Integrative virtual and empirical screening to identify multi-target flavonoid quercetin-3-O-glucuronide for reducing MRSA pathogenicity.

OBJECTIVES: Methicillin-resistant Staphylococcus aureus (MRSA) poses a major clinical challenge due to escalating antibiotic resistance. This study aimed to evaluate the antivirulence activity of quercetin-3-O-glucuronide (Q3G) against MRSA and to elucidate its molecular targets and therapeutic efficacy in vitro and in vivo.

METHODS: Enzymatic assays were conducted to determine the inhibitory effects of Q3G on sortase A (SrtA) and caseinolytic peptidase P (ClpP) in MRSA USA300. Virulence-associated phenotypes and gene expression were assessed in vitro. Target engagement was analyzed using structural approaches. Therapeutic efficacy was evaluated in Galleria mellonella and murine pneumonia and skin infection models.

RESULTS: Q3G inhibited SrtA and ClpP with IC₅₀ values of 10.64 and 7.11 µg/mL, respectively, without affecting bacterial growth. It markedly reduced adhesion, invasion, biofilm formation, toxin secretion including Hla and PVL, hemolytic activity, urease function, and virulence gene expression. Structural analysis confirmed direct binding to key residues of both targets. In vivo, 40 mg/kg Q3G significantly improved survival in wax borers and pneumonia mice, reduced skin lesion severity, and decreased bacterial burden by approximately 30%.

CONCLUSIONS: Q3G acts as a dual-target antivirulence agent that attenuates MRSA pathogenicity without bactericidal pressure, offering a growth-independent strategy with reduced potential for resistance development.