Dataset: Flow cytometric counts from grazing saturation culture experiment using single prey (Isochrysis galbana) and predator (Ochromonas danica) from March to April 2020

This dataset has not been validatedPreliminary and in progressVersion 1 (2023-08-03)Dataset Type:experimental

Principal Investigator: Stephen D. Archer (Bigelow Laboratory for Ocean Sciences)

Co-Principal Investigator: Nicole J. Poulton (Bigelow Laboratory for Ocean Sciences)

BCO-DMO Data Manager: Dana Stuart Gerlach (Woods Hole Oceanographic Institution)


Project: EAGER: A Saturation Approach to Microzooplankton Grazing Rate Determination (Grazing Saturation)


Abstract

Flow cytometric counts of the nanoeukaryote prey Isochrysis galbana, the mixotrophic predator Ochromonas danica and 2 um green fluorescent bead abundance in laboratory culture based experiments to demonstrate the saturation approach.

The culture experiments used the mixotrophic chrysophyte Ochromonas danica, ~ 6 to 9 µm in diameter, as a predator, and the prymnesiophyte Isochrysis galbana, ~ 4 to 6 µm in diameter, as prey. The experiments followed the methods of 'Tests Using Single Prey and Predator Combinations' as described in Archer et al. (2022). 

  • preyIsochyris galbana; AphiaID=573884
  • predatorOchromonas danica, CCMP 1391; AphiaID=1305802 (recommended name of Chlorochromonas danica).

Culture experiment
The cultures were maintained in sterile L1 media in a 21˚C incubator with a 14 h light/10 h dark cycle and light levels of ~ 90 µE m-2 sec-1. Prior to the start of the experiments, O. danica was transitioned from K media and rice to sterile L1 media and then fed every other day on I. galbana. The prey, I. galbana was maintained in semi-continuous growth through regular transfers (2 - 3 days). Similar ratios of predator-to-prey were generated in tubes containing a total volume of 40 ml. Fluorescent polystyrene microspheres (beads) of 2 µm in diameter (Fluoresbrite Plain YG microspheres, Polysciences, Inc., Warrington, PA), were used as surrogate prey.  To minimize clumping, the beads were blocked in a solution of 1 % bovine serum albumin overnight then centrifuged for 5 minutes at 2000 rpm, after which the pellet was resuspended in 0.2 µm filtered seawater. A new solution of beads was prepared at the start of each experiment. During the incubations, the experimental tubes were rotated at 1.2 rpm on a plankton wheel to keep particles in suspension. Two subsamples of 1 ml were removed from each tube at the start (T0) and the final time point (T24) after ~24 hours, for flow cytometric analysis.

Note: It was not possible to accurately obtain Ochromonas danica abundance at the final time point (T24

Flow Cytometric Measurements
Particles were excited with a 488 nm blue excitation laser (100 mW). Data acquisition was triggered on forward scatter (FSC). Signals were recorded from detectors with bandpass filters for right angle light scatter and fluorescence emission in red (692 nm/80 nm band pass) indicative of chlorophyll a, orange for phycoerythrin (593/52 nm), and green (525/35 nm). To ensure accurate calibration of the flow cytometer, ZE5 QC beads (Bio-Rad, Hercules, CA, USA) were run daily. 


Related Datasets

IsRelatedTo

Dataset: Grazing saturation culture experiment #1 using single prey (M.pusilla) and predator (O.danica)
Archer, S. D., Poulton, N. J. (2023) Flow cytometric counts from grazing saturation culture experiment using single prey (Micromonas pusilla) and predator (Ochromonas danica) in October 2019. Biological and Chemical Oceanography Data Management Office (BCO-DMO). (Version 1) Version Date 2023-08-02 http://lod.bco-dmo.org/id/dataset/905469
IsRelatedTo

Dataset: Natural waters experiment with picoeukaryotes and Synechococcus
Archer, S. D., Poulton, N. J. (2023) Flow cytometric counts of picoeukaryotes, Synechococcus, and beads using natural waters from the Gulf of Maine during Jul-Aug 2019 and Jun-Jul 2021. Biological and Chemical Oceanography Data Management Office (BCO-DMO). (Version 1) Version Date 2023-08-04 http://lod.bco-dmo.org/id/dataset/905568

Related Publications

Methods

Archer, S. D., Lubelczyk, L. C., Kunes, M., McPhee, K., Dawydiak, W., Staiger, M., Posman, K. M., & Poulton, N. J. (2022). Saturation Approach to Determine Grazing Mortality in Picoeukaryote and Synechococcus Populations. Frontiers in Marine Science, 9. https://doi.org/10.3389/fmars.2022.844620
Methods

Durand, M. D., & Olson, R. J. (1996). Contributions of phytoplankton light scattering and cell concentration changes to diel variations in beam attenuation in the equatorial Pacific from flow cytometric measurements of pico-, ultra- and nanoplankton. Deep Sea Research Part II: Topical Studies in Oceanography, 43(4–6), 891–906. https://doi.org/10.1016/0967-0645(96)00020-3
Software

FlowJo, LLC. (2023) FlowJo™ Software Version 10.6 [software application] Becton, Dickinson and Company.
Software

R Core Team (2021). R: A language and environment for statistical computing. R v4.0.5. (March 2021) R Foundation for Statistical Computing, Vienna, Austria. URL https://www.R-project.org/