Synergistic effect of the rifaximin-berberine combination against Klebsiella pneumoniae: RfaH targeting supported by...

The escalating crisis of antimicrobial resistance (AMR) among Gram-negative pathogens, particularly Klebsiella pneumoniae (KP), necessitates innovative strategies to enhance the efficacy of existing antibiotics. Synergistic drug combinations present a promising approach to improve therapeutic outcomes and delay the emergence of resistance. This study investigates the synergistic interaction between the natural alkaloid berberine chloride and the repurposed antibiotic rifaximin against KP. Integrated in vitro and in silico analyses reveal significant bactericidal synergy between the two agents, mediated through concurrent inhibition of the transcriptional anti-termination factor RfaH, a key regulator of virulence and capsule biosynthesis. Molecular docking and dynamics simulations demonstrate that both compounds cooperatively bind to the RfaH pocket, stabilizing an inactive ternary complex without major structural disruption. Functional assays confirm that the combination effectively suppresses RfaH-dependent capsule production at lower concentrations compared to monotherapy. These findings suggest that RfaH is a viable target for combinatorial inhibition and provide a plausible mechanistic foundation for the berberine-rifaximin synergy. This work supports the rational development of dual-targeting anti-virulence strategies to combat multidrug-resistant KP infections.