In the present study, the effect of the high and low virulence S. iniae strains on oxidative activity and cytotoxic effects of trout spleen macrophages were studied. Spleen macrophages were collected from trout. Luminol was used as chemiluminigenic probe. Phorbolmyristate acetate (PMA) also was used as positive control activating agent of respiratory burst and significantly stimulated production of reactive oxygen species (ROS) by normal macrophages. Sensitivity of S. Iniae strains to H2O2 and cytotoxicity of bacterial strains for trout macrophages was also determined using the antibacterial effect of H₂O₂ and the release of lactate dehydrogenase from infected macrophages, respectively. The low virulence strain was killed approximately 6 times more by H₂O₂ compared to the high virulence strain. Results indicate that, high virulence in S. iniae appears to be correlated with higher macrophage cytotoxicity and resistance to ROS.