The TLR2 ligand of O. tsutsugamushi is sensitive to proteinase K, H2O2, and NaOH. (A to D) HEK293 cells transfected with human TLR2 were stimulated with heat-inactivated and crude lysates of O. tsutsugamushi pretreated with the indicated compound. IL-8 concentrations were measured 18 h later by ELISA. Representative results from one of two independent experiments are shown (n = 4 replicates, mean ± SEM). ***, P < 0.001 by one way-ANOVA; ns, no significant difference.