Possible physiological mechanisms for production of hydrogen peroxide by the ichthyotoxic flagellate Heterosigma akashiwo

Blooms of the toxic red tide phytoplankton Heterosigma akashiwo (Raphidophy ceae) are responsible for substantial losses within the aquaculture industr y. The toxicological mechanisms of H.akashiwo blooms are complex and to dat e, heavily debated. One putative type of ichthyotoxin includes the producti on of reactive oxygen species (ROS) that could alter gill structure and fun ction, resulting in asphyxiation. In this study, we investigated the potent ial of H.akashiwo to produce extracellular hydrogen peroxide, and have inve stigated which cellular processes are responsible for this production. With in all experiments, H.akashiwo produced substantial amounts of hydrogen per oxide (up to 7.6 pmol min(-1) 10(4) cells(-1)), resulting in extracellular concentrations of similar to 0.5 mu mol l(-1) H2O2. Measured rates of hydro gen peroxide production were directly proportional to cell density, but at higher cell densities, accuracy of H2O2 detection was reduced. Whereas ligh t intensity did not alter H2O2 production, rates of production were stimula ted when temperature was elevated. Hydrogen peroxide production was not onl y dependent on growth phase, but also was regulated by the availability of iron in the medium. Reduction of total iron to 1 nmol l(-1) enhanced the pr oduction of H2O2 relative to iron replete conditions (10 mu mol l(-1) iron) . From this, we collectively conclude that production of extracellular H2O2 by H.akashiwo occurs through a metabolic pathway that is not directly link ed to photosynthesis.