Staphylococcus aureus has evolved a sophisticated regulatory system to control its virulence. One of the main roles of this interconnected network is to sense and respond to diverse environmental signals by altering the synthesis of virulence components required for survival in the host, including cell surface adhesins, extracellular enzymes and toxins. The accessory gene regulator (agr), a quorum sensing system that detects the local concentration of a cyclic peptide signaling molecule, is one of the well-studied of these S. aureus regulatory mechanisms. By using this system, S. aureus is able to sense its own population density and translate this information into a specific pattern of gene expression. In addition to Agr, this pathogen senses specific stimuli through various two-component systems and synchronizes responses with alternative sigma factors and cytoplasmic regulators of the SarA protein family. These different regulatory mechanisms combine host and environmental information into a network that guarantees the best possible response of pathogens to changing circumstances. In this article, an overview of the most significant and thoroughly studied regulatory systems of S. aureus is provided, along with a summary of their roles in host interactions.

Novel strategies to counter multidrug-resistant pathogens such as methicillin-resistant Staphylococcus aureus are urgently required. The antimicrobial properties of fatty acids (FAs) have long been recognized and offer significant promise as viable alternatives to, or potentiators of, conventional antibiotics. In this review, we examine the interplay between FAs and S. aureus, specifically detailing the underlying molecular mechanisms responsible for FA-mediated inhibition and the counteracting staphylococcal systems evolved to withstand FA onslaught. Finally, we present an update on the recent therapeutic FA applications to combat S. aureus infection, either as a monotherapy or in combination with antibiotics or host-derived antimicrobial peptides. Given the frequency of interaction between FAs and S. aureus during host colonization and infection, understanding FA mode of action and deciphering S. aureus FA resistance strategies are central in rationally designing future anti-staphylococcal FAs and FA-combination therapies.

Staphylococcus aureus is a versatile pathogen capable of causing a wide range of infections, from minor skin infections to life-threatening invasive diseases. The pathogenicity of S. aureus is attributed to its ability to produce various virulence factors, including adhesion and biofilm-related proteins. Understanding the prevalence and distribution of these genes among S. aureus isolates from different sources is crucial for devising effective strategies to combat biofilm-associated contamination. In this study, we conducted a comprehensive network meta-analysis to assess the prevalence of adhesion and biofilm-related genes in S. aureus isolates and investigate the impact of the isolate source on their occurrence. A systematic search of multiple databases was performed, and a total of 53 relevant studies were included. The prevalence of adhesion and biofilm-related genes in S. aureus isolates was determined, with the highest prevalence observed for clfB (p-estimate = 85.4, 95% confidence interval [CI] 78-90.6), followed by eno (p-estimate = 81.1, 95% CI 61.7-91.9), and icaD (p-estimate = 77, 95% CI 68.6-83.6). Conversely, bap and bbp genes exhibited the lowest prevalence rates (p-estimate = 6.7 and 18.7, respectively). The network meta-analysis allowed us to examine the pairwise co-study of adhesion and biofilm-related genes in S. aureus isolates. The most frequently co-studied gene pairs were icaA-icaD (30 times) and fnbA-fnbB (25 times). Subgroup analysis showed that the occurrence of icaC and icaB genes was significantly lower in animal isolates compared to human and food isolates (p < 0.05). It is worth noting that there was limited data available for the analysis of sasG, bbp, bap, eno, and fib genes. In conclusion, the study revealed varying prevalence rates of adhesion and biofilm-related genes in S. aureus isolates. Genes such as clfB, eno, and icaD were found to be highly prevalent, while bap and bbp were less common. Limited existing data on the prevalence of genes like sasG, bbp, bap, eno, and fib highlights the need for further research to determine their exact prevalence rates. Our results contribute to a better understanding of S. aureus pathogenesis and can facilitate the development of effective strategies for the prevention and treatment of S. aureus infections.

Canine superficial pyoderma is a common bacterial skin infection of dogs, generally caused by Staphylococcus pseudintermedius. The C4 strain of Staphylococcus felis was recently discovered to have strong antimicrobial activity against S. pseudintermedius in mice.

