Abstract
In Escherichia coli, the role of RND-type drug transporters other than the major efflux pump AcrB has largely remained undeciphered (particularly in multidrug resistant pathogens), because genetic engineering in such isolates is challenging. The present study aimed to explore the capability of the AcrB homolog MdtF to contribute to the extrusion of noxious compounds and to multidrug resistance in an E. coli clinical isolate with demonstrated expression of this efflux pump. An mdtF/acrB double-knockout was engineered, and susceptibility changes with drugs from various classes were determined in comparison to the parental strain and its acrB and tolC single-knockout mutants. The potential of MdtF to participate in the export of agents with different physicochemical properties was additionally assessed using accumulation and real-time efflux assays with several fluorescent dyes. The results show that there was limited impact to the multidrug resistant phenotype in the tested E. coli strain, while the RND-type transporter remarkably contributes to the efflux of all tested dyes. This should be considered when evaluating the efflux phenotype of clinical isolates via dye accumulation assays. Furthermore, the promiscuity of MdtF should be taken into account when developing new antibiotic agents.
Highlights
Knowledge about resistance mechanisms is crucial for future drug development in particular against Gram-negative pathogens
We aimed to explore the MdtF functionality and its contribution to drug resistance in an multidrug resistance (MDR) E. coli isolate with verified expression of this RND-type efflux pump [3,11]
In order to evaluate the contribution of MdtF to total TolC-dependent efflux in MDR E. coli strains, we aimed to compare an mdtF/acrB double-knockout of the patient isolate KUN9180 with the acrB (KUN∆acrB) and tolC single-knockout (KUN∆tolC) mutants from previous studies [3,11]
Summary
Knowledge about resistance mechanisms is crucial for future drug development in particular against Gram-negative pathogens. The importance of RND (resistance nodulation cell division)-type efflux pumps for the emergence of multidrug resistance (MDR) in Gram-negative pathogens has been demonstrated, among others, for Escherichia coli [2,3,4], Klebsiella [5], Enterobacter [6] Acinetobacter [7], and Pseudomonas aeruginosa strains [4]. As shown recently with the latter, in isolates lacking carbapenemases, the overexpression of efflux pumps appeared as the predominant reason for carbapenem and multidrug resistance [8]. In contrast to Pseudomonas aeruginosa strains (in which several different transporters of the RND superfamily contributes to MDR), AcrB has been the only multidrug efflux transporter with a proven impact in E. coli. One reason is that they are not found significantly expressed in laboratory strains [9] (in which most of the experiments were carried out), whereas results from MDR clinical isolates have remained underrepresented
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