Staphylococcus aureus is a well-known colonizer of the human skin and nose, but also a human pathogen that causes a wide spectrum of diseases. It is well established that S. aureus secretes an arsenal of virulence factors that have evolved to circumvent the human immune system. A major group of S. aureus virulence factors is the bi-component β-barrel pore-forming toxins, also known as leukocidins. These pore-forming toxins target specific cells of the innate and adaptive immune system by interacting with specific receptors expressed on the cell membrane. Even though still heavily debated, clinical and epidemiological studies suggest the involvement of one of the bi-component toxin, Panton-Valentine Leukocidin (PVL), as an important factor contributing to the epidemic spread and increased virulence of CA-MRSA strains. However, the host- and cell-specificity of PVL and other leukocidins, and the lack of adequate in vivo models, fuels the controversy and impairs the appropriate assessment of their role in S. aureus pathophysiology. Currently, the mechanisms of pore-formation and the contribution of PVL and other leukocidins to S. aureus pathophysiology are incompletely understood. This review summarizes our current understanding of leukocidin pore-formation, knowledge gaps, and highlights recent findings identifying novel host-factors involved in the toxin-host interface. As a result, this review furthers emphasizes the complexity behind S. aureus leukocidin cytotoxicity and the challenges associated in the quest to study and understand these major virulence factors.

Staphylococcal bi-component pore-forming toxins, also known as leukocidins, target and lyse human phagocytes in a receptor-dependent manner. S-components of the leukocidins Panton-Valentine leukocidin (PVL), γ-haemolysin AB (HlgAB) and CB (HlgCB), and leukocidin ED (LukED) specifically employ receptors that belong to the class of G-protein coupled receptors (GPCRs). Although these receptors share a common structural architecture, little is known about the conserved characteristics of the interaction between leukocidins and GPCRs. In this study, we investigated host cellular pathways contributing to susceptibility towards S. aureus leukocidin cytotoxicity. We performed a genome-wide CRISPR/Cas9 library screen for toxin-resistance in U937 cells sensitized to leukocidins by ectopic expression of different GPCRs. Our screen identifies post-translational modification (PTM) pathways involved in the sulfation and sialylation of the leukocidin-receptors. Subsequent validation experiments show differences in the impact of PTM moieties on leukocidin toxicity, highlighting an additional layer of refinement and divergence in the staphylococcal host-pathogen interface. Leukocidin receptors may serve as targets for anti-staphylococcal interventions and understanding toxin-receptor interactions will facilitate the development of innovative therapeutics. Variations in the genes encoding PTM pathways could provide insight into observed differences in susceptibility of humans to infections with S. aureus.

Bacterial pathogens have evolved to secrete strong anti-inflammatory proteins that target the immune system. It was long speculated whether these virulence factors could serve as therapeutics in diseases in which abnormal immune activation plays a role. We adopted the secreted chemotaxis inhibitory protein of Staphylococcus aureus (CHIPS) as a model virulence factor-based therapeutic agent for diseases in which C5AR1 stimulation plays an important role. We show that the administration of CHIPS in human C5AR1 knock-in mice successfully dampens C5a-mediated neutrophil migration during immune complex-initiated inflammation. Subsequent CHIPS toxicology studies in animal models were promising. However, during a small phase I trial, healthy human volunteers showed adverse effects directly after CHIPS administration. Subjects showed clinical signs of anaphylaxis with mild leukocytopenia and increased C-reactive protein concentrations, which are possibly related to the presence of relatively high circulating anti-CHIPS antibodies and suggest an inflammatory response. Even though our data in mice show CHIPS as a potential anti-inflammatory agent, safety issues in human subjects temper the use of CHIPS in its current form as a therapeutic candidate. The use of staphylococcal proteins, or other bacterial proteins, as therapeutics or immune-modulators in humans is severely hampered by pre-existing circulating antibodies.

Bacterial pathogens have evolved to secrete strong anti-inflammatory proteins that target the immune system. It was long speculated whether these virulence factors could serve as therapeutics in diseases in which abnormal immune activation plays a role. We adopted the secreted Chemotaxis Inhibitory Protein of Staphylococcus aureus (CHIPS) as a model virulence factor-based therapeutic agent for diseases in which C5aR1 stimulation plays an important role. We show that administration of CHIPS in human C5aR1 knock-in mice successfully dampens C5a mediated neutrophil migration during immune complex-initiated inflammation. Subsequent CHIPS toxicology studies in animal models were promising. However, during a small phase-I trial, healthy human volunteers showed adverse effects directly after CHIPS administration. Subjects showed clinical signs of anaphylaxis with mild leukocytopenia and increased C-reactive protein concentrations, suggesting an inflammatory response, which are believed to be related to the presence of relatively high circulating anti-CHIPS antibodies. Even though our data in mice shows CHIPS as a potential anti-inflammatory agent, safety issues in human subjects temper the use of CHIPS in its current form as a therapeutic candidate. The use of staphylococcal proteins, or other bacterial proteins, as therapeutics or immune-modulators in humans is severely hampered by pre-existing circulating antibodies.