Extracellular production and degradation of superoxide in the coral Stylophora pistillata and cultured Symbiodinium

Reactive oxygen species (ROS) are thought to play a major role in cell death pathways and bleaching in scleractinian corals. Direct measurements of ROS in corals are conspicuously in short supply, partly due to inherent problems with ROS quantification in cellular systems. In this study we characterized the dynamics of the reactive oxygen species superoxide anion radical (O(2)(-)) in the external milieu of the coral Stylophora pistillata. Using a sensitive, rapid and selective chemiluminescence-based technique, we measured extracellular superoxide production and detoxification activity of symbiont (non-bleached) and aposymbiont (bleached) corals, and of cultured Symbiodinium (from clades A and C). Bleached and non-bleached Stylophora fragments were found to produce superoxide at comparable rates of 10(-11)-10(-9) mol O(2)(-) mg protein(-1) min(-1) in the dark. In the light, a two-fold enhancement in O(2)(-) production rates was observed in non-bleached corals, but not in bleached corals. Cultured Symbiodinium produced superoxide in the dark at a rate of . Light was found to markedly enhance O(2)(-) production. The NADPH Oxidase inhibitor Diphenyleneiodonium chloride (DPI) strongly inhibited O(2)(-) production by corals (and more moderately by algae), possibly suggesting an involvement of NADPH Oxidase in the process. An extracellular O(2)(-) detoxifying activity was found for bleached and non-bleached Stylophora but not for Symbiodinium. The O(2)(-) detoxifying activity was partially characterized and found to resemble that of the enzyme superoxide dismutase (SOD). The findings of substantial extracellular O(2)(-) production as well as extracellular O(2)(-) detoxifying activity may shed light on the chemical interactions between the symbiont and its host and between the coral and its environment. Superoxide production by Symbiodinium possibly implies that algal bearing corals are more susceptible to an internal build-up of O(2)(-), which may in turn be linked to oxidative stress mediated bleaching.