Construction and antibiotic resistance of a Salmonella typhimurium strain overexpressing baeR

Abstract:

[Background] Salmonella typhimurium is a major zoonotic pathogen, and its multi-drug resistance is becoming increasingly serious. The two-component system can regulate the resistance of S. typhimurium. [Objective] We constructed the baeR-overexpressing strain and complementary strain of S. typhimurium to study the effect of BaeSR on the antibiotic resistance of S. typhimurium. [Methods] a complementary strain (CRcbaeR∆baeSR∆acrB) and an overexpression strain (CRpbaeR∆baeSR∆acrB) were constructed based on a baeSR and acrB double-deletion strain of S. typhimurium (CR∆baeSR∆acrB), and their antimicrobial susceptibility and biological characteristics were tested. RNA-Seq was used to screen the differentially expressed genes (DEGs) related to drug resistance, and RT-qPCR was conducted to verify some drug-related genes. [Results] The baeR-overexpressing strain and complementary strain were successfully constructed. Compared with that in CR∆baeSR∆acrB, the minimum inhibitory concentrations (MICs) of ofloxacin, enrofloxacin, florfenicol, mequindox, ceftazidime, ceftiofur, amoxicillin, and ampicillin increased by 2-256 folds while that of spectinomycin and apramycin decreased by 50% in CRpbaeR∆baeSR∆acrB. The MIC of ceftazidime, ceftiofur, florfenicol, and enrofloxacin was two-fold higher in CRcbaeR∆baeSR∆acrB than in CR∆baeSR∆acrB. The growth curves of CR∆baeSR∆acrB, CRpbaeR∆baeSR∆acrB, and CRcbaeR∆baeSR∆acrB showed no significant difference. However, the biofilm-forming ability and motility of CRpbaeR∆baeSR∆acrB and CRcbaeR∆baeSR∆acrB were significantly higher than those of CR∆baeSR∆acrB (P<0.01). A total of 743 DEGs were identified between CRpbaeR∆baeSR∆acrB and CR∆baeSR∆acrB, including 724 upregulated genes and 19 downregulated genes. Bioinformatics analysis showed that the DEGs were mainly enriched in pathways such as β-lactam resistance, quorum sensing, and two-component system. A total of 3 073 DEGs were identified between CRcbaeR∆baeSR∆acrB and CR∆baeSR∆acrB, including 1 467 upregulated genes and 1 606 downregulated genes. These DEGs were mainly enriched in pathways such as carbon metabolism, biosynthesis of amino acids, and biosynthesis of antibiotics. A total of 15 drug resistance-related DEGs were screened out, mainly involving efflux pump, biofilm, and two-component system. Five out of the 15 DEGs were randomly selected for RT-qPCR, which showed consistent results and confirmed the reliability of RNA-Seq results. [Conclusion] This study indicates that baeR affects the drug resistance of S. typhimurium by regulating its biofilm-forming ability and motility. RNA-Seq screened out 15 resistance-related DEGs, indicating that baeR overexpression can affect the drug resistance of S. typhimurium by regulating the expression of the genes associated with efflux pump, biofilm and flagella.