Abstract: Objective To observe the difference in IL-1 production and cytotoxicity induced by recombinant suilysin(rSLY) and non-hemolytic mutant of SLY, rSLY353L in mouse peritoneal macrophages (PM) for the better understanding of the mechanism of inflammasome activation induced by SLY. Methods Lactate dehydrogenase (LDH) release assay was conducted to determine the cytotoxicity of rSLY and rSLY353L. IL-1 production was determined with ELISA. Results The SLY at different concentrations (0.1-10 g) induced significant cytotoxicity in PM in a dose and time dependent manner, whereas rSLY353L did not induce detectable cytotoxicity. In contrast to rSLY353L, 1 g/ml of rSLY induced 750 pg/ml of IL-1 and caused less than 20% of cytotoxicity. 5 g/ml of SLY induced higher level of IL-1 and higher cytotoxicity at the same time. The peak time of IL-1 production by PM induced by rSLY is 20 h after treatment, then the production of IL-1 decreased and the cytotoxicity increased with incubation time. Conclusion SLY (1 g/ml) induced significant IL-1 production and low cytotoxicity by PM at 20 h after treatment which is the optimized condition for further study to understand rSLY-induced inflammasome activation mechanism.