Functional Study of : a Lineage-Specific Two-Component System Positively Regulating Biofilm Formation.

PURPOSE

Biofilms significantly contribute to the persistence and antibiotic resistance of infections. However, the regulatory mechanisms governing biofilm formation of remain not fully elucidated. This study aimed to investigate the function of the lineage-specific two-component system, , in biofilm regulation and pathogenicity.

METHODS

Bioinformatic analysis was conducted to assess the prevalence of across various lineages and to examine its structural features. The impact of on pathogenicity was evaluated using in vivo mouse models, including skin abscess, bloodstream infection, and nasal colonization models. Crystal violet staining and confocal laser scanning microscopy were utilized to examine the impact of on biofilm formation. Mechanistic insights into -mediated biofilm regulation were investigated by quantifying polysaccharide intercellular adhesin (PIA) production, extracellular DNA (eDNA) release, autolysis assays, and RT-qPCR.

RESULTS

The prevalence of varied among different lineages, with notably low carriage rates in ST398 and ST59 lineages. Deletion of in NCTC8325 strain resulted in decreased susceptibility to β-lactam and glycopeptide antibiotics. Although did not significantly affect acute pathogenicity, the Δ mutant exhibited significantly reduced nasal colonization and biofilm-forming ability. Overexpression of in naturally -lacking strains (ST398 and ST59) enhanced biofilm formation, suggesting a lineage-independent effect. Phenotypic assays further revealed that the Δ mutant showed reduced PIA production, decreased eDNA release, and lower autolysis rates. RT-qPCR indicated significant downregulation of , and genes, along with upregulation of , whereas autolysis-related genes remained unchanged.

CONCLUSION

The two-component system positively regulates biofilm formation in a lineage-independent manner, primarily by modulating PIA synthesis via the operon. These findings provide new insights into the molecular mechanisms of biofilm formation in and highlight as a potential target for therapeutic strategies aimed at combating biofilm-associated infections.