Reactive oxygen species play an important role in the oxygen-dependent microbicidal system of mammalian leukocytes. In bivalves, however, details of the correlation between generation of reactive oxygen molecules and intracellular killing of bacteria by hemocytes has remained unclear. Focusing on the role of reactive oxygen molecules, we investigated phagocytosis-associated bactericidal system of the Pacific oyster hemocytes. Oyster agranulocytes generate superoxide anion as the first oxygen metabolite by a specific reactive oxygen-forming system. This system and NADPH-oxidase one are similar in functional aspects. Under anaerobic condition, oyster hemocytes showed reduced bactericidal activity against 6 species of bacteria among 14 species tested. By oyster hemocytes, a carotenoid-deficient mutant of Micrococcus luteus is killed more sensitive than wildtype strain under aerobic condition. The killing effects on the mutant decrease significantly in anaerobic atmosphere, however. From these findings, oyster hemocytes have an oxygen-dependent cytotoxic system, although the activity might not be sufficient for a bactericidal system. Bactericidal activity of reactive oxygen is strongly inhibited by addition of catalase. It is suggested that hydrogen peroxide serves an important role in the bactericidal action of oyster hemocytes. Hydroxyl radical is often detected for metal-conjugated form such as crypto-OH radical. In granulocytes, intracellular oxidants are generated by a non-enzyme catalyzing system, which is thought to be regulated by transition metals.