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Quantification of the role of reactive oxygen species, phycotoxins and fatty acids in fish toxicity by harmful marine microalgae remains inconclusive. An  in vitro  fish gill (from rainbow trout  Oncorhynchus mykiss ) assay was used to simultaneously assess the effect in superoxide dismutase, catalase and lactate dehydrogenase enzymatic activities caused by seven species of ichthyotoxic microalgae ( Chattonella marina ,  Fibrocapsa japonica ,  Heterosigma akashiwo ,  Karenia mikimotoi ,  Alexandrium catenella ,  Karlodinium veneficum ,  Prymnesium parvum ). Quantification of superoxide production by these algae was also performed. The effect of purified phycotoxins and crude extracts was compared, and the effect of fatty acids is discussed. The raphidophyte  Chattonella  was the most ichthyotoxic (gill cell viability down to 35%) and also the major producer of superoxide radicals (14 pmol cell -1  hr -1 ) especially after cell lysis. The raphidophyte  Heterosigma  and dinoflagellate  Alexandrium  were the least toxic and had low superoxide production, except when  A .  catenella  was lysed (5.6 pmol cell -1  hr -1 ). Catalase showed no changes in activity in all the treatments. Superoxide dismutase (SOD) and lactate dehydrogenase exhibited significant activity increases of ≤23% and 51.2% TCC (total cellular content), respectively, after exposure to  C .  marina , but SOD showed insignificant changes with remaining algal species. A strong relationship between gill cell viability and superoxide production or superoxide dismutase was not observed. Purified brevetoxins PbTx-2 and -3 (from  Karenia brevis , LC 50  of 22.1 versus 35.2 μg mL -1 ) and karlotoxin KmTx-2 (from  Karlodinium ; LC 50  = 380 ng mL -1 ) could almost entirely account for the fish killing activity by those two dinoflagellates. However, the paralytic shellfish toxins (PST) GTX1&4, C1&C2, and STX did not account for  Alexandrium  ichthyotoxicity. Only aqueous extracts of  Alexandrium  were cytotoxic (≤65% decrease of viability), whereas crude methanol and acetone extracts of  Chattonella ,  Fibrocapsa ,  Heterosigma ,  Karlodinium  and  Prymnesium  decreased cell viability down to 0%. These and our previous findings involving the role of fatty acids confirm that superoxide radicals are only partially involved in ichthyotoxicity and point to a highly variable contribution by other compounds such as lipid peroxidation products (e.g. aldehydes).

Progress in Understanding Algal Bloom-Mediated Fish Kills: The Role of Superoxide Radicals, Phycotoxins and Fatty Acids