Limited Multidrug Resistance Efflux Pump Overexpression among Multidrug-Resistant Escherichia coli Strains of ST131.

Gram-negative bacteria partly rely on efflux pumps to facilitate growth under stressful conditions and to increase resistance to a wide variety of commonly used drugs. In recent years, Escherichia coli sequence type 131 (ST131) has emerged as a major cause of extraintestinal infection frequently exhibiting a multidrug resistance (MDR) phenotype. The contribution of efflux to MDR in emerging E. coli MDR clones, however, is not well studied. We characterized strains from an international collection of clinical MDR E. coli isolates by MIC testing with and without the addition of the AcrAB-TolC efflux inhibitor 1-(1-naphthylmethyl)-piperazine (NMP). MIC data for 6 antimicrobial agents and their reversion by NMP were analyzed by principal-component analysis (PCA). PCA revealed a group of 17 MDR E. coli isolates (n = 34) exhibiting increased susceptibility to treatment with NMP, suggesting an enhanced contribution of efflux pumps to antimicrobial resistance in these strains (termed enhanced efflux phenotype [EEP] strains). Only 1/17 EEP strains versus 12/17 non-EEP MDR strains belonged to the ST131 clonal group. Whole-genome sequencing revealed marked differences in efflux-related genes between EEP and control strains, with the majority of notable amino acid substitutions occurring in AcrR, MarR, and SoxR. Quantitative reverse transcription-PCR (qRT-PCR) of multiple efflux-related genes showed significant overexpression of the AcrAB-TolC system in EEP strains, whereas in the remaining strains, we found enhanced expression of alternative efflux proteins. We conclude that a proportion of MDR E. coli strains exhibit an EEP, which is linked to an overexpression of the AcrAB-TolC efflux pump and a distinct array of genomic variations. Members of ST131, although highly successful, are less likely to exhibit the EEP.

Contribution of AcrAB-TolC to multidrug resistance in an Escherichia coli sequence type 131 isolate.

Drug efflux by resistance-nodulation-cell division (RND)-type transporters, such as AcrAB-TolC of Escherichia coli, is an important resistance mechanism in Gram-negative bacteria; however, its contribution to multidrug resistance (MDR) in clinical isolates is poorly defined. We inactivated acrB of a sequence type 131 E. coli human isolate that showed high-level MDR, but had no mutations within the known efflux-associated local or global regulators. The resistance profile of the acrB deletion mutant revealed significantly increased susceptibility to drugs from seven antibiotic classes, including agents usually inactive against Gram-negative bacteria, notably the new oxazolidinone, tedizolid (512-fold enhanced susceptibility). AcrB deficiency reduced, but did not abolish, the efflux of dyes, which indicates the activity of at least one more efflux transporter. The findings demonstrate the efficacy of AcrAB-TolC-mediated broad-spectrum drug efflux, including agents primarily developed for Gram-positive pathogens, in a clinical isolate representative of a globally-emerging lineage. The results illustrate the need to develop molecules modified to impede their transport by AcrAB-TolC and its homologues and new efflux inhibitors.