Further Insight into the Mechanism of Human PMN Lysis following Phagocytosis of Staphylococcus aureus.

ABSTRACT

Staphylococcus aureus is an important human pathogen that can cause a variety of diseases ranging from mild superficial skin infections to life-threatening conditions like necrotizing pneumonia, endocarditis, and septicemia. Polymorphonuclear leukocytes (PMNs; neutrophils in particular herein) are essential for host defense against S. aureus infections, and the microbe is phagocytosed readily. Most ingested bacteria are killed, but some S. aureus strains-such as the epidemic USA300 strain-have an enhanced ability to cause PMN lysis after phagocytosis. Although progress has been made, the mechanism for lysis after phagocytosis of S. aureus remains incompletely determined. Here, we tested the hypothesis that disruption of phagosome integrity and escape of S. aureus from the PMN phagosome into the cytoplasm precedes PMN lysis. We used USA300 wild-type and isogenic deletion strains to evaluate and/or verify the role of selected S. aureus molecules in this cytolytic process. Compared to the wild-type USA300 strain, Δagr, Δhla, ΔlukGH, and Δpsm strains each caused significantly less lysis of human PMNs 3 h and/or 6 h after phagocytosis, consistent with previous studies. Most notably, confocal microscopy coupled with selective permeabilization assays demonstrated that phagosome membrane integrity is largely maintained prior to PMN lysis after S. aureus phagocytosis. We conclude that PMN lysis does not require escape of S. aureus from the phagosome to the cytoplasm and that these are independent phenomena. The findings are consistent with the ability of S. aureus (via selected molecules) to trigger lysis of human PMNs by an undetermined signaling mechanism. IMPORTANCE S. aureus strain USA300 has the ability to cause rapid lysis of human neutrophils after phagocytosis. Although this phenomenon likely contributes to the success of USA300 as a human pathogen, our knowledge of the mechanism remains incomplete. Here, we used a selective permeabilization assay coupled with confocal microscopy to demonstrate that USA300 is contained within human neutrophil phagosomes until the point of host cell lysis. Thus, consistent with a process in macrophages, S. aureus fails to escape into the neutrophil cytoplasm prior to cytolysis.