Fig. 4.

Introduction of WalK(S221P) to S. aureus increased vancomycin resistance and attenuated bacterial virulence. E-test of vancomycin susceptibilities in (A) MRSA strains USA300 and K-USA300, and (B) MSSA strains Newman and K-Newman. RT-qPCR detection of the expression levels of virulence factors in (C) USA300 and K-USA300, and (D) Newman and K-Newman. Statistical significance was calculated by 2way ANOVA. * P < 0.05, ** P < 0.01, *** P < 0.001, and ns indicates no significance. (E) Hemolytic experiments. Sheep blood plates were inoculated with USA300 and K-USA300 or strains Newman and K-Newman, cultured at 37 °C overnight, then kept at 4 °C for 24 h and photographed. Western blot analysis of Hla and Hlb levels in (F) USA300 and K-USA300, and (H) Newman and K-Newman. Molecular weights of the protein marker are indicated on the left. LC, loading control. Evaluation of gray values of the expressed Hla and Hlb using ImageJ software. The relative values of Hla/LC and Hlb/LC in (G) USA300 and K-USA300, and (I) Newman and K-Newman were indicated. Statistical significance was calculated by 2way ANOVA. *** P < 0.001.