Factors influencing in vitro killing of bacteria by hemocytes of the eastern oyster (Crassostrea virginica)

A tetrazolium dye reduction assay was used to study factors governing the k illing of bacteria by oyster hemocytes, In vitro tests were performed on ba cterial strains by using hemocytes from oysters collected from the same loc ation in winter and summer. Vibrio parahaemolyticus strains, altered in mot ility or colonial morphology (opaque and translucent), and Listeria monocyt ogenes mutants lacking catalase, superoxide dismutase, hemolysin, and phosp holipase activities were examined in winter and summer. Vibrio vulnificus s trains, opaque and translucent (with and without capsules), were examined o nly in summer. Among V. parahaemolyticus and L. monocytogenes, significantl y (P < 0.05) higher levels of killing by hemocytes were observed in summer than in winter. L. monocytogenes was more resistant than V. parahaemolyticu s or V. vulnificus to the bactericidal activity of hemocytes. In winter, bo th translucent strains of V. parahaemolyticus showed significantly (P < 0.0 5) higher susceptibility to killing by hemocytes than did the wild-type opa que strain. In summer, only one of the V. parahaemolyticus translucent stra ins showed significantly (P < 0.05) higher susceptibility to killing by hem ocytes than did the wild-type opaque strain. No significant differences (P > 0.05) in killing by hemocytes were observed between opaque (encapsulated) and translucent (nonencapsulated) pairs of V. vulnificus. Activities of 19 hydrolytic enzymes were measured in oyster hemolymph collected in winter a nd summer. Only one enzyme, esterase (C4), showed a seasonal difference in activity (higher in winter than in summer). These results suggest that diff erences existed between bacterial genera in their ability to evade killing by oyster hemocytes, that a trait(s) associated with the opaque phenotype m ay have enabled V. parahaemolyticus to evade killing by the oyster's cellul ar defense, and that bactericidal activity of hemocytes was greater in summ er than in winter.