不同条件下伤寒沙门菌Vi荚膜多糖的表达调控研究

Regulation of Vi capsular polysaccharide expression in Salmonella enterica serovar Typhi under different conditions

为探究不同培养条件对伤寒沙门菌Vi荚膜多糖表达的影响及Vi荚膜多糖对细菌在巨噬细胞内生存能力的影响,本研究将伤寒沙门菌在LB培养条件下给予不同应激或在不同培养基中培养,通过实时荧光定量反转录聚合酶链反应(real time fluorescent quantitative reverse transcription polymerase chain reaction,real time RT-PCR)检测Vi荚膜多糖相关基因的转录水平,并采用流式细胞术检测Vi荚膜多糖的表达水平。将野生(wild type,WT)株和Vi荚膜多糖缺陷(ΔvexE)株分别感染巨噬细胞,观察其胞内生存能力。结果显示,加入氧应激、Mg^(2+)应激、抗菌肽应激后,tviA、tviB、vexE基因表达下调,加入酸应激和在SOB、LPM、M9培养基中培养后,tviA、tviB、vexE基因表达上调;除氧应激外,其余应激组流式细胞术结果与real time RTPCR结果一致。与WT株相比,ΔvexE株在巨噬细胞内生存能力明显下降。结果表明,酸应激及SOB、LPM、M9培养基促进了Vi荚膜多糖表达,Mg^(2+)应激、抗菌肽应激则抑制了Vi荚膜多糖表达,氧应激对Vi荚膜多糖表达的影响有待进一步研究;Vi荚膜多糖有利于伤寒沙门菌在巨噬细胞内生存。

The aim of this research was to study the influence of different culture conditions on the expression of Vi capsular polysaccharide and the effect of Vi capsular polysaccharide in the viability of Salmonella enterica serovar Typhi(S.Typhi)in macrophages.S.Typhi was cultured in LB medium under different stresses or in different media,and real time fluorescent quantitative reverse transcription polymerase chain reaction(real time RT-PCR)was used to detect the transcription of Vi-related genes and flow cytometry was used to analyze the expression of Vi.Macrophages were infected with wild type(WT)strain and Vi mutant strain(ΔvexE)to observe the effect of Vi in the viability of bacteria.The results showed that the expressions of tviA,tviB and vexE were down-regulated in responses to oxygen,Mg^(2+)and antimicrobial peptide stresses,and up-regulated under acid stress,and in SOB,LPM,M9 media.Except for oxygen stress,the results of flow cytometry were consistent with real time RT-PCR.The survival rate ofΔvexE strain in macrophages was significantly lower than that of WT strain.It is concluded that acid stress,SOB,LPM and M9 media promote Vi expression.Mg^(2+)and antimicrobial peptide stresses inhibit Vi expression.The influence of oxygen stress on Vi expression is still unclear.Vi could enhance the intracellular survival of S.Typhi in macrophages.