We aimed to evaluate in vitro if this antimicrobial activity was maintained using a novel canine skin explant model.

Punch biopsies (8 mm) of skin from recently euthanised dogs were collected and placed into six-well plates on top of an agarose pedestal.

Histological examination of the skin explants showed an intact dermal-epidermal organisation and a stratum corneum that was successfully colonised by S. pseudintermedius after topical application. The number of colony forming units of S. pseudintermedius showed a 2 log increase after 24 h colonisation, indicating that the explant supported bacterial growth. By contrast, co-treatment with S. felis C4 live bacteria and its sterile protein product significantly reduced the growth of a methicillin-susceptible (ST540, p = 0.0357) and a methicillin-resistant (MR) strain (ST71, p = 0.0143) of S. pseudintermedius. No detectable bacteria were recovered from or visualised on skin 24 h posttreatment with the S. felis C4 sterile protein product.

Using a novel canine explant model, we demonstrate that the S. felis C4 strain inhibits the growth of S. pseudintermedius and that it is a promising candidate for a new probiotic therapy to treat cutaneous infections caused by S. pseudintermedius, including MR strains.

Our previous studies revealed the existence of a Universal Receptive System that regulates interactions between cells and their environment. This system is composed of DNA- and RNA-based Teazeled receptors (TezRs) found on the surface of prokaryotic and eukaryotic cells, as well as integrases and recombinases. In the current study, we aimed to provide further insight into the regulatory role of TezR and its loss in Staphylococcus aureus gene transcription. To this end, transcriptomic analysis of S. aureus MSSA VT209 was performed following the destruction of TezRs. Bacterial RNA samples were extracted from nuclease-treated and untreated S. aureus MSSA VT209. After destruction of the DNA-based-, RNA-, or combined DNA- and RNA-based TezRs of S. aureus, 103, 150, and 93 genes were significantly differently expressed, respectively. The analysis revealed differential clustering of gene expression following the loss of different TezRs, highlighting individual cellular responses following the loss of DNA- and RNA-based TezRs. KEGG pathway gene enrichment analysis revealed that the most upregulated pathways following TezR inactivation included those related to energy metabolism, cell wall metabolism, and secretion systems. Some of the genetic pathways were related to the inhibition of biofilm formation and increased antibiotic resistance, and we confirmed this at the phenotypic level using in vitro studies. The results of this study add another line of evidence that the Universal Receptive System plays an important role in cell regulation, including cell responses to the environmental factors of clinically important pathogens, and that nucleic acid-based TezRs are functionally active parts of the extrabiome.

Staphylococcus aureus is an important pathogen in healthcare facilities, with its resistance to a number of antibiotics currently being a global concern. In this report the presence of S.aureus, resistance gene virulence and antibiotic susceptibility profiles were determined in the mobile phones of senior nursing students. S.aureus was isolated in 11.84% (9/76) of the samples. Furthermore, 44.44% of the mobile phones carried the mecA (MRSA) gene, while none carried the vanA gene. Virulence genes identified were 100% hla, 88.89% hlb, 22.22% tst and sec, and 11.11% sea. The antibiogram revealed that 33.33% of the strains were resistant to cefoxitin and 44.44% showed inducible resistance to clindamycin (ICRSA). The mobile phones of senior nursing students represent an important reservoir of drug-resistant and virulent strains of S.aureus, which could act as infectious foci for the transmission of this pathogen.

The effect of cosmetic ingredients on growth and virulence factor expression in Staphylococcus aureus may vary between culture medium and skin. Researchers have used an in vitro skin model with human heel callus to assess bacterial survival and growth on the stratum corneum of the epidermis. Here, we reconstituted a skin model using keratin as a base (instead of callus) and compared it with brain heart infusion (BHI) medium. We investigated the effects of five cosmetic ingredients (macadamia nut oil, sodium myristoyl methyl taurate, methyl p-hydroxybenzoate, 2-phenoxyethanol, and zinc oxide) on growth and virulence factor expression in S. aureus. Interestingly, the survival pattern of S. aureus in our skin model was similar to that reported in models using callus. Upon the addition of cosmetic ingredients to BHI or skin model medium, the sensitivity of S. aureus to these cosmetic ingredients differed between the two media. Notably, after adding the two tested cosmetic ingredients, the expression level of staphylococcal enterotoxin A in S. aureus reduced significantly in skin model medium compared with that in the BHI medium. Additionally, the expression levels of other S. aureus virulence factors (RNAIII, icaA, and hlb) differed between the two media. These findings suggest that our skin model is a valuable tool for evaluating the effects of cosmetic ingredients on growth and virulence factor expression in S. aureus.

