Effects of Epiphytic Biofilm Activity on the Photosynthetic Activity, pH and Inorganic Carbon Microenvironment of Seagrass Leaves (Zostera marina L.)

Abstract

Epiphytic biofilms on seagrass leaves can lead to extreme microenvironmental conditions for the encapsulated leaf limiting both its photosynthesis and respiration. Yet, little is known about how the biological activity of the biofilm itself changes the seagrass phyllosphere microenvironment and dynamics. We used microsensors to measure O2 concentrations and pH gradients and calculate fluxes of O2, CO2 and bicarbonate (HCO3−) around seagrass leaves (Z. marina L.) covered with artificial, inactive biofilms and natural epiphytic biofilms. A sterilized seawater-agar matrix was used to make an artificial “inactive” biofilm on seagrass leaves with the same thickness as the natural leaf epiphytic biofilm, which impeded turbulent exchange of gases but did not have microbial activity. We compared the concentration profiles and fluxes of O2 and inorganic carbon of the “active” and “inactive” biofilm to investigate the effect of microbial activity and molecular diffusion in seagrass leaf biofilms. In light, the O2 flux of leaves with inactive biofilm was only 31% of the leaves with active biofilm, indicating that the photosynthesis of the microbial community in the biofilm makes up the majority of O2 production in the leaf microenvironment. During darkness, the O2 concentration profiles and O2 fluxes were almost identical in the “active” and “inactive” biofilms. The pH profiles showed the same trend with an increase in pH of ~1.0 in the “active” biofilms and ~0.3 pH units in the “inactive” biofilms in the light, and both showing a decrease of ~0.3 pH units in darkness compared to the bulk seawater. Our measurements thus demonstrate strong photosynthesis in the epiphyte layer driving phyllosphere basification and inorganic carbon limitation. The calculated CO2 concentration on the leaf surface decreased to 0.09 μmol L-1 in the epiphytic biofilm in the light compared to leaf surface CO2 concentrations of 13.8 μmol L-1 on bare seagrass leaves, and the CO2 influxes were only 3.0% and 5.4% of O2 effluxes for leaves with “active” and “inactive” biofilm, respectively. Calculations also showed that HCO3− influxes in light accounted for 91-97% of the total inorganic carbon influx to the seagrass leaf, although the HCO3− utilization via CO2 concentration mechanisms is energy-consuming. Besides increasing mass transfer impedance, leaf epiphytic biofilm activity thus strongly affects the seagrass leaf microenvironment in the light by inducing higher O2 concentration and pH, increasing CO2 limitation and reducing the leaf photosynthetic efficiency.