One pulse, one light curve: Fast characterization of the light response of microphytobenthos biofilms using chlorophyll fluorescence

Abstract

Research on microphytobenthos (MPB) photosynthesis has increasingly relied on pulse amplitude modulation fluorometry, often through the generation of light response curves of the relative electron transport of PSII (rETR), generated through the sequential exposure to a range of actinic irradiances. However, fast, vertical phototactic responses by the motile diatoms that dominate these biofilms occur during the generation of sequential light curves. Light-induced vertical migration is known to confound the characterization of the inherent physiological response of the cells forming the biofilm. The alternative approach of shortening the duration of the light curve protocol prevents the reaching of a physiological steady state and results in light curves heavily dependent on initial light conditions. This study tested the application to MPB of a recently introduced method allowing for the fast generation of non-sequential light curves, based on steady state and temporally independent measurements (“single-pulse light curves,” SPLC). This method was compared to the commonly used sequential protocols regarding its ability to (1) measure a steady-state light response, (2) minimize the effects of vertical migration, and (3) minimize the effects of photoacclimation state induced by initial conditions. The results indicate that the SPLC method can be optimized by applying one light exposure period of just 60 s prior to the generation of the light curve, resulting in a clearly advantageous alternative to the sequential protocols. It effectively allows characterizing the steady-state light response of MPB samples while avoiding the effects of light-induced vertical migration and of the initial light conditions on light curve parameters.