Methicillin-susceptible Staphylococcus aureus (MSSA) accounts for ∼40% of staphylococcal infections in China. However, the molecular characterization of MSSA is not well described. In this study, 124 MSSA strains collected in 2013 from a comprehensive teaching hospital in Chongqing, Southwestern China, were subjected to antibiotics susceptibility testing and molecular typing, including multilocus sequence typing, staphylococcal protein A ( spa ) gene typing, accessory gene regulator ( agr ) typing, pulsed-field gel electrophoresis (PFGE) typing, Panton–Valentine leukocidin ( pvl ) gene detection, and antibiotic-resistant gene detection. MSSA strains exhibited high genetic heterogeneity. A total of 10 PFGE groups, 26 sequence types, and 47 spa types were identified. Type I (62.9%) was the most frequent agr type, followed by type II (15.3%), type IV (11.3%), and type III (10.5%). The prevalence of pvl genes was 27.4% (34/124). Notably, 44.4% (55/124) of MSSA strains were multidrug resistance (MDR), and MDR isolates were mostly resistant to penicillin, erythromycin, and clindamycin. The resistance gene blaZ was present in 84.7% of strains, ermC was present in 85.5% of strains, ermA was present in 28.2% of strains, tetK was present in 16.1% of strains, tetM was present in 6.5% of strains, and aacA-aphD was present in 2.6% of strains. These data demonstrated the high prevalence of MDR MSSA in Chongqing, thereby indicating the need to control MSSA infection.

Staphylococcus aureus is a vital pathogen causing clinical infections. Capsules are important virulence factors for S. aureus. This study investigates the regulatory mechanisms underlying capsule production in S. aureus. Bacterial strains XN108 and Newman were used, and combined approaches like RNA sequencing (RNA-seq), RT-qPCR, transmission electron microscopy (TEM), gene reporter, and electrophoretic mobility shift assay (EMSA) were performed to test the role and mechanism of WalK(S221P) mutation in S. aureus capsule production. RNA-seq showed an increased expression of cap genes in the WalK(S221P)-carried S. aureus XN108 relative to the mutation-cured XN108-R. TEM and capsular polysaccharide determination demonstrated that XN108 produced more capsules than XN108-R did. Similar results were presented in the WalK(S221P)-contained K-Newman versus the wild-type Newman. RT-qPCR screening showed an increasing expression of the mgrA gene in XN108 versus XN108-R. Gene reporter and EMSA analysis revealed that WalK(S221P) mutation promoted S. aureus capsule production through MgrA. Deletion of mgrA decreased the WalK(S221P)-mediated capsule yield. Moreover, WalK(S221P) mutation remarkably increased the tolerance of S. aureus to whole blood killing and microphage phagocytosis. Overall, these data provide mechanistic insights into the effect of WalK(S221P) on the capsule production of S. aureus, which may set down foundations for future S. aureus virulence investigations.

Staphylococcus aureus is a major opportunistic pathogen that causes a wide spectrum of human and animal diseases. Capsule is one of the key virulence factors of S. aureus. Approximately 90% of S. aureus isolates produce capsular polysaccharides (CPs) that envelop the entire bacterial cells. CPs can suppress the phagocytosis of S. aureus by innate immune cells, promote intracellular survival, and reduce the killing efficacy of antimicrobial agents. As a result, CPs are versatile candidates for vaccine development. The synthesis of S. aureus CPs is controlled by the cap operon, which contains 16 genes encoding functional enzymes involved in the biosynthesis and transport of CPs. During S. aureus growth, cap operon expression is strictly regulated. The control region of the cap operon is characterized by two promoters: a housekeeping sigma factor A (SigA)-dependent promoter and a stress-responsive sigma factor B-dependent promoter. Many transcription factors, including both positive and negative regulatory molecules, are timely involved in the regulatory network of S. aureus capsule synthesis. Moreover, environmental conditions such as medium composition and carbon dioxide levels further modulate CP production. In this review, the structure and function of S. aureus CPs are introduced, and the biosynthesis of S. aureus CPs is discussed. Moreover, the regulation of S. aureus CP production is summarized, the factors affecting CP production are outlined, and the preparation of S. aureus CPs is reviewed. The advanced information presented here may provide useful references for further research on the production and function of S. aureus CPs.

The emergence of vancomycin-intermediate Staphylococcus aureus (VISA) threatens the efficacy of this last-line antibiotic. The GraSR two-component system is frequently mutated in VISA strains. Here, we demonstrate that the GraS(T136I) point mutation, identified in the clinical VISA isolate XN108, is a key determinant of reduced vancomycin susceptibility. Introducing this mutation into the susceptible strain Newman increased the vancomycin MIC from 1.5 to 4 mg/L, while its reversion in XN108 decreased the MIC from 12 to 8 mg/L. The mutation conferred common phenotypes, including thickened cell wall, decreased autolysis, and reduced cell surface negative charge via upregulation of the dltABCD operon and mprF. Notably, the GraS(T136I) mutation also upregulated virulence genes (efb, hlb, sbi, hld) and enhanced hemolytic activity. Interestingly, despite this hypervirulent profile, the mutant showed impaired long-term survival within macrophages. Our study reveals that a single GraSR mutation can co-regulate vancomycin resistance and virulence, offering new insights into the adaptation of S. aureus to antibiotic pressure.

The emergence of vancomycin-intermediate Staphylococcus aureus (VISA) threatens the efficacy of this last-line antibiotic. The GraSR two component system is frequently mutated in VISA strains. Here, we demonstrate that the GraS(T136I) point mutation, identified in the clinical VISA isolate XN108, is a key determinant of reduced vancomycin susceptibility. Introducing this mutation into the susceptible strain Newman increased the vancomycin MIC from 1.5 to 4 mg/L, while its reversion in XN108 decreased the MIC from 12 to 8 mg/L. The mutation conferred common phenotypes, including thickened cell wall, decreased autolysis, and reduced cell surface negative charge via upregulation of the dltABCD operon and mprF. Notably, GraS(T136I) mutation also upregulated virulence genes (efb, hlb, sbi, hld) and enhanced hemolytic activity. Interestingly, despite this hypervirulence profile, the mutant showed impaired long term survival within macrophages. Our study reveals that a single GraSR mutation can co-regulate vancomycin resistance and virulence, offering new insights into the adaptation of S. aureus to antibiotic pressure.

Antibiotic-resistant Staphylococcus aureus infections pose a formidable clinical challenge. Cold atmospheric plasma (CAP) has emerged as a promising disinfection technology; however, the underlying bacterial response mechanisms and potential resistance to CAP remain incompletely elucidated. Here, we validated the broad-spectrum and efficient bactericidal activity of CAP against both drug-sensitive and drug-resistant clinical isolates of S. aureus. Mechanistically, CAP primarily eliminated bacteria by inducing oxidative damage to proteins and lipids, thereby disrupting internal cellular structures. The addition of exogenous free radical scavengers significantly attenuated CAP-mediated oxidative killing. Transcriptomic analysis and RT-qPCR validation further revealed upregulation of redox-related genes in S. aureus following CAP exposure. Deletion of key antioxidant genes, particularly ahpC and mrgA, markedly enhanced bacterial susceptibility to CAP. Virtual screening and surface plasmon resonance analysis identified two small-molecular inhibitors, C4515 and A8226, which specifically target AhpC and MrgA, respectively, and potentiated CAP bactericidal efficacy. Collectively, this study delineates the molecular interactions between CAP and S. aureus, and underscores the potential of CAP-based strategies for combating drug-resistant bacterial infections.