产肠毒素霍乱弧菌的快速鉴定及基因分型

Rapid detection of the toxigenic vibrio cholerae and genotyping

  • 摘要: 目的: 为适应临床诊断和食品检测的需要,探讨快速检测霍乱弧菌及其分型的方法。方法: 用聚合酶链反应(PCR)检测霍乱弧菌肠毒素(CTX) A2 B亚单位靶序列基因,结合随机扩增多态性DNA (RAPD),对霍乱弧菌进行基因分型。结果: 检测的3株O139群霍乱弧菌、3株ElTor生物型和4株古典生物型均含有(CTX) A2 B亚单位基因,核苷酸序列分析证实其同源(97 1%~98 9%)。RAPD中用普通引物将不同弧菌分成2种基因谱;即①O139群和ElTor型;②古典型。用重复序列引物进一步将霍乱弧菌分成3型,即①O139群;②ElTor型;③古典型。重复实验结果稳定。结论: 用PCR结合RAPD检测霍乱弧菌并对其进行分型,操作简便、快速、敏感而且准确,对临床诊断、食品检测和流行病学调查具有一定的实用性。

     

    Abstract: Objective: To study a method quickly to indentify toxigenic vibrio cholerae and genotyping for meeting the requirement of clinical diagnosis and test of foods.Methods: Polymerase chain reaction (PCR)was used to detect cholera toxin(CTX) A2 B subunit gene of a target and randomly amplified polymorphic DNA (RAPD) to play a role of genotype.Results: 3 isolates of V cholerae strains O139, 3 isolates O1 biotype El Tor, 2 isolates O1 biotype classical containedthe 566bp segment of genomic sequences of ctx A2 B which were homologous (97 1%~98 9%)confirmed by nuceotide sequence analysis The isolates of V cholerae were divided into 2 kinds of genetic fingerprint by RAPD with the common primers, that is (1) V cholerae O139 and V cholerae O1 El Tor biotype, (2)V Cholerae O1 classical,and 3 kinds of genetic fingerprint with the repeative primers, that is (1)V cholerae O139,(2)V cholerae O1 El Tor biotype, (3)V Cholerae O1 classical The results of test was stable.Conclusion: It was simple, rapid, sensitive and accurate that PCR and RAPD were used to detect toxigenic V cholerae and identify their genetic pattern, which play a utilitary role in the clinical diagnosis,test of foods and the epidemiologic investigation

     

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