Antimicrobial resistance is a global public health crisis that undermines the efficacy of treatments for infectious diseases, contributing to higher healthcare costs. Among Gram-positive bacteria, Staphylococcus aureus poses significant challenges due to its ability to develop resistance to multiple antibiotics, particularly in food and healthcare settings. Biofilm formation by S. aureus further enhances its resistance and pathogenicity. This study investigated the effects of 126 halogenated compounds on S. aureus biofilms, identifying five potent halogenated phenols. Among these, 2,4,6-triiodophenol (2,4,6-TIP) emerged as the most effective, exhibiting strong biofilm inhibition at a minimum inhibitory concentration (MIC) of 5 μg mL-1. Additionally, 2,4,6-TIP demonstrated efficacy against biofilms formed by methicillin-resistant S. aureus MW2 and various Gram-negative bacteria, including Vibrio parahaemolyticus and uropathogenic Escherichia coli (UPEC), as well as the fungal species Candida albicans. It also prevented the formation of polymicrobial biofilms involving S. aureus and C. albicans. Beyond its antibiofilm properties, 2,4,6-TIP was effective in controlling key virulence factors in S. aureus, such as metabolic, hemolysis and protease activities. It also reduced swimming motility in V. parahaemolyticus and UPEC, and impaired hyphal formation in C. albicans. Transcriptomic analysis further revealed that 2,4,6-TIP significantly repressed the gene expression of RNAIII, a key regulator of biofilm and virulence production in S. aureus. Furthermore, in silico analysis, plant and nematode models showed that 2,4,6-TIP exhibited reduced toxicity compared to phenol. These findings unveiled the strong antimicrobial potential of 2,4,6-TIP and suggest a broad-spectrum capacity to target the virulent characteristics of medically important pathogens. It also highlights that strategic halogenation may play a critical role in enhancing the activity of phenolic compounds while alleviating their toxicity profiles. t.

The adhesion of bacteria to surfaces is associated with physicochemical and biological interactions. The present investigations provide new results about the differential adhesion levels of skin bacteria using a representative 3D skin model which mainly relies on the different physicochemical properties of the respective surfaces. Modulation of the adhesion of bacteria and thus their colonization, may occur by adjusting the physicochemical properties of the epidermal and bacterial surfaces. Lewis acid and hydrophobicity were the most strongly correlated parameters with the antiadhesion properties of the tested compounds. Modulation of physicochemical properties appears to be the primary driver of reduced Staphylococcus aureus adhesion in this study, with no significant changes observed in the expression of genes associated with classical adhesion pathways.

Our previous studies revealed the existence of a Universal Receptive System that regulates interactions between cells and their environment. This system is composed of DNA- and RNA-based Teazeled receptors (TezRs) found on the surface of prokaryotic and eukaryotic cells, as well as integrases and recombinases.. In the current study, we aimed to provide further insight into the regulatory role of TezR and its loss in Staphylococcus aureus gene transcription. To this end, transcriptomic analysis of S. aureus MSSA VT209 was performed following the destruction of TezRs. Bacterial RNA samples were extracted from nuclease-treated and untreated S. aureus MSSA VT209. After destruction of the DNA-based-, RNA-, or combined DNA- and RNA-based TezRs of S. aureus , 103, 150, and 93 genes were significantly differently expressed, respectively. The analysis revealed differential clustering of gene expression following the loss of different TezRs, highlighting individual cellular responses following the loss of DNA- and RNA-based TezRs. KEGG pathway gene enrichment analysis revealed that the most upregulated pathways following TezR inactivation included those related to energy metabolism, cell wall metabolism, and secretion systems. Some of the genetic pathways were related to the inhibition of biofilm formation and increased antibiotic resistance, and we confirmed this at the phenotypic level using in vitro studies. The results of this study add another line of evidence that the Universal Receptive System plays an important role in cell regulation, including cell responses to the environmental factors of clinically important pathogens, and that nucleic acid-based TezRs are functionally active parts of the extrabiome.

