6 research outputs found
Comparison of techniques used to count single-celled viable phytoplankton
Author Posting. © The Author(s), 2010. This is the author's version of the work. It is posted here by permission of Springer for personal use, not for redistribution. The definitive version was published in Journal of Applied Phycology 24 (2012): 751-758, doi:10.1007/s10811-011-9694-z.Four methods commonly used to count phytoplankton were evaluated based upon the precision of concentration
estimates: Sedgewick Rafter and membrane filter direct counts, flow cytometry, and flow-based imaging cytometry
(FlowCAM). Counting methods were all able to estimate the cell concentrations, categorize cells into size classes,
and determine cell viability using fluorescent probes. These criteria are essential to determine whether discharged
ballast water complies with international standards that limit the concentration of viable planktonic organisms based
on size class. Samples containing unknown concentrations of live and UV-inactivated phytoflagellates (Tetraselmis
impellucida) were formulated to have low concentrations (<100 ml-1) of viable phytoplankton. All count methods
used chlorophyll a fluorescence to detect cells and SYTOX fluorescence to detect non-viable cells. With the
exception of one sample, the methods generated live and non-viable cell counts that were significantly different
from each other, although estimates were generally within 100% of the ensemble mean of all subsamples from all
methods. Overall, percent coefficient of variation (CV) among sample replicates was lowest in membrane filtration
sample replicates, and CVs for all four counting methods were usually lower than 30% (although instances of ~60%
were observed). Since all four methods were generally appropriate for monitoring discharged ballast water,
ancillary considerations (e.g., ease of analysis, sample processing rate, sample size, etc.) become critical factors for
choosing the optimal phytoplankton counting method.This study was supported by the U.S. Coast Guard Research and Development Center under contract HSCG32-07-
X-R00018. Partial research support to DMA and DMK was provided
through NSF International Contract 03/06/394, and Environmental Protection Agency Grant RD-83382801-0