EV71和Cox A16型肠道病毒RT-PCR检测方法的研究

    刘胜牙，朱玉兰，王佃鹏，古莉冰，甄胜西，徐媛，赵倩，高朝贤

    深圳国际旅行卫生保健中心，广东 深圳 518033

    摘要：目的 建立咽拭子标本中EV71和CoxA16型肠道病毒逆转录聚合酶链反应（RT-PCR）检测方法。方法 根据GENBANK数据库中EV71和CoxA16型肠道病毒的基因序列，利用CLUSTAL软件进行序列比对分析，选择基因组的保守序列，并借助引物设计生物信息学软件，设计筛选出一套针对EV71和CoxA16型肠道病毒的RT-PCR扩增的引物。从特异性、最低检测限、重复性等方面进行评估，建立了EV71和CoxA16型肠道病毒RT-PCR检测方法。结果 RT-PCR方法对EV71和COXA16病毒的最低检测量分别为63 TCID50/ml和32 TCID50 /ml。其他病原微生物及人类基因组在该系统无反应信号，显示方法特异性良好。结论RT-PCR对手足口病病原体检测准确性高、特异性强，具有快速、经济等特点，适合手足口病的早期诊断。

    关键词:手足口病；肠道病毒71型；柯萨奇病毒A16型；逆转录聚合酶链反应

    中图分类号：R512.5　文献标识码：B

    Study on RT-PCR detecting method for humanenterovirus 71 and

    Coxsackie A16 type virus

    LIU Sheng-ya, ZHU Yu-lan, WANG Dian-peng, GULi-bing, ZHEN Sheng-xi, XU Yuan, ZHAO Qian, GAO Chao-xian

    Shenzhen International Travel Health CareCenter, Shenzhen, Guangdong 518033, China

    Abstract:   Objective  To study a RT-PCR method for detecting EV71 and Cox A16 virus fromthroat swab specimen． Methods   Specific primers weredesigned based on highly conserved region of EV71 and Cox A16depending on bioinformatics software．Specificity, minimum detectionlimits and repeatability of the method wereassessed.    Results   The minimumdetection limits of the method for EV71 and Cox A16 virus were 63TCID50/ml and 32 TCID50/ml, respectively. Other pathogenicorganisms and human genome in the method didn't show any responsesignal, displaying good specificity of the method.  Conclusion   Our results indicate RT-PCR offers a rapid,sensitive, specific and cheap method to detect pathogen ofhand-food and mouth disease.

    Key words:   Hand-foot and mouthdisease；Human enterovirus 71；Coxsackie virus A16；Reversetranscription-PCR

    《中国国境卫生检疫杂志》2012年8月刊