Staphylococcus aureus has been involved in transfusion-transmitted fatalities associated with platelet concentrates (PCs) due to its heightened pathogenicity enhanced by genome-encoded virulence and antibiotic resistance genes. This may be facilitated by mobile genetic elements (MGEs) that can cause rearrangements. Several factors contribute to S. aureus virulence, including the type VII secretion system (T7SS), composed of six core genes conserved across S. aureus strains. In this study, we conducted comparative genome analyses of five S. aureus isolates from PCs (CI/BAC/25/13 /W, PS/BAC/169/17 /W and PS/BAC/317/16 /W were detected during PCs screening with the BACT/ALERT automated culture system, and ATR-20003 and CBS2016-05 were missed during screening and caused septic transfusion reactions). Multiple alignments of the genomes revealed evidence of rearrangements involving phage Sa3int in PS/BAC/169/17 /W and PS/BAC/317/16 /W. While the former had undergone translocation of its immune evasion cluster (IEC), the latter had lost part of the phage, leaving behind the IEC. This observation highlights S. aureus genome plasticity. Unexpectedly, strain CBS2016-05 was found to encode a pseudo-type VII secretion system (T7SS) that had lost five of the conserved core genes (esxA, esaA, essA, esaB and essB) and contained a 5' truncated essC. Since these genes are essential for the function of the T7SS protein transport machinery, which plays a key role in S. aureus virulence, CBS2016-05 probably compensates by recruiting other export mechanisms and/or alternative virulence factors, such as neu-tralizing immunity proteins. This study unravels genome rearrangements in S. aureus isolated from PCs and reports the first S. aureus isolate lacking conserved T7SS core genes.

Staphylococcus aureus acts both as a colonizing commensal bacterium and invasive pathogen. Nasal colonization is associated with an increased risk of infection caused by the identical strain. In patients with atopic dermatitis (AD), the degree of S. aureus colonization is associated with the severity of the disease. Here, we comparatively analyzed the in vivo transcriptional profile of S. aureus colonizing the nose and non-diseased skin (non-lesional skin) as opposed to the diseased skin (lesional skin-defined here as infection) of 12 patients with AD. The transcriptional profile during the asymptomatic colonization of the nose closely resembled that of the lesional skin samples for many of the genes studied, with an elevated expression of the genes encoding adhesion-related proteins and proteases. In addition, the genes that modify and remodel the cell wall and encode proteins that facilitate immune evasion showed increased transcriptional activity. Notably, in a subgroup of patients, the global virulence regulator Agr (accessory gene regulator) and downstream target genes were inactive during nasal colonization but upregulated in the lesional and non-lesional skin samples. Taken together, our results demonstrate a colonization-like transcriptional profile on diseased skin and suggest a role for the peptide quorum sensing system Agr during the transition from asymptomatic nasal colonization to skin colonization/infection.

Staphylococcus aureus is a Gram-positive pathogen responsible for the majority of skin and soft tissue infections (SSTIs). S. aureus colonizes the anterior nares of approximately 20%-30% of the population and transiently colonizes the skin, thereby increasing the risk of developing SSTIs and more serious infections. Current laboratory models that mimic the skin surface environment are expensive, require substantial infrastructure, and limit the scope of bacterial physiology studies under human skin conditions. To overcome these limitations, we developed a cost-effective, open-source, chemically defined media recipe termed skin-like medium (SLM) that incorporates key aspects of the human skin surface environment and supports growth of several staphylococcal species. We utilized SLM to investigate the transcriptional response of methicillin-resistant Staphylococcus aureus (MRSA) following growth in SLM compared to a commonly used laboratory media. Through RNA-seq analysis, we observed the upregulation of several virulence factors, including genes encoding functions involved in adhesion, proteolysis, and cytotoxicity. To further explore these findings, we conducted quantitative reverse transcription-PCR (qRT-PCR) experiments to determine the influence of media composition, pH, and temperature on the transcriptional response of key factors involved in adhesion and virulence. We also demonstrated that MRSA primed in SLM adhered better to human corneocytes and demonstrated adhesin-specific phenotypes that previously required genetic manipulation. This improved adherence to corneocytes was dependent on both acidic pH and growth in SLM. These results support the potential utility of SLM as an in vitro model for assessing staphylococcal physiology and metabolism on human skin.

