病例来源霍乱弧菌毒力基因特征与Ⅵ型分泌系统功能研究

韩雨; 王堃锟; 苏雅丽; 张凤伟; 段然; 李晓瑞; 肖悦; 逄波; 梁未丽

doi:10.3784/jbjc.202502200112

病例来源霍乱弧菌毒力基因特征与Ⅵ型分泌系统功能研究

Virulence gene characteristics and type Ⅵ secretion system functionality in case-derived Vibrio cholerae strains

摘要

摘要:
目的分析13株病例来源霍乱弧菌Ⅵ型分泌系统（T6SS）功能表达与毒力因子分布特征。
方法通过生物信息学方法分析菌株的序列型（ST）分型、弧菌致病岛、CTXΦ与T6SS组成。采用Western Blot（WB）和细菌竞争实验检测T6SS功能活性及其介导的种间或种内竞争。
结果 13株病例来源霍乱弧菌，均为O1/O139血清群，其中5株携带ctxB10，为产毒株，8株为非产毒株。 13株菌共包括3种ST，ST75型（8株）、ST173型（3株）和ST1481型（2株）。不同ST具有不同的毒力岛与T6SS效应蛋白模块特征：ST75型菌株均含有弧菌致病岛1（VPI-1）和弧菌致病岛2（VPI-2），T6SS为A₁A₁A₁型；ST173型菌株仅含有VPI-1毒力岛，T6SS为C₁A₂A₁型；ST1481型菌株不含VPI毒力岛，T6SS为C₇A₂H₃型。 T6SS功能方面，除1株菌表达但不分泌Hcp外，其余均可正常表达并分泌Hcp，并介导霍乱弧菌对大肠埃希菌的杀伤作用。所有菌株T6SS启动子的SNP45位点均为T。
结论 13株病例来源霍乱弧菌，除1株T6SS失活外，其余菌株T6SS呈组成型表达，介导菌间竞争作用，赋予宿主更强的生态位竞争优势。 ST75型菌株T6SS效应蛋白模块为A₁A₁A₁亚型，能够与第七次大流行El Tor型菌株（7PET）共存，这可能是ST75型霍乱弧菌近年来检出率增加的原因之一。

Abstract:
Objective To analyze the functional expression of the type Ⅵ secretion system （T6SS） and the distribution characteristics of virulence factor in 13 clinical strains of Vibrio cholerae.
Methods The sequence type （ST）, Vibrio pathogenicity island, CTXΦ, and T6SS composition were analyzed through bioinformatics. Western Blot （WB） and bacterial competition assay were used to assess the functional activity of T6SS and its role in interspecies or intraspecies competition.
Results All the 13 clinical strains of V. cholerae belong to O1/O139 serogroups, including 5 toxigenic strains carrying ctxB10 and 8 non-toxigenic strains. These strains are classified into three STs: ST75 （8 strains）, ST173 （3 strains）, and ST1481 （2 strains）. The strains with different STs had distinct virulence islands and T6SS sets: ST75 strains all contain Vibrio pathogenicity island-1 （VPI-1） and Vibrio pathogenicity island-2 （VPI-2）, and their T6SS effector module sets are A₁A₁A₁ subtype. ST173 strains only contain VPI-1, with a C₁A₂A₁ subtype T6SS. ST1481 strains do not contain VPI, and possessing a C₇A₂H₃ subtype T6SS. Regarding T6SS function, all the strains except one expressed and secreted Hcp, mediating the killing effect against Escherichia coli. The SNP45 locus in the T6SS promoter is T in all the strains.
Conclusion Among the 13 clinical V. cholerae isolates, all except one exhibited constitutive expression of T6SS, mediating interbacterial competition and enhancing the ecological niche competitiveness of the host. Notably, the T6SS effector module set of ST75 strains is the A₁A₁A₁ subtype, which can coexist with the seventh pandemic El Tor （7PET） strains, potentially accounting for the increased detection rate of V. cholerae ST75 in recent years.

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