CpxR overexpression increases the susceptibility of acrB and cpxR double-deleted Salmonella enterica serovar Typhimurium to colistin

Abstract

Background: Colistin has been used as the last therapeutic resort for treatment of MDR Gram-negative bacteria infections in humans. The two-component system CpxAR has been reported to contribute to the MDR of bacteria. There may be a more complex network mediated by CpxAR contributing to colistin susceptibility than previously understood. Methods: A series of AcrB or CpxR deletion mutants of a multidrug-susceptible standard strain of Salmonella enterica serovar Typhimurium (Salmonella Typhimurium) was constructed in our previous study. MICs of colistin were determined by the 2-fold serial broth microdilution method. Time–kill and survival assays were carried out with various concentrations of colistin. Growth curves and starvation survival were measured by OD600 or cfu count in LB and M9-glucose (0.2%) minimum media. Quantitative RT–PCR was used to determine the mRNA expression levels of target genes. Results: The results showed that the MIC of colistin for the CpxR-overexpressed strain JSDacrBDcpxR::kan/pcpxR was dramatically decreased (0.05 mg/L) by 16-fold compared with JS (0.8 mg/L) and JSDacrBDcpxR::kan (0.8 mg/L). Colistin time–kill and survival assays showed that JSDacrBDcpxR::kan/pcpxR was more susceptible to colistin (0.05 mg/L), but had a considerably higher survivability regarding prolonged starvation stress compared with JSDacrBDcpxR::kan. Furthermore, the expression levels of colistin resistance-related genes (phoP, phoQ, pmrB, pmrC, pmrH and pmrD) were found to be remarkably down-regulated and the negative regulatory protein mgrB was significantly up-regulated. Conclusions: This study demonstrated that CpxR may regulate the colistin susceptibility of Salmonella Typhimurium through the PmrAB and PhoPQ regulatory systems.