Active Fluorescence Assays of Phytoplankton Physiological State: Population and Individual Cell Measurements

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Olson9819206Active fluorescence techniques such as Fast Repetition Rate (FRR) fluorometry are becoming commonly used to monitor phytoplankton condition in the sea, but many questions remain about the relationships between these measurements of photosynthetic potential and ecologically relevant phytoplankton properties such as growth rate. Photosynthetic parameters such as the fraction of functional reaction centers, the functional absorption cross section of photosystem 2 (PS2), and the rate of electron transfer between PS2 and PSI appear to be valuable diagnostic indicators of specific suboptimal conditions, but it is not clear how quantitatively we can interpret these indicators, or how important a role heterogeneity among phytoplankton species and individuals might play in such interpretation. Olson and Sosik therefore will conduct a series of studies designed to investigate the effects of environmental conditions on phytoplankton growth and active fluorescence parameters, and to determine the extent of variability in active fluorescence responses among phytoplankton species and individual cells in natural populations. Specifically, Olson and Sosik will investigate relationships between nitrogen-, iron-, and light-limitation on phytoplankton growth and photosynthetic parameters as measured by active fluorescence techniques. Observations of wide areas of ocean in which active fluorescence indicates suboptimal phytoplankton condition, and experiments in which growth rate appears to be uncoupled from active fluorescence parameters raise questions about these relationships. Batch and continuous cultures of phytoplankton will be used to study transient and steady-state limitation (including supraoptimal light intensities) of cell growth and the accompanying effects on photochemical energy conversion efficiency of PS2 (i.e., the fraction of functional photosynthetic reaction centers), the functional absorption cross section of PS2, and the rate of electron transfer from PS2 to PS 1. Fluorescence responses of natural populations of phytoplankton to changing environmental conditions will be assessed through observations of local waters, transects between Woods Hole and the Sargasso Sea in spring and summer, and through bottle incubation experiments. The approach the investigators will use focuses on examining inter- and intra-species heterogeneity in active fluorescence parameters, and in active fluorescence responses to environmental conditions. Olson and Sosik will analyze the samples described above by both FRR fluorometry (a bulk method) and individual cell "pump-during-probe" (PDP) techniques. A PDP flow cytometer will be used to measure pico- and nanophytoplankton populations and individual larger cells, and a PDP microfluorometer will be used to analyze individual cells of selected species larger than 5 gm. The distribution of active fluorescence parameters both within and between species will thus be assessed in pure cultures under different growth conditions, including transient and steady state situations. Application of both bulk and individual cell techniques to natural samples will allow the investigators to determine the influence of various phytoplankton groups on the bulk measurements, and analysis of the distribution of properties among individual cells may enable them to deduce the recent nutrient history of the phytoplankton.The combination of bulk and individual-cell measurements of active fluorescence will provide new insights into the regulation and growth of marine phytoplankton, including a new way to evaluate the dynamics of nutrient availability.