CAJ | 학술논문

　　副溶血弧菌和霍乱弧菌（（non-O1/non-O139型），不仅对水产动物具有广泛和强的致病作用，而且危害人类的健康，是人和水产动物共患病的病原菌。以副溶血弧菌的 toxR 基因及霍乱弧菌的lolB 基因设计2对引物，建立了副溶血弧菌和霍乱弧菌的 PCR 同步检测方法，相应的扩增目的片段分别为368 bp 和519 bp 。在 PCR 反应体系中副溶血弧菌和霍乱弧菌可扩增出目的基因片段，5株对照菌无任何扩增条带，表明所建立的双重 PCR 检测方法特异性较强，且所设计的2对引物间无交叉干扰现象；灵敏性试验结果表明，副溶血弧菌的检测最低限1．75×102 cfu/mL ，霍乱弧菌的检测最低限1．25×102 cfu/mL ，具有较好的灵敏性；人工染菌水产品检测结果显示，人工染菌的8种水产品均为阳性检测结果，而未染菌组均为阴性。本试验建立的双重 PCR 法特异性强、灵敏性高，可用于副溶血弧菌和霍乱弧菌引起的水产动物疾病的诊断及水产品的安全检测。
　　부용혈호균화곽란호균（（non-O1/non-O139형），불부대수산동물구유엄범화강적치병작용，이차위해인류적건강，시인화수산동물공환병적병원균。이부용혈호균적 toxR 기인급곽란호균적lolB 기인설계2대인물，건립료부용혈호균화곽란호균적 PCR 동보검측방법，상응적확증목적편단분별위368 bp 화519 bp 。재 PCR 반응체계중부용혈호균화곽란호균가확증출목적기인편단，5주대조균무임하확증조대，표명소건립적쌍중 PCR 검측방법특이성교강，차소설계적2대인물간무교차간우현상；령민성시험결과표명，부용혈호균적검측최저한1．75×102 cfu/mL ，곽란호균적검측최저한1．25×102 cfu/mL ，구유교호적령민성；인공염균수산품검측결과현시，인공염균적8충수산품균위양성검측결과，이미염균조균위음성。본시험건립적쌍중 PCR 법특이성강、령민성고，가용우부용혈호균화곽란호균인기적수산동물질병적진단급수산품적안전검측。
V ibrio parahaemolyticus and V . cholerae were known to be responsible not only for aquatic animals ,but also to be recognized as pathogenic bacteria for human .In this study ,two pairs of specific primers were designed based on toxR gene for V . parahaemolyticus and lolB gene for V .cholerae ,and simultaneous detection of the both V ibrio was established by a dulplex polymerase chain reaction (PCR) , amplification of 368 bp and 519 bp gene fragments .The results showed that two purposed gene fragments from mixed chromosomal DNA of V . parahaemolyticus and V . cholerae in one PCR reaction were simultaneously amplified by two PCR primers ,and no positive reaction was detected in 5 other controlled pathogenic bacteria , showing strong specificity , and no interference between the two primers . The sensitivity test of the dulplex PCR revealed that the two primers detected V .parahaemolyticus at a level of as few as 1 .75 × 102 cfu/mL ,and V .cholerae at a level of as few as 1 .25 × 102 cfu/mL .The positive amplified results were observed in the 8 aquatic products infected artificially ,and negative amplifi cation in the no-infected 8 aquatic products . The findings prove that the dulplex PCR is a specific and sensitive method for simultaneous detection of the both V ibrio in one PCR reaction ,and can be used as the detection of food safety ,and diagnose of aquatic animals diseases caused by the both V ibrio .