Single-Cell Analysis of CX3CR1+ Cells Reveals a Pathogenic Role for BIRC5+ Myeloid Proliferating Cells Driven by Staphylococcus aureus Leukotoxins.

Our previous studies identified a population of stem cell-like proliferating myeloid cells within inflamed tissues that could serve as a reservoir for tissue macrophages to adopt different activation states depending on the microenvironment. By lineage-tracing cells derived from CX3CR1+ precursors in mice during infection and profiling by single-cell RNA sequencing, in this study, we identify a cluster of BIRC5+ myeloid cells that expanded in the liver during chronic infection with either the parasite Schistosoma mansoni or the bacterial pathogen Staphylococcus aureus. In the absence of tissue-damaging toxins, S. aureus infection does not elicit these BIRC5+ cells. Moreover, deletion of BIRC5 from CX3CR1-expressing cells results in improved survival during S. aureus infection. Hence the combination of single-cell RNA sequencing and genetic fate-mapping CX3CR1+ cells revealed a toxin-dependent pathogenic role for BIRC5 in myeloid cells during S. aureus infection.

Staphylococcus aureus induces a muted host response in human blood that blunts the recruitment of neutrophils.

Staphylococcus aureus is an opportunistic pathogen and chief among bloodstream-infecting bacteria. S. aureus produces an array of human-specific virulence factors that may contribute to immune suppression. Here, we defined the response of primary human phagocytes following infection with S. aureus using RNA-sequencing (RNA-Seq). We found that the overall transcriptional response to S. aureus was weak both in the number of genes and in the magnitude of response. Using an ex vivo bacteremia model with fresh human blood, we uncovered that infection with S. aureus resulted in the down-regulation of genes related to innate immune response and cytokine and chemokine signaling. This muted transcriptional response was conserved across diverse S. aureus clones but absent in blood exposed to heat-killed S. aureus or blood infected with the less virulent staphylococcal species Staphylococcus epidermidis. Notably, this signature was also present in patients with S. aureus bacteremia. We identified the master regulator S. aureus exoprotein expression (SaeRS) and the SaeRS-regulated pore-forming toxins as key mediators of the transcriptional suppression. The S. aureus-mediated suppression of chemokine and cytokine transcription was reflected by circulating protein levels in the plasma. Wild-type S. aureus elicited a soluble milieu that was restrictive in the recruitment of human neutrophils compared with strains lacking saeRS. Thus, S. aureus blunts the inflammatory response resulting in impaired neutrophil recruitment, which could promote the survival of the pathogen during invasive infection.

Identification of a domain critical for Staphylococcus aureus LukED receptor targeting and lysis of erythrocytes.

Leukocidin ED (LukED) is a pore-forming toxin produced by Staphylococcus aureus, which lyses host cells and promotes virulence of the bacteria. LukED enables S. aureus to acquire iron by lysing erythrocytes, which depends on targeting the host receptor Duffy antigen receptor for chemokines (DARC). The toxin also targets DARC on the endothelium, contributing to the lethality observed during bloodstream infection in mice. LukED is comprised of two monomers: LukE and LukD. LukE binds to DARC and facilitates hemolysis, but the closely related Panton-Valentine leukocidin S (LukS-PV) does not bind to DARC and is not hemolytic. The interaction of LukE with DARC and the role this plays in hemolysis are incompletely characterized. To determine the domain(s) of LukE that are critical for DARC binding, we studied the hemolytic function of LukE-LukS-PV chimeras, in which areas of sequence divergence (divergence regions, or DRs) were swapped between the toxins. We found that two regions of LukE's rim domain contribute to hemolysis, namely residues 57-75 (DR1) and residues 182-196 (DR4). Interestingly, LukE DR1 is sufficient to render LukS-PV capable of DARC binding and hemolysis. Further, LukE, by binding DARC through DR1, promotes the recruitment of LukD to erythrocytes, likely by facilitating LukED oligomer formation. Finally, we show that LukE targets murine Darc through DR1 in vivo to cause host lethality. These findings expand our biochemical understanding of the LukE-DARC interaction and the role that this toxin-receptor pair plays in S. aureus pathophysiology.

Clumping factor B is an important virulence factor during Staphylococcus aureus skin infection and a promising vaccine target.

