Deciphering the roles of AcrAB-TolC efflux pump in promoting the transmission of antibiotic resistance

The dissemination of antimicrobial resistance has emerged as a formidable challenge to global public health. Plasmid-mediated conjugative transfer is the primary mechanism driving the rapid spread of resistance. Bacterial multidrug efflux pumps not only directly confer antibiotic resistance phenotypes via reducing intracellular drug concentrations, but also are involved in the modulation of microbial pathogenesis. However, it remains unclear whether efflux pumps participate in plasmid transfer and the spread of drug resistance, especially in the absence of antibiotic pressure. In this study, we elucidate the pivotal roles of efflux pumps, such as AcrAB-TolC, in the horizontal transfer of resistance plasmids. Notably, deletion of acrA, acrB, and tolC significantly diminished the conjugative transfer of RP4-7 and clinically relevant resistance plasmids. Upon complementation, the conjugative frequency of these resistance plasmids was restored to levels comparable to those of the control group. Mechanistic investigations revealed that AcrAB-TolC functions to maintain bacterial cell physiological state and membrane homeostasis, beyond its role as an efflux pump. Specifically, the absence of the acrB gene resulted in reduced cell-to-cell interactions, increased DNA damage, diminished energy metabolism, and impaired activity of the quorum sensing system. Using AcrB protein as a receptor, we identify chlorpromazine, a phenothiazine anti-psychotic drug, as a potent horizontal gene transfer inhibitor, which effectively prevents the spread of ARGs both in vitro and in vivo. Collectively, our findings underscore the role of efflux pumps in plasmid-mediated conjugative transfer of ARGs, offering novel insights into the mechanisms by which efflux pumps drive the spread of antibiotic resistance.