霍乱弧菌和副溶血弧菌分离株的gyrB基因系统发育分析

依据gyrB基因部分编码序列构建系统发育树以分类和鉴别霍乱弧菌和副溶血弧菌,并探讨其种系发生关系。扩增并测序13株霍乱弧菌、8株副溶血弧菌、2株嗜水气单胞菌及1株类志贺邻单胞菌的gyrB基因(编码DNA促旋酶B亚单位)序列,并采用距离法与最大似然法构建系统发育树。两种方法所构建的树结构完全一致,霍乱弧菌、副溶血弧菌、嗜水气单胞菌及类志贺邻单胞菌各自形成一个独立的簇。其中,霍乱肠毒素基因(ctxA)阳性的霍乱弧菌(8株O139群与2株O1群ElTor型)聚类成一分枝;3株副溶血弧菌临床株(1株2002年流行株,2株2004年分离株)与1日本菌株及2001年1株自环境分离的毒力株聚类。系统发育分析靶分子gyrB基因可以良好区分上述4种常见病原菌。产毒O139群霍乱弧菌与产毒O1群ElTor型霍乱弧菌关系密切。副溶血弧菌环境毒力株与本地区临床主要流行株在系统发育关系上较为接近,可能是潜在的致病菌。

Classification and identification of Vibrio cholerae and Vibrio parahaemolyticus isolates based on gyrB gene phylogenetic analysis

In order to validate the usefulness of gyrB genotype for the classification and identification of Vibrio cholerae and Vibrio parahaemolyticus isolates, the phylogenetic analysis of 13 V. cholerae, 8 V. parahaemolyticus, 2 Aeromonas hydrophila and 1 Plesiomonas shigelloides strains was carried out using the partial coding sequence of gyrB, a gene that encodes the B subunit of DNA gyrase (topoisomerase type II ) in bacteria. These strains were separately clustered at species level and typed by the DNA sequences of reference strains from GenBank. CtxA positive V. cholerae strains including 8 clincical isolates of 0139 and 2 clinical isolates of 01 formed one cluster. Four V. parahaemolyticus strains of 1 isolate from 2002 Zhejiang outbreak patient ( tdh positive), 2 clinical isoltates from 2004 and 1 strain from Japan were grouped with an environmental isolate ( trh positive) from 2001. GyrB genotype is applicable to species identification of V. cholerae, V. parahaemolyticus, A. hydrophila and P. shigelloides isolates. The ctxA positive 0139 and 01 group of V. cholerae are closely related, as reflected by gyrB sequence divergence. Furthermore, the toxigenic V. parahaemolyticus strain isolated from environments may be the potential pathogen to the local prevalent and sporadic cases.