Staphylococcus aureus expresses a number of cell wall-anchored proteins that mediate adhesion and invasion of host cells and tissues and promote immune evasion, consequently contributing to the virulence of this organism. The cell wall-anchored protein clumping factor B (ClfB) has previously been shown to facilitate S. aureus nasal colonization through high affinity interactions with the cornified envelope in the anterior nares. However, the role of ClfB during skin and soft tissue infection (SSTI) has never been investigated. This study reveals a novel role for ClfB during SSTIs. ClfB is crucial in determining the abscess structure and bacterial burden early in infection and this is dependent upon a specific interaction with the ligand loricrin which is expressed within the abscess tissue. Targeting ClfB using a model vaccine that induced both protective humoral and cellular responses, leads to protection during S. aureus skin infection. This study therefore identifies ClfB as an important antigen for future SSTI vaccines.

A Small Membrane Stabilizing Protein Critical to the Pathogenicity of Staphylococcus aureus.

Staphylococcus aureus is a major human pathogen, and the emergence of antibiotic-resistant strains is making all types of S. aureus infections more challenging to treat. With a pressing need to develop alternative control strategies to use alongside or in place of conventional antibiotics, one approach is the targeting of established virulence factors. However, attempts at this have had little success to date, suggesting that we need to better understand how this pathogen causes disease if effective targets are to be identified. To address this, using a functional genomics approach, we have identified a small membrane-bound protein that we have called MspA. Inactivation of this protein results in the loss of the ability of S. aureus to secrete cytolytic toxins, protect itself from several aspects of the human innate immune system, and control its iron homeostasis. These changes appear to be mediated through a change in the stability of the bacterial membrane as a consequence of iron toxicity. These pleiotropic effects on the ability of the pathogen to interact with its host result in significant impairment in the ability of S. aureus to cause infection in both a subcutaneous and sepsis model of infection. Given the scale of the effect the inactivation of MspA causes, it represents a unique and promising target for the development of a novel therapeutic approach.

IL-10 Plays Opposing Roles during Staphylococcus aureus Systemic and Localized Infections.

IL-10 is a potent anti-inflammatory mediator that plays a crucial role in limiting host immunopathology during bacterial infections by controlling effector T cell activation. Staphylococcus aureus has previously been shown to manipulate the IL-10 response as a mechanism of immune evasion during chronic systemic and biofilm models of infection. In the present study, we demonstrate divergent roles for IL-10 depending on the site of infection. During acute systemic S. aureus infection, IL-10 plays an important protective role and is required to prevent bacterial dissemination and host morbidity by controlling effector T cells and the associated downstream hyperactivation of inflammatory phagocytes, which are capable of host tissue damage. CD19+CD11b+CD5+ B1a regulatory cells were shown to rapidly express IL-10 in a TLR2-dependent manner in response to S. aureus, and adoptive transfer of B1a cells was protective during acute systemic infection in IL-10-deficient hosts. In contrast, during localized s.c. infection, IL-10 production plays a detrimental role by facilitating bacterial persistence via the same mechanism of controlling proinflammatory T cell responses. Our findings demonstrate that induction of IL-10 has a major influence on disease outcome during acute S. aureus infection. Too much IL-10 at one end of the scale may suppress otherwise protective T cell responses, thus facilitating persistence of the bacteria, and at the other end, too little IL-10 may tend toward fatal host-mediated pathology through excessive activation of T cells and associated phagocyte-mediated damage.

The Staphylococcus aureus Cell Wall-Anchored Protein Clumping Factor A Is an Important T Cell Antigen.

Staphylococcus aureus has become increasingly resistant to antibiotics, and vaccines offer a potential solution to this epidemic of antimicrobial resistance. Targeting of specific T cell subsets is now considered crucial for next-generation anti-S. aureus vaccines; however, there is a paucity of information regarding T cell antigens of S. aureus This study highlights the importance of cell wall-anchored proteins as human CD4+ T cell activators capable of driving antigen-specific Th1 and Th17 cell activation. Clumping factor A (ClfA), which contains N1, N2, and N3 binding domains, was found to be a potent human T cell activator. We further investigated which subdomains of ClfA were involved in T cell activation and found that the full-length ClfA N123 and N23 were potent Th1 and Th17 activators. Interestingly, the N1 subdomain was capable of exclusively activating Th1 cells. Furthermore, when these subdomains were used in a model vaccine, N23 and N1 offered Th1- and Th17-mediated systemic protection in mice upon intraperitoneal challenge. Overall, however, full-length ClfA N123 is required for maximal protection both locally and systemically.