Staphylococcus aureus is the major cause of skin diseases, and its increased prevalence in skin colonization and infections present a need to understand its physiology in this environment. The work presented here outlines S. aureus upregulation of colonization and virulence factors using a newly developed medium that strives to replicate the human skin surface environment and demonstrates roles for adhesins clumping factor A (ClfA), serine-rich repeat glycoprotein adhesin (SraP), and the fibronectin binding proteins (Fnbps) in human corneocyte adherence.

Staphylococcus aureus is a significant human pathogen with a formidable propensity for antibiotic resistance. Worldwide, it is the leading cause of skin and soft tissue infections (SSTI), septic arthritis, osteomyelitis, and infective endocarditis originating from both community- and healthcare-associated settings. Although often grouped by methicillin resistance, both methicillin-resistant (MRSA) and methicillin-sensitive (MSSA) strains are known to cause significant pathologies and injuries. Virulence factors and growing resistance to antibiotics play major roles in the pathogenicity of community-associated strains. In our study, we examined the genetic variability and acquired antibiograms of 122 S. aureus clinical isolates from SSTI, blood, and urinary tract infections originating from pediatric patients within the southeast region of Virginia, USA. We identified a suite of clinically relevant virulence factors and evaluated their prevalence within these isolates. Five genes (clfA, spA, sbi, scpA, and vwb) with immune-evasive functions were identified in all isolates. MRSA isolates had a greater propensity to be resistant to more antibiotics as well as significantly more likely to carry several virulence factors compared to MSSA strains. Further, the carriage of various genes was found to vary significantly based on the infection type (SSTI, blood, urine).

Staphylococcus aureus is a Gram-positive pathogen responsible for the majority of skin and soft tissue infections (SSTIs). S. aureus colonizes the anterior nares of approximately 20-30% of the population and transiently colonizes the skin, thereby increasing the risk of developing SSTIs and more serious infections. Current laboratory models that mimic the skin surface environment are expensive, require substantial infrastructure, and limit the scope of bacterial physiology studies under human skin conditions. To overcome these limitations, we developed a cost-effective, open-source, chemically defined media recipe termed skin-like media (SLM) that incorporates key aspects of the human skin surface environment and supports growth of several Staphylococcal species. We utilized SLM to investigate the transcriptional response of methicillin-resistant S. aureus (MRSA) following growth in SLM compared to a commonly used laboratory media. Through RNA-seq analysis, we observed the upregulation of several virulence factors, including genes encoding functions involved in adhesion, proteolysis, and cytotoxicity. To further explore these findings, we conducted qRT-PCR experiments to determine the influence of media composition, pH, and temperature on the transcriptional response of key factors involved in adhesion and virulence. We also demonstrated that MRSA primed in SLM adhered better to human corneocytes and demonstrated adhesin-specific phenotypes that previously required genetic manipulation. These results support the potential utility of SLM as an in vitro model for assessing Staphylococcal physiology and metabolism on human skin.

Staphylococcus aureus is the major cause of skin diseases, and its increased prevalence in skin colonization and infections present a need to understand its physiology in this environment. The work presented here outlines S. aureus upregulation of colonization and virulence factors using a newly developed media that strives to replicate the human skin surface environment, and demonstrates roles for adhesins ClfA, SraP, and Fnbps in human corneocyte adherence.

