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The opportunistic human pathogenStaphylococcus aureuscauses serious infectious diseases ranging from superficial skin and soft tissue infections to necrotizing pneumonia and sepsis. While classically regarded as extracellular pathogen,S. aureusis able to invade and survive within human cells. Host cell exit is associated with cell death, tissue destruction and spread of infection. The exact molecular mechanism employed byS. aureusto escape the host cell is still unclear. In this study, we performed a genome-wide shRNA screen and identified the calcium signaling pathway to be involved in intracellular infection.S. aureusinduced a massive cytosolic Ca2+-increase in epithelial host cells after invasion and intracellular replication of the pathogen. This was paralleled by decrease in endoplasmic reticulum Ca2+-concentration. Additionally, calcium ions from the extracellular space contributed to the cytosolic Ca2+-increase. As a consequence, we observed that the cytoplasmic Ca2+-rise led to increase in mitochondrial Ca2+-concentration, the activation of calpains and caspases and eventually to cell lysis ofS. aureus-infected cells. Our study therefore suggests that intracellularS. aureusdisturbs the host cell Ca2+-homeostasis and induces cytoplasmic Ca2+-overload, which results in both apoptotic and necrotic cell death in parallel or succession.ImportanceDespite being regarded as an extracellular bacterium, the pathogenStaphylococcus aureuscan invade and survive within human cells. The intracellular niche is considered as hide-out from the host immune system and antibiotic treatment and allows bacterial proliferation. Subsequently, the intracellular bacterium induces host cell death, which may facilitate spread of infection and tissue destruction. So far, host cell factors exploited by intracellularS. aureusto promote cell death are only poorly characterized. We performed a genome-wide screen and found the calcium signaling pathway to play a role inS. aureusinvasion and cytotoxicity. The intracellular bacterium induces a cytoplasmic and mitochondrial Ca2+-overload, which results in host cell death. Thus, this study firstly showed how an intracellular bacterium perturbs the host cell Ca2+-homeostasis.

Staphylococcus aureusis a major human pathogen, which can invade and survive in non-professional and professional phagocytes. Intracellularity is thought to contribute to pathogenicity and persistence of the bacterium. Upon internalization by epithelial cells, cytotoxicS. aureusstrains can escape from the phagosome, replicate in the cytosol and induce host cell death. Here, we identified a staphylococcal cysteine protease to induce cell death by intracellularS. aureusafter translocation into the host cell cytoplasm. We demonstrated that loss of staphopain A function leads to delayed onset of host cell death and prolonged intracellular replication ofS. aureusin epithelial cells. Overexpression of staphopain A in a non-cytotoxic strain facilitated intracellular killing of the host cell even in the absence of detectable intracellular replication. Moreover, staphopain A contributed to efficient colonization of the lung in a mouse pneumonia model. Our study suggests that staphopain A is utilized byS. aureusto mediate escape from the host cell and thus contributes to tissue destruction and dissemination of infection.Author SummaryStaphylococcus aureusis a well-known antibiotic-resistant pathogen that emerges in hospital and community settings and can cause a variety of diseases ranging from skin abscesses to lung inflammation and blood poisoning. The bacterium asymptomatically colonizes the upper respiratory tract and skin of about one third of the human population and takes advantage of opportune conditions, like immunodeficiency or breached barriers, to cause infection. AlthoughS. aureusis not regarded as a professional intracellular bacterium, it can be internalized by human cells and subsequently exit the host cells by induction of cell death, which is considered to cause tissue destruction and spread of infection. The bacterial virulence factors and underlying molecular mechanisms involved in the intracellular lifestyle ofS. aureusremain largely unknown. We identified a bacterial cysteine protease to contribute to host cell death mediated by intracellularS. aureus. Staphopain A induced killing of the host cell after translocation of the pathogen into the cell cytosol, while bacterial proliferation was not required. Further, the protease enhanced survival of the pathogen during lung infection. These findings reveal a novel, intracellular role for the bacterial protease staphopain A.

Staphylococcus aureus is a notorious opportunistic pathogen causing a plethora of diseases. Recent research established that once phagocytosed by neutrophils and macrophages, a certain percentage of S. aureus is able to survive within these phagocytes which thereby even may contribute to dissemination of the pathogen. S. aureus further induces its uptake by otherwise non-phagocytic cells and the ensuing intracellular cytotoxicity is suggested to lead to tissue destruction, whereas bacterial persistence within cells is thought to lead to immune evasion and chronicity of infections. We here review recent work on the S. aureus host pathogen interactions with a focus on the intracellular survival of the pathogen.