AME 37:159-169 (2004)  -  doi:10.3354/ame037159

ABSTRACT: To determine nutrient availability and growth rates of phytoplankton on a population level, improvement of discriminative power in fluorescence-activated cell sorting is required. We have combined fluorescence of the endogenous photosynthetic pigments chlorophyll a and phycocyanin with a phosphate deficiency (P-deficiency) related stain (enzyme-labelled fluorescence, ELF-97) that yields green fluorescent precipitates at the site of phosphatase activity, referred to as ELF alcohol (ELFA) fluorescence, to sort phytoplankton from Lake Loosdrecht (The Netherlands). Stable isotope labelling with ¹³C-enriched CO₂ enabled assessment of specific growth rates of sorted populations by pyrolytic methylation-gas chromatography and in-line compound specific isotope-ratio mass spectrometry. The dominant population in the lake, the filamentous cyanobacterium Limnothrix sp., was growing under P-deficiency in spring, but the availability of phosphate increased in summer. Continuous flow of phosphate-rich medium into a laboratory-scale enclosure of lake water resulted in washout of the cells with ELFA fluorescence, and increased growth rates. In addition, this study revealed population heterogeneity within the cluster of phycocyanin-containing cyanobacteria. ELFA fluorescence thus reflects the level of P-deficiency of freshwater cyanobacteria and micro-algae, but is modulated by metabolic activity of cells within the population.

KEY WORDS: Alkaline phosphatase · Enzyme-labelled fluorescence · Filamentous cyanobacteria · Fluorescence-activated cell sorting · Population-specific growth rate · Phytoplankton · Stable isotope labelling