated without the addition of nutrients.
Copepod grazing set up
The most reliable method to quantify feeding rates of mesozooplankton on both
phytoplankton and non-pigmented microzooplankton, is the analysis of particle removal
in bottle incubations (Båmstedt et al., 2000). Because of interferences with
microzooplankton grazing activity, especially when both micro- and mesozooplankton
prey upon the same species, it is necessary to simultaneously estimate the
microzooplankton grazing rates in separate dilution experiments (Nejstgaard et al.,
1997, Nejstgaard et al., 2001). Thus, for copepod grazing experiments three 2.3 L
bottles per mesocosm (100% undiluted water with added nutrients) were prepared along
with the dilutions and 25 female copepods of the species Temora longicornis were
added to each bottle (~11 copepod L -1 ). This copepod concentration was at the upper
limit of in situ densities in the period March-April (Greve et al., 2004). T. longicornis is
known to be a selective and omnivorous grazer feeding on phytoplankton and
microzooplankton in size classes > 20 µm (Tackx et al., 1990, Maar et al., 2004,
Gentsch et al., 2009). Its role in the planktonic food web makes T. longicornis a key
species and therefore it was selected as copepod grazer in our experiment. These
copepods were caught by vertical net hauls at Helgoland Roads and transferred to the
laboratory immediately. Only actively swimming females of T. longicornis were sorted
out under a dissecting microscope and acclimated to mesocosm conditions for 24 hours
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