Staphylococcal alpha-phenol soluble modulins contribute to neutrophil lysis after phagocytosis

Staphylococcus aureus community-acquired (CA) MRSA strains are highly virulent and can cause infections in otherwise healthy individuals. The most important mechanism of the host for clearing S.aureus is phagocytosis by neutrophils and subsequent killing of the pathogen. Especially CA-MRSA strains are very efficient in circumventing this neutrophil killing. Interestingly, only a relative small number of virulence factors have been associated with CA-MRSA, one of which are the phenol soluble modulins (PSMs). We have recently shown that the PSMs are functionally inhibited by serum lipoproteins, indicating that PSMs may exert their cytolytic function primarily in the intracellular environment. To further investigate the intracellular role of the PSMs we measured the effect of the -type and -type PSMs on neutrophil killing after phagocytosis. Using fluorescently labelled S.aureus, we measured bacterial survival after phagocytosis in a plate reader, which was employed next to flow cytometry and time-lapse microscopy. Phagocytosis of the CA-MRSA strain MW2 by human neutrophils resulted in rapid host cell death. Using mutant strains of MW2, we demonstrated that in the presence of serum, the intracellular expression of only the psm operon is both necessary and sufficient for both increasedneutrophil cell death and increased survival of S.aureus. Our results identify PSM peptides as prominent contributors to killing of neutrophils after phagocytosis, a finding with major implications for our understanding of S.aureus pathogenesis and strategies for S.aureus vaccine development.