Memory Th1 Cells Are Protective in Invasive Staphylococcus aureus Infection.

Mechanisms of protective immunity to Staphylococcus aureus infection in humans remain elusive. While the importance of cellular immunity has been shown in mice, T cell responses in humans have not been characterised. Using a murine model of recurrent S. aureus peritonitis, we demonstrated that prior exposure to S. aureus enhanced IFNγ responses upon subsequent infection, while adoptive transfer of S. aureus antigen-specific Th1 cells was protective in naïve mice. Translating these findings, we found that S. aureus antigen-specific Th1 cells were also significantly expanded during human S. aureus bloodstream infection (BSI). These Th1 cells were CD45RO+, indicative of a memory phenotype. Thus, exposure to S. aureus induces memory Th1 cells in mice and humans, identifying Th1 cells as potential S. aureus vaccine targets. Consequently, we developed a model vaccine comprising staphylococcal clumping factor A, which we demonstrate to be an effective human T cell antigen, combined with the Th1-driving adjuvant CpG. This novel Th1-inducing vaccine conferred significant protection during S. aureus infection in mice. This study notably advances our understanding of S. aureus cellular immunity, and demonstrates for the first time that a correlate of S. aureus protective immunity identified in mice may be relevant in humans.

Prophage-encoded methyltransferase drives adaptation of community-acquired methicillin-resistant Staphylococcus aureus.

We recently described the evolution of a community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) USA300 variant responsible for an outbreak of skin and soft tissue infections. Acquisition of a mosaic version of the Φ11 prophage (mΦ11) that increases skin abscess size was an early step in CA-MRSA adaptation that primed the successful spread of the clone. The present report shows how prophage mΦ11 exerts its effect on virulence for skin infection without encoding a known toxin or fitness genes. Abscess size and skin inflammation were associated with DNA methylase activity of an mΦ11-encoded adenine methyltransferase (designated pamA). pamA increased expression of fibronectin-binding protein A (fnbA; FnBPA), and inactivation of fnbA eliminated the effect of pamA on abscess virulence without affecting strains lacking pamA. Thus, fnbA is a pamA-specific virulence factor. Mechanistically, pamA was shown to promote biofilm formation in vivo in skin abscesses, a phenotype linked to FnBPA's role in biofilm formation. Collectively, these data reveal a novel mechanism-epigenetic regulation of staphylococcal gene expression-by which phage can regulate virulence to drive adaptive leaps by S. aureus.

Secreted mammalian DNases protect against systemic bacterial infection by digesting biofilms.

Extracellular DNase DNASE1L3 maintains tolerance to self-DNA in humans and mice, whereas the role of its homolog DNASE1 remains controversial, and the overall function of secreted DNases in immunity is unclear. We report that deletion of murine DNASE1 neither caused autoreactivity in isolation nor exacerbated lupus-like disease in DNASE1L3-deficient mice. However, combined deficiency of DNASE1 and DNASE1L3 rendered mice susceptible to bloodstream infection with Staphylococcus aureus. DNASE1/DNASE1L3 double-deficient mice mounted a normal innate response to S. aureus and did not accumulate neutrophil extracellular traps (NETs). However, their kidneys manifested severe pathology, increased bacterial burden, and biofilm-like bacterial lesions that contained bacterial DNA and excluded neutrophils. Furthermore, systemic administration of recombinant DNASE1 protein during S. aureus infection rescued the mortality of DNase-deficient mice and ameliorated the disease in wild-type mice. Thus, DNASE1 and DNASE1L3 jointly facilitate the control of bacterial infection by digesting extracellular microbial DNA in biofilms, suggesting the original evolutionary function of secreted DNases as antimicrobial agents.

Single-cell analysis of CX3CR1 + cells reveal a pathogenic role for BIRC5 + myeloid proliferating cells driven by Staphylococcus aureus leukotoxins.