The recurrence of biofilm-associated infections (BAIs) remains high after implant-associated surgery. Biofilms on the implant surface reportedly shelter bacteria from antibiotics and evade innate immune defenses. Moreover, little is currently known about eliminating residual bacteria that can induce biofilm reinfection. Herein, novel "interference-regulation strategy" based on bovine serum albumin-iridium oxide nanoparticles (BIONPs) as biofilm homeostasis interrupter and immunomodulator via singlet oxygen (¹ O₂ )-sensitized mild hyperthermia for combating BAIs is reported. The catalase-like BIONPs convert abundant H₂ O₂ inside the biofilm-microenvironment (BME) to sufficient oxygen gas (O₂ ), which can efficiently enhance the generation of ¹ O₂ under near-infrared irradiation. The ¹ O₂ -induced biofilm homeostasis disturbance (e.g., sigB, groEL, agr-A, icaD, eDNA) can disrupt the sophisticated defense system of biofilm, further enhancing the sensitivity of biofilms to mild hyperthermia. Moreover, the mild hyperthermia-induced bacterial membrane disintegration results in protein leakage and ¹ O₂ penetration to kill bacteria inside the biofilm. Subsequently, BIONPs-induced immunosuppressive microenvironment re-rousing successfully re-polarizes macrophages to pro-inflammatory M1 phenotype in vivo to devour residual biofilm and prevent biofilm reconstruction. Collectively, this ¹ O₂ -sensitized mild hyperthermia can yield great refractory BAIs treatment via biofilm homeostasis interference, mild-hyperthermia, and immunotherapy, providing a novel and effective anti-biofilm strategy.

Staphylococcus aureus bears a type 7b secretion system (T7bSS) that assembles in the bacterial envelope to promote the secretion of WXG-like proteins and toxic effectors bearing LXG domains. Cognate immunity proteins bind cytosolic effectors to mute their toxicity prior to secretion. T7b-secreted factors have been associated with the pathogenesis of staphylococcal disease and intraspecies competition. We identified earlier strain WU1, an S. aureus ST88 isolate that caused outbreaks of skin and soft tissue infections in mouse breeding facilities. WU1 was also found to persistently colonize the nasopharynx of animals, suggesting a strong host adaptation. In this manner, WU1 colonization and infectivity in mice resembles that of methicillin-sensitive and -resistant S. aureus strains in humans, where nasal carriage is a major risk factor for invasive infections. Here, animals were colonized with wild-type or T7-deficient WU1 strains or combinations thereof. Absence of the T7bSS did not affect colonization in the nasopharynx of animals, and although fluctuations were observed in weekly samplings, the wild-type strain did not replace the T7-deficient strain in cocolonization experiments. Bloodstream infection with a T7b-deficient strain resulted in enhanced survival and reduced bacterial loads and abscesses in soft tissues compared to infection with wild-type WU1. Together, experiments using a mouse-adapted strain suggest that the T7bSS of S. aureus is an important contributor to the pathogenesis of invasive disease.

The pathogenic bacterium Staphylococcus aureus is the most common pathogen isolated in skin-and-soft-tissue infections (SSTIs) in the United States. Most S. aureus SSTIs are caused by the epidemic clone USA300 in the USA. These infections can be serious; in 2019, SSTIs with S. aureus were associated with an all-cause, age-standardized mortality rate of 0.5 globally. Clinical presentations of S. aureus SSTIs vary from superficial infections with local symptoms to monomicrobial necrotizing fasciitis, which can cause systemic manifestations and may lead to serious complications or death. In order to cause skin infections, S. aureus employs a host of virulence factors including cytolytic proteins, superantigenic factors, cell wall-anchored proteins, and molecules used for immune evasion. The immune response to S. aureus SSTIs involves initial responders such as keratinocytes and neutrophils, which are supported by dendritic cells and T-lymphocytes later during infection. Treatment for S. aureus SSTIs is usually oral therapy, with parenteral therapy reserved for severe presentations; it ranges from cephalosporins and penicillin agents such as oxacillin, which is generally used for methicillin-sensitive S. aureus (MSSA), to vancomycin for methicillin-resistant S. aureus (MRSA). Treatment challenges include adverse effects, risk for Clostridioides difficile infection, and potential for antibiotic resistance.

