Dark adaptation and ability of pulse-amplitude modulated (PAM) fluorometry to identify nutrient limitation in the bloom-forming cyanobacterium, Microcystis aeruginosa (Kützing)

Abstract

Harmful algal blooms in inland waters are widely linked to excess phosphorus (P) loading, but increasing evidence shows that their growth and formation can also be influenced by nitrogen (N) and iron (Fe). Deficiency in N, P, and Fe differentially affects cellular photosystems and is manifested as changes in photosynthetic yield (F_v/F_m). While F_v/F_m has been increasingly used as a rapid and convenient in situ gauge of nutrient deficiency, there are few rigorous comparisons of instrument sensitivity and ability to resolve specific nutrient stresses. This study evaluated the application of F_v/F_m to cyanobacteria using controlled experiments on a single isolate and tested three hypotheses: i) single F_v/F_m measurements taken with different PAM fluorometers can distinguish among limitation by different nutrients, ii) measurements of F_v/F_m made by the addition of DCMU are comparable to PAM fluorometers, and iii) dark adaptation is not necessary for reliable F_v/F_m measurements. We compared F_v/F_m taken from the bloom-forming Microcystis aeruginosa (UTEX LB 3037) grown in nutrient-replete treatment (R) and N-, P-, and Fe-limited treatments (LN, LP, LFe, respectively), using three pulse-amplitude modulated (PAM) fluorometers and the chemical photosynthesis inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), and evaluated the effects of dark adaptation prior to PAM measurement. There were significant differences in F_v/F_m estimates among PAM fluorometers for light- versus dark-adapted cell suspensions over the whole experiment (21 days), which were all significantly higher than the DCMU-based measurements. However, dark adaptation had no effect on F_v/F_m when comparing PAM-based values across a single nutrient treatment. All F_v/F_m methods could distinguish LN and LP from R and LFe treatments but none were able to resolve LFe from R, or LN from LP cultures. These results indicated that for most PAM applications, dark adaptation is not necessary, and furthermore that single measurements of F_v/F_m do not provide a robust measurement of nutrient limitation in Microcystis aeruginosa UTEX LB 3037, and potentially other, common freshwater cyanobacteria.

Keywords: Cyanobacteria; Microcystis aeruginosa; Nutrient limitation; Photosynthetic yield; Pulse-amplitude modulated fluorometry.