Our previous studies identified a population of stem cell-like proliferating myeloid cells within inflamed tissues that could serve as a reservoir for tissue macrophages to adopt different activation states depending on the microenvironment. By lineage tracing cells derived from CX3CR1 + precursors in mice during infection and profiling by scRNA-seq, here we identify a cluster of BIRC5 + myeloid cells that expanded in the liver during either chronic infection with the parasite Schistosoma mansoni or the bacterial pathogen Staphylococcus aureus . In the absence of tissue damaging toxins, S. aureus infection does not elicit these BIRC5 + cells. Moreover, deletion of BIRC5 from CX3CR1 expressing cells results in improved survival during S. aureus infection. Hence, the combination of scRNA-Seq and genetic fate mapping CX3CR1 + cells revealed a toxin dependent pathogenic role for BIRC5 in myeloid cells during S. aureus infection.

Capsular Polysaccharide Is Essential for the Virulence of the Antimicrobial-Resistant Pathogen Enterobacter hormaechei.

Nosocomial infections caused by multidrug-resistant (MDR) Enterobacter cloacae complex (ECC) pathogens are on the rise. However, the virulence strategies employed by these pathogens remain elusive. Here, we study the interaction of ECC clinical isolates with human serum to define how this pathogen evades the antimicrobial action of complement, one of the first lines of host-mediated immune defense. We identified a small number of serum-sensitive strains, including Enterobacter hormaechei strain NR3055, which we exploited for the in vitro selection of serum-resistant clones. Comparative genomics between the serum-sensitive NR3055 strain and the isolated serum-resistant clones revealed a premature stop codon in the wzy gene of the capsular polysaccharide biosynthesis locus of NR3055. The complementation of wzy conferred serum resistance to NR3055, prevented the deposition of complement proteins on the bacterial surface, inhibited phagocytosis by human neutrophils, and rendered the bacteria virulent in a mouse model of peritonitis. Mice exposed to a nonlethal dose of encapsulated NR3055 were protected from subsequent lethal infections by encapsulated NR3055, whereas mice that were previously exposed to unencapsulated NR3055 succumbed to infection. Thus, capsule is a key immune evasion determinant for E. hormaechei, and it is a potential target for prophylactics and therapeutics to combat these increasingly MDR human pathogens. IMPORTANCE Infections caused by antimicrobial resistant bacteria are of increasing concern, especially those due to carbapenem-resistant Enterobacteriaceae pathogens. Included in this group are species of the Enterobacter cloacae complex, regarding which there is a paucity of knowledge on the infection biology of the pathogens, despite their clinical relevance. In this study, we combine techniques in comparative genomics, bacterial genetics, and diverse models of infection to establish capsule as an important mechanism of Enterobacter pathogens to resist the antibacterial activity of serum, a first line of host defense against bacterial infections. We also show that immune memory targeting the Enterobacter capsule protects against lethal infection. The further characterization of Enterobacter infection biology and the immune response to infection are needed for the development of therapies and preventative interventions targeting these highly antibiotic resistant pathogens.

Rewilding of laboratory mice enhances granulopoiesis and immunity through intestinal fungal colonization.

The paucity of blood granulocyte populations such as neutrophils in laboratory mice is a notable difference between this model organism and humans, but the cause of this species-specific difference is unclear. We previously demonstrated that laboratory mice released into a seminatural environment, referred to as rewilding, display an increase in blood granulocytes that is associated with expansion of fungi in the gut microbiota. Here, we find that tonic signals from fungal colonization induce sustained granulopoiesis through a mechanism distinct from emergency granulopoiesis, leading to a prolonged expansion of circulating neutrophils that promotes immunity. Fungal colonization after either rewilding or oral inoculation of laboratory mice with Candida albicans induced persistent expansion of myeloid progenitors in the bone marrow. This increase in granulopoiesis conferred greater long-term protection from bloodstream infection by gram-positive bacteria than by the trained immune response evoked by transient exposure to the fungal cell wall component ß-glucan. Consequently, introducing fungi into laboratory mice may restore aspects of leukocyte development and provide a better model for humans and free-living mammals that are constantly exposed to environmental fungi.

Multivalent human antibody-centyrin fusion protein to prevent and treat Staphylococcus aureus infections.

