鼠伤寒沙门氏菌baeR过表达株的构建及其耐药性

背景]鼠伤寒沙门氏菌（Salmonella typhimurium）是一种重要的人兽共患病原菌,其多重耐药性问题日益严重,双组分系统可调控鼠伤寒沙门氏菌的耐药性。目的]通过构建鼠伤寒沙门氏菌baeR过表达株及回补株探究BaeSR双组分系统对鼠伤寒沙门氏菌耐药性的影响。方法]在BaeSR双组分系统和AcrB外排泵双缺失株（CRΔbaeSRΔacrB）的基础上构建baeR过表达株（CRpbaeRΔbaeSRΔacrB）及baeR回补株（CRcbaeRΔbaeSRΔacrB）,测定双缺失株、回补株和过表达株的最小抑菌浓度（minimum inhibitory concentration,MIC）,并对其生长特性、生物膜形成能力及运动性进行分析。采用转录组学技术筛选与耐药相关的差异表达基因,RT-qPCR验证耐药相关基因。结果]构建了鼠伤寒沙门氏菌baeR过表达株和baeR回补株。与双缺失株相比,过表达株对氧氟沙星、恩诺沙星、氟苯尼考、乙酰甲喹、头孢他啶、头孢噻呋、阿莫西林和氨苄西林的MIC分别升高2-256倍,对大观霉素、安普霉素的MIC下降了50%;与双缺失株相比,回补株对头孢他啶...

Construction and antibiotic resistance of a Salmonella typhimurium strain overexpressing baeR

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 acr B double-deletion strain of S.typhimurium(CR?baeSR?acr B),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?acr B,CRpbaeR?baeSR?acr B,and CRcbaeR?baeSR?acr B showed no significant difference.However,the biofilm-forming ability and motility of CRpbaeR?baeSR?acr B and CRcbaeR?baeSR?acr B were significantly higher than those of CR?baeSR?acrB(P<0.01).A total of 743 DEGs were identified between CRpbaeR?baeSR?acr B and CR?baeSR?acr B,including 724 upregulated genes and 19 downregulated genes.Bioinformatics analysis showed that the DEGs were mainly enriched in pathways such asβ-lactam resistance,quorum sensing,and two-component system.A total of 3073 DEGs were identified between CRcbaeR?baeSR?acrB and CR?baeSR?acrB,including 1467 upregulated genes and 1606 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.