BaeR overexpression enhances the susceptibility of acrB deleted Salmonella enterica serovar Typhimurium to polymyxin

Abstract

The mechanism of polymyxin resistance is complex, and the modification of lipopolysaccharide mediated by two-component system is one of the main cause of polymyxin resistance. To date, no studies have reported the contribution of the BaeSR two-component system to the polymyxin resistance of Salmonella. In this study, baeR, acrB single and double gene deletion strains of Salmonella typhimurium (AT-P128) which induced polymyxin resistance in vitro were constructed by using CRISPR/Cas9 gene editing technology, and the baeR gene was overexpressed in the acrB single gene deletion strain by the pUC19 plasmid. The susceptibility of different strains to polymyxin was determined by broth dilution method. Time–kill assay was carried out with different concentrations of polymyxin. The difference of gene expression among strains was compared by transcriptome sequencing (RNA-seq) and RT-qPCR. As a result, the MIC of the BaeR overexpression strain (AT-P128ΔacrB/pbaeR) to polymyxin was significantly reduced by 8-fold compared with the other tested strains. The growth curve results showed no significant change in the growth rate of the strain before and after gene deletion and overexpression. The time-kill assay showed that AT-P128ΔacrB/pbaeR was more susceptible under different concentrations of polymyxin. RNA-seq and RT-qPCR results showed that the expression levels of several polymyxin resistance-related genes including phoPQ, pmrD, pmrAB, arnT, eptB, lpxD, pagC and pagL changed significantly. These results indicate that overexpression of baeR in the context of the acrB gene deletion increases the polymyxin susceptibility of the strain and affects the expression level of polymyxin resistance genes, providing insight into the polymyxin resistance mechanism of S. typhimurium.