Treating and preventing infections by antimicrobial-resistant bacterial pathogens is a worldwide problem. Pathogens such as Staphylococcus aureus produce an array of virulence determinants, making it difficult to identify single targets for the development of vaccines or monoclonal therapies. We described a human-derived anti-S. aureus monoclonal antibody (mAb)-centyrin fusion protein ("mAbtyrin") that simultaneously targets multiple bacterial adhesins, resists proteolysis by bacterial protease GluV8, avoids Fc engagement by S. aureus IgG-binding proteins SpA and Sbi, and neutralizes pore-forming leukocidins via fusion with anti-toxin centyrins, while maintaining Fc- and complement-mediated functions. Compared with the parental mAb, mAbtyrin protected human phagocytes and boosted phagocyte-mediated killing. The mAbtyrin also reduced pathology, reduced bacterial burden, and protected from different types of infections in preclinical animal models. Finally, mAbtyrin synergized with vancomycin, enhancing pathogen clearance in an animal model of bacteremia. Altogether, these data establish the potential of multivalent mAbs for treating and preventing S. aureus diseases.

Microbiome-Independent Effects of Antibiotics in a Murine Model of Nosocomial Infections.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most common causes of hospital-acquired pneumonia. To better manage patients with MRSA pneumonia, we require a greater understanding of the host-pathogen interactions during infection. MRSA research focuses on highly virulent and cytotoxic strains, which demonstrate robust phenotypes in animal models of infection. However, nosocomial infections are often caused by hospital-acquired MRSA (HA-MRSA) isolates that exhibit low cytotoxicity and few or no phenotypes in mice, thereby confounding mechanistic studies of pathogenesis. Consequently, virulence pathways utilized by HA-MRSA in nosocomial pneumonia are largely unknown. Here, we report that conditioning mice with broad-spectrum antibiotics lowers the barrier to pneumonia, thereby transforming otherwise avirulent HA-MRSA isolates into lethal pathogens. HA-MRSA isolates are avirulent in gnotobiotic mice, mimicking results in conventional animals. Thus, the observed enhanced susceptibility to infection in antibiotic-treated mice is not due to depletion of the microbiota. More generally, we found that antibiotic conditioning leads to increased susceptibility to infection by diverse antimicrobial-resistant (AMR) pathogens of low virulence. Treatment with antibiotics leads to dehydration and malnutrition, suggesting a potential role for these clinically relevant and reducible hospital complications in susceptibility to pathogens. In sum, the model described here mitigates the impact of low virulence in immunocompetent mice, providing a convenient model to gain fundamental insight into the pathogenesis of nosocomial pathogens. IMPORTANCE Antimicrobial-resistant (AMR) pathogens are responsible for over 2.8 million infections and over 35,000 deaths per year in the United States. To study these microbes, animal models that are susceptible to these pathogens are required. However, many of these pathogens exhibit low virulence in conventional mice, which has negatively impacted mechanistic studies. Here, we show that mice treated with antibiotics in their drinking water become exquisitely susceptible to low-virulence AMR pathogens. Surprisingly, the increased susceptibility was independent of the impact of antibiotics on the microbiome and seems to be due to an unintended consequence of antibiotic treatment: weight loss due to dehydration and caloric restriction. Unlike other models used to sensitize mice to low-virulence pathogens, our model does not reduce phagocyte numbers. Thus, here, we describe an immunocompetent mouse model to facilitate the identification of novel targets and accelerate the development of preventives and therapeutics to combat infections by AMR pathogens.

The tempo and mode of gene regulatory programs during bacterial infection.

Innate immune recognition of bacterial pathogens is a key determinant of the ensuing systemic response, and host or pathogen heterogeneity in this early interaction can impact the course of infection. To gain insight into host response heterogeneity, we investigate macrophage inflammatory dynamics using primary human macrophages infected with Group B Streptococcus. Transcriptomic analysis reveals discrete cellular states within responding macrophages, one of which consists of four sub-states, reflecting inflammatory activation. Infection with six additional bacterial species-Staphylococcus aureus, Listeria monocytogenes, Enterococcus faecalis, Yersinia pseudotuberculosis, Shigella flexneri, and Salmonella enterica-recapitulates these states, though at different frequencies. We show that modulating the duration of infection and the presence of a toxin impacts inflammatory trajectory dynamics. We provide evidence for this trajectory in infected macrophages in an in vivo model of Staphylococcus aureus infection. Our cell-state analysis defines a framework for understanding inflammatory activation dynamics in response to bacterial infection.

Human OTULIN haploinsufficiency impairs cell-intrinsic immunity to staphylococcal α-toxin.

