Dataset: Cell Abundance Bioassay Experiment
Deployment: SAV-19-02

Sampling and analytical procedures:

 Bioassay experiments were conducted at station 1 and stations 3. At each station, inorganic and organic phosphate amendments were performed on seawater with and without nitrogen enrichment  (NH₄Cl, NaNO₃).  Bioassay Experiments consisted in incubating, over an incubation period of 48h, surface seawater (5m) with inorganic or organic phosphate compounds (20 µM; final concentration of P) including, polyphosphate (polyp), inorganic phosphate (Pi), nucleotides (ATP or AMP) and methylphosphonate (Mepn). In each incubation experiment, a control treatment (surface seawater) was included.

 Samples for cell abundance (2 mL) were subsampled from experimental incubation bottle,  fixed with 0.5% glutaraldehyde (final concentration), flash frozen in liquid nitrogen and then transferred into a -80 °C freezer until analysis. Frozen samples were thawed at room temperature and were analyzed using the Guava EasyCyte HT flow cytometer (Millipore). Instrument specific beads were used to calibrate the cytometer.

Samples were analyzed at a low flow rate (0.24 µL s^-1) during 3 min. For heterotrophic bacteria cell counts, samples were incubated with SYBR Green II solution 1:10 (Molecular Probes) for 15 min in the dark, in order to stain the nucleic acids. Bacterial cells were detected and enumerated based on diagnostic plots of forward scatter vs. green fluorescence.  Group-specific phytoplankton were distinguished based on plots of forward scatter vs. orange fluorescence (phycoerythrin containing Synechococcus sp.), and SSC vs. red fluorescence (eukaryotes).

Instrument:  Cell abundance of both phytoplankton and heterotrophic bacterial cell abundance were determined using the Guava EasyCyte HT flow cytometer (Millipore).

Location:  Northwestern Atlantic surface waters. Depth: surface-50 m.