Staphylococcus aureus is a clinically important bacterial pathogen that has become resistant to treatment with most routinely used antibiotics. Alternative strategies, such as vaccination and phage therapy, are therefore actively being investigated to prevent or combat staphylococcal infections. Vaccination requires that vaccine targets are expressed at sufficient quantities during infection so that they can be targeted by the host's immune system. While our knowledge of in vitro expression levels of putative vaccine candidates is comprehensive, crucial in vivo expression data are scarce and promising vaccine candidates during in vitro assessment often prove ineffective in preventing S. aureus infection. Here, we show how a newly developed high-throughput quantitative reverse transcription-PCR (qRT-PCR) assay monitoring the expression of 84 staphylococcal genes encoding mostly virulence factors can inform the selection and design of effective vaccine candidates against staphylococcal infections. We show that this assay can accurately quantify mRNA expression levels of these genes in several host organs relying only on very limited amounts of bacterial mRNA in each sample. We selected two highly expressed genes, lukE and lukD, encoding pore-forming leukotoxins, to inform the design of detoxified recombinant proteins and showed that immunization with recombinant genetically detoxified LukED antigens conferred protection against staphylococcal skin infection in mice. Consequently, knowledge of in vivo-expressed virulence determinants can be successfully deployed to identify and select promising candidates for optimized design of effective vaccine antigens against S. aureus. Notably, this approach should be broadly applicable to numerous other pathogens. IMPORTANCE Vaccination is an attractive strategy for preventing bacterial infections in an age of increased antimicrobial resistance. However, vaccine development frequently suffers significant setbacks when candidate antigens that show promising results in in vitro experimentation fail to protect from disease. An alluring strategy is to focus resources on developing bacterial virulence factors that are expressed during disease establishment or maintenance and are critical for bacterial in-host survival as vaccine targets. While expression profiles of many virulence factors have been characterized in detail in vitro, our knowledge of their in vivo expression profiles is still scarce. Here, using a high-throughput qRT-PCR approach, we identified two highly expressed leukotoxins in a murine infection model and showed that genetically detoxified derivatives of these elicited a protective immune response in a murine skin infection model. Therefore, in vivo gene expression can inform the selection of promising candidates for the design of effective vaccine antigens.

Staphylococcus aureus is a versatile pathogen known to cause hospital- and community-acquired, foodborne, and zoonotic infections. The clinical infections by S. aureus cause an increase in morbidity and mortality rates and treatment costs, aggravated by the emergence of drug-resistant strains. As a multi-faceted pathogen, it is imperative to consolidate the knowledge on its pathogenesis, including the mechanisms of virulence regulation, development of antimicrobial resistance, and biofilm formation, to make it amenable to different treatment strategies. Nanomaterials provide a suitable platform to address this challenge, with the potential to control intracellular parasitism and multidrug resistance where conventional therapies show limited efficacy. In a nutshell, the first part of this review focuses on the impact of S. aureus on human health and the role of virulence factors and biofilms during pathogenesis. The second part discusses the large diversity of nanoparticles and their applications in controlling S. aureus infections, including combination with antibiotics and phytochemicals and the incorporation of antimicrobial coatings for biomaterials. Finally, the limitations and prospects using nanomaterials are highlighted, aiming to foster the development of novel nanotechnology-driven therapies against multidrug-resistant S. aureus.

Staphylococci are commensals of human skin and mucous membranes, but some species can also cause serious infections. Host niches during both colonization and infection differ greatly and are characterized by specific environmental conditions (pH, temperature, oxygen, nutrient availability, and microbiota) that can affect gene expression and virulence of microbes. To successfully occupy extremely different habitats at different anatomical sites, Staphylococci are equipped with a variety of regulatory elements that allow specific adaptation to the changing environments. Not surprisingly, gene expression in vivo can be significantly different from the expression pattern observed in vitro. Niche specific stimuli that influence the bacterial ability to either cause infection or maintain colonization are only partially understood. Here, we describe habitat specific conditions and discuss the available literature analyzing staphylococcal gene expression, focusing on Staphylococcus aureus and S. epidermidis during colonization of the nose and skin.

