Production of extracellular superoxide and hydrogen peroxide by five marine species of harmful bloom-forming algae

Abstract

Harmful bloom-forming algae include some of the most prolific microbial producers of extracellular reactive oxygen species (ROS). However, the taxonomic diversity of ROS production, the underlying physiological mechanisms and ecophysiological roles of ROS cycling are not completely characterized among phytoplankton taxa that form harmful algal blooms (HABs). This study examines the extracellular production of the ROS superoxide and hydrogen peroxide by five marine HAB species: Chattonella marina, Heterosigma akashiwo, Karenia brevis, Pseudo-nitzschia sp. and Aureococcus anophagefferens. All species produced extracellular superoxide and hydrogen peroxide. Rates of ROS production per cell spanned several orders of magnitude and varied inversely with cell density, suggesting a potential signaling role for extracellular ROS. ROS production was also detected in the spent media of all cultures except K. brevis, indicating the presence of cell-free ROS-generating constituents, such as enzymes or metabolites, which could be further investigated as molecular targets for tracking ROS production in laboratory and field settings. Finally, ratios of superoxide to hydrogen peroxide production could not be accounted for by superoxide dismutation alone, except in the case of K. brevis, indicating a diversity of ROS production and degradation pathways that may ultimately help illuminate the functions of HAB-derived ROS.

Keywords: Aureococcus anophagefferens; Karenia brevis; Pseudo-nitzschia; hydrogen peroxide; reactive oxygen species; superoxide.

Fig. 1.

Cell-associated rates of extracellular superoxide (O₂⁻) and hydrogen peroxide (H₂O₂) production by the organisms examined in this study.

Fig. 2.

Cell density dependence of extracellular superoxide (O₂⁻) and hydrogen peroxide (H₂O₂) production. Spearman’s rank correlation coefficient (ρ) and associated P-value describe the degree of monotonicity in the relationship between cell-normalized superoxide production rates and cell density.

Fig. 3.

Comparison of superoxide and hydrogen peroxide production rates in diluted cell suspensions and cell-free filtrates. C. marina (Cm), Pseudo-nitzschia sp. (P-n), A. anophagefferens (Aa), K. brevis (Kb), and H. akashiwo (Ha). Error bars represent the standard error of the mean of biological replicates. The diagonal line represents the 2:1 molar ratio of superoxide to hydrogen peroxide production expected from the dismutation of superoxide.