副溶血性弧菌luxS缺失株的构建及其生物学特性

Construction and Biological Characteristics of a luxS-Deleted Mutant of Vibrio parahaemolyticus

本实验通过同源重组技术构建副溶血性弧菌(Vibrio parahaemolyticus)luxS突变株,比较野生株和突变株的生长能力、4,5-二羟基-2,3-戊二酮(4,5-dihydroxy-2,3-pentanedione,DPD)含量、生物被膜形成能力、运动能力、药物敏感性、胞外蛋白分泌能力以及群体感应相关基因的表达量,研究LuxS/AI-2群体感应系统LuxS基因缺失后对副溶血性弧菌生物学特性的影响。结果显示,敲除luxS基因并不影响菌株的药物敏感性和生长能力;与野生株相比,突变株的DPD含量、泳动能力显著降低,而生物被膜形成能力、培养24 h的胞外蛋白酶分泌能力显著增强;实时聚合酶链式反应结果显示突变株的鞭毛合成基因flgM的表达量显著上升,而溶血素分泌相关基因toxS、鞭毛合成基因flaA、外膜蛋白基因ompW的表达量均显著下降。本实验可为进一步研究副溶血性弧菌LuxS/AI-2群体感应系统对其生物学特性的调控机制提供参考,有助于预防和控制副溶血性弧菌。

In this study,a luxS-deleted mutant of Vibrio parahaemolyticus was successfully constructed by homologous recombination technology,and the growth capacity,4,5-dihydroxy-2,3-pentanedione(DPD)production,biofilm formation,motility,drug susceptibility,extracellular protein-secreting capacity,and quorum sensing-related gene expression of the wild and mutant strains were compared to understand the effects of luxS gene deletion in the LuxS/AI-2 quorum sensing system on the biological characteristics of V.parahaemolyticus.The results showed that the deletion of the luxS gene did not affect the drug susceptibility or growth capacity of the strain.Compared with the wild-type strain,the DPD production and mobility of the mutant strain was significantly reduced,while the capacity of biofilm formation and extracellular protease secretion after 24 hours of incubation was significantly enhanced.The real-time polymerase chain reaction(PCR)results showed that the expression of the flagellar synthesis gene flgM significantly increased,while the expression of the hemolysin secretion-related gene toxS,the flagella synthesis gene flaA and the outer membrane protein gene ompW decreased significantly in the mutant strain.This study provides a basis for further research on the regulation mechanism of the LuxS/AI-2 quorum sensing system on the biological characteristics of V.parahaemolyticus,which will be helpful for the prevention and control of V.parahaemolyticus.