Bacterial survival on, and interactions with, human skin may explain the epidemiological success of MRSA strains. We evaluated the bacterial counts for 27 epidemic and 31 sporadic MRSA strains on 3D epidermal models based on N/TERT cells (NEMs) after 1, 2 and 8 days. In addition, the expression of antimicrobial peptides (hBD-2, RNase 7), inflammatory cytokines (IL-1β, IL-6) and chemokine IL-8 by NEMs was assessed using immunoassays and the expression of 43 S. aureus virulence factors was determined by a multiplex competitive Luminex assay. To explore donor variation, bacterial counts for five epidemic and seven sporadic MRSA strains were determined on 3D primary keratinocyte models (LEMs) from three human donors. Bacterial survival was comparable on NEMs between the two groups, but on LEMs, sporadic strains showed significantly lower survival numbers compared to epidemic strains. Both groups triggered the expression of immune factors. Upon interaction with NEMs, only the epidemic MRSA strains expressed pore-forming toxins, including alpha-hemolysin (Hla), gamma-hemolysin (HlgB), Panton-Valentine leucocidin (LukS) and LukED. Together, these data indicate that the outcome of the interaction between MRSA and human skin mimics, depends on the unique combination of bacterial strain and host factors.

The study aimed to examine the influence of a rotating magnetic field (RMF) of two different frequencies (5 and 50 Hz) on the expression of regulatory (agrA, hld, rot) and staphylococcal enterotoxin (SE-sea, sec, sel) genes as well as the production of SEs (SEA, SEC, SEL) by the Staphylococcus aureus FRI913 strain cultured on a medium supplemented with a subinhibitory concentration of trans-anethole (TA). Furthermore, a theoretical model of interactions between the bacterial medium and bacterial cells exposed to RMF was proposed. Gene expression and SEs production were measured using quantitative real-time PCR and ELISA techniques, respectively. Based on the obtained results, it was found that there were no significant differences in the expression of regulatory and SE genes in bacteria simultaneously cultured on a medium supplemented with TA and exposed to RMF at the same time in comparison to the control (unexposed to TA and RMF). In contrast, when the bacteria were cultured on a medium supplemented with TA but were not exposed to RMF or when they were exposed to RMF of 50 Hz (but not to TA), a significant increase in agrA and sea transcripts as compared to the unexposed control was found. Moreover, the decreased level of sec transcripts in bacteria cultured without TA but exposed to RMF of 50 Hz was also revealed. In turn, a significant increase in SEA and decrease in SEC and SEL production was observed in bacteria cultured on a medium supplemented with TA and simultaneously exposed to RMFs. It can be concluded, that depending on SE and regulatory genes expression as well as production of SEs, the effect exerted by the RMF and TA may be positive (i.e., manifests as the increase in SEs and/or regulatory gene expression of SEs production) or negative (i.e., manifests as the reduction in both aforementioned features) or none.

Staphylococcus aureus is an opportunistic pathogen that can cause life-threatening bloodstream infections such as sepsis and endocarditis. In recent years, the emergence and increase of methicillin-resistant and multidrug-resistant S. aureus has posed a great challenge to the antibiotic treatment of infectious diseases. Anti-virulence strategies targeting virulence factors are an effective new therapy for the treatment of S. aureus infections.

In this study, we constructed a NWMN2330 deletion mutant (Newman-ΔNWMN2330) and a complement (Newman-ΔNWMN2330-C) of S. aureus Newman to study the role of NWMN2330 in the virulence of S. aureus. Through transcriptome sequencing, it was found that the expression of 224 genes in Newman-ΔNWMN2330 was significantly different (>2-fold) compared with S. aureus Newman, and these differentially expressed genes were related to multiple functions of S. aureus. And we found that NWMN2330 could positively regulate the expression of S. aureus hla gene. Therefore, the deletion mutant Newman-ΔNWMN2330 exhibited lower hemolytic activity and lower α-toxin production than Newman. Newman-ΔNWMN2330 also exhibited lower lethality and pathogenicity in worm survival experiments and nude mouse skin abscess model. RT-qPCR results showed that compared with the wild-type strain, the expression of saeRS and hla in Newman-ΔNWMN2330 strain was significantly reduced at the mRNA level, which preliminarily indicated that NWMN2330 promoted the expression of hla by up-regulating saeRS.

In general, our results indicated that NWMN2330 may be associated with the virulence of Staphylococcus aureus by increasing the expression of hla and saeRS.