Development of CmeABC Efflux Pump- Based Intervention Strategies Against Campylobacter

CmeABC, a multidrug efflux pump, contributes to Campylobacter resistance to a broad spectrum of antimicrobial agents and is also essential for Campylobacter colonization by mediation of bile resistance. We hypothesize that inhibition of CmeABC will not only control antibiotic resistance but also increase the susceptibility of Campylobacter to in vivo bile salts, consequently decreasing the colonization level of Campylobacter. Using both in vitro and in vivo systems, we examined the effect of an efflux pump inhibitor (EPI) MC-207,110 on the susceptibility of Campylobacter to various antimicrobials. Presence of the EPI resulted in 2- to 2048-fold reduction in the MICs of antimicrobials known to be substrates of CmeABC pump in all Campylobacter strains. Particularly, the MICs of selected bile salts were dramatically decreased 64- to 512-fold when the EPI was used. The intrinsic and acquired resistance of C. jejuni to macrolide was decreased significantly (32- to 64-fold reduction in the MIC of erythromycin) in the presence of the EPI while the MICs of fluoroquinolones were only slightly decreased (2-4 folds). Investigation of 57 clinical Campylobacter isolates of various origins further showed that the EPI decreased the MICs of erythromycin (2- to 512-fold) in all isolates. Compared to wild-type strains, the isogenic CmeB mutants displayed much lower magnitude of reduction in the MICs of antimicrobials in the presence of the EPI. The inhibitory effect of the EPI was does-dependents and as low as 0.5 μg/ml of the EPI resulted in decreased MIC of antimicrobials in C. jejuni. Presence of the EPI decreased the frequency of emergence of erythromycin-resistant mutants in C. jejuni (<10^-11), which is well below normal frequency of approximate 10^-8. Notably, MIC of erythromycin was also greatly decreased (> 4-fold) in CmeB mutants in the presence of EPI, suggesting the existence of other pump(s) involved in macrolide resistance in C. jejuni. Chicken colonization study demonstrated that oral administration of EPI dramatically reduced the colonization of Campylobacter in the intestine. In addition, anti-CmeC antibodies also enhanced the susceptibility of C. jejuni to bile salt, suggesting immune intervention by targeting CmeC may be another effective strategy to inhibit CmeABC efflux pump. Together, these findings indicate that inhibition of CmeABC by specific EPI or antibodies is a promising approach to control antibiotic resistance and colonization of Campylobacter in human and animals.