Antibacterial and anti-biofilm activity of kushenol N against methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) poses a serious clinical challenge due to antibiotic resistance and biofilm formation. Here, we demonstrate that kushenol N (KN), a natural flavonoid, exhibits potent antibacterial activity against Gram-positive bacteria, including multiple MRSA isolates (MICs of 8-32 μg/mL), and anti-biofilm activity against MRSA USA300. KN disrupted bacterial membranes and inhibited biofilm formation in a dose-dependent manner, reducing biomass by ∼85 % at 1/2 MIC and lowering extracellular DNA to ∼18 % of control. Key biofilm-associated genes (sarA, atl, agrA, clfA) were downregulated by 0.39-0.69-fold in MRSA USA300. KN displayed low cytotoxicity and minimal hemolytic activity (<6 %). In vivo, KN improved survival in Galleria mellonella (up to 100 % at 12 h) and murine infection models (up to 83 % at 75 mg/kg), while reducing bacterial loads and tissue damage, approaching the efficacy of vancomycin. These results highlight KN as a promising dual-function antibacterial and anti-biofilm agent, while the antivirulence effects were specifically observed in MRSA USA300, supporting further preclinical development and evaluation across additional MRSA strains.