The molecular basis of interindividual clinical variability upon infection with Staphylococcus aureus is unclear. We describe patients with haploinsufficiency for the linear deubiquitinase OTULIN, encoded by a gene on chromosome 5p. Patients suffer from episodes of life-threatening necrosis, typically triggered by S. aureus infection. The disorder is phenocopied in patients with the 5p- (Cri-du-Chat) chromosomal deletion syndrome. OTULIN haploinsufficiency causes an accumulation of linear ubiquitin in dermal fibroblasts, but tumor necrosis factor receptor-mediated nuclear factor κB signaling remains intact. Blood leukocyte subsets are unaffected. The OTULIN-dependent accumulation of caveolin-1 in dermal fibroblasts, but not leukocytes, facilitates the cytotoxic damage inflicted by the staphylococcal virulence factor α-toxin. Naturally elicited antibodies against α-toxin contribute to incomplete clinical penetrance. Human OTULIN haploinsufficiency underlies life-threatening staphylococcal disease by disrupting cell-intrinsic immunity to α-toxin in nonleukocytic cells.

The cell envelope of Staphylococcus aureus selectively controls the sorting of virulence factors.

Staphylococcus aureus bi-component pore-forming leukocidins are secreted toxins that directly target and lyse immune cells. Intriguingly, one of the leukocidins, Leukocidin AB (LukAB), is found associated with the bacterial cell envelope in addition to secreted into the extracellular milieu. Here, we report that retention of LukAB on the bacterial cells provides S. aureus with a pre-synthesized active toxin that kills immune cells. On the bacteria, LukAB is distributed as discrete foci in two distinct compartments: membrane-proximal and surface-exposed. Through genetic screens, we show that a membrane lipid, lysyl-phosphatidylglycerol (LPG), and lipoteichoic acid (LTA) contribute to LukAB deposition and release. Furthermore, by studying non-covalently surface-bound proteins we discovered that the sorting of additional exoproteins, such as IsaB, Hel, ScaH, and Geh, are also controlled by LPG and LTA. Collectively, our study reveals a multistep secretion system that controls exoprotein storage and protein translocation across the S. aureus cell wall.

Targeting leukocidin-mediated immune evasion protects mice from Staphylococcus aureus bacteremia.

Staphylococcus aureus is responsible for various diseases in humans, and recurrent infections are commonly observed. S. aureus produces an array of bicomponent pore-forming toxins that target and kill leukocytes, known collectively as the leukocidins. The contribution of these leukocidins to impair the development of anti-S. aureus adaptive immunity and facilitate reinfection is unclear. Using a murine model of recurrent bacteremia, we demonstrate that infection with a leukocidin mutant results in increased levels of anti-S. aureus antibodies compared with mice infected with the WT parental strain, indicating that leukocidins negatively impact the generation of anti-S. aureus antibodies in vivo. We hypothesized that neutralizing leukocidin-mediated immune subversion by vaccination may shift this host-pathogen interaction in favor of the host. Leukocidin-immunized mice produce potent leukocidin-neutralizing antibodies and robust Th1 and Th17 responses, which collectively protect against bloodstream infections. Altogether, these results demonstrate that blocking leukocidin-mediated immune evasion can promote host protection against S. aureus bloodstream infection.

The Role of Staphylococcus aureus Virulence Factors in Skin Infection and Their Potential as Vaccine Antigens.

Staphylococcus aureus (S. aureus) causes the vast majority of skin and soft tissue infections (SSTIs) in humans. S. aureus has become increasingly resistant to antibiotics and there is an urgent need for new strategies to tackle S. aureus infections. Vaccines offer a potential solution to this epidemic of antimicrobial resistance. However, the development of next generation efficacious anti-S. aureus vaccines necessitates a greater understanding of the protective immune response against S. aureus infection. In particular, it will be important to ascertain if distinct immune mechanisms are required to confer protection at distinct anatomical sites. Recent discoveries have highlighted that interleukin-17-producing T cells play a particularly important role in the immune response to S. aureus skin infection and suggest that vaccine strategies to specifically target these types of T cells may be beneficial in the treatment of S. aureus SSTIs. S. aureus expresses a large number of cell wall-anchored (CWA) proteins, which are covalently attached to the cell wall peptidoglycan. The virulence potential of many CWA proteins has been demonstrated in infection models; however, there is a paucity of information regarding their roles during SSTIs. In this review, we highlight potential candidate antigens for vaccines targeted at protection against SSTIs.