Inhibitory effects of Quercetagetin on the growth, biofilm formation, and virulence factors of bovine mastitis-associated Escherichia coli

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Introduction Bovine mastitis caused by Escherichia coli ( E. coli ) poses a significant challenge to the dairy industry, primarily due to biofilm-mediated antibiotic tolerance. Methods This study investigated the antibacterial and anti-biofilm activities of Quercetagetin (QG), a natural flavonoid, against highly virulent and multidrug-resistant (MDR) E. coli isolates, rigorously screened and selected from a large clinical cohort of 136 bovine mastitis cases. Results QG exhibited potent antibacterial activity with Minimum Inhibitory Concentrations (MICs) ranging from 0.5 to 1 mg/mL. Notably, QG not only prevented initial biofilm formation (Minimum Biofilm Inhibitory Concentration; MBIC = 0.5 mg/mL) but also effectively eradicated mature biofilms at a low concentration (Minimum Biofilm Eradication Concentration; MBEC = 1 mg/mL). Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) confirmed that QG disrupted biofilm architecture, stripped away the extracellular polymeric substance (EPS), and induced bacterial cell lysis. Phenotypic assays revealed that QG significantly ( P < 0.0001) suppressed swimming, swarming, and twitching motilities and inhibited the synthesis of cellulose and poly-N-acetylglucosamine (PGA). Mechanistically, qRT-PCR analysis demonstrated that QG downregulated the expression of flagellar genes ( fliA , fliG ) and the master biofilm regulator csgD . Discussion Given that the transcription of these genes is positively regulated by the second messenger cyclic dimeric guanosine monophosphate (c-di-GMP), these transcriptional findings support a hypothesis consistent with c-di-GMP pathway involvement, suggesting that QG may impair biofilm integrity by modulating components of this downstream regulatory network. Collectively, these results suggest QG as a promising non-antibiotic candidate compound for controlling E. coli mastitis.