环介导等温扩增法快速检测手足口病病原体  

Rapid and Economical Detection of Pathogens of Hand-foot-and-mouth Disease by Reverse Transcription Loop-mediate Amplification

作　　者：戴颖[1] 刘峰涛[2] 何书[1] 汪奇伟[1] 

机构地区：[1]河南大学淮河医院,河南开封475003 [2]河南大学医学院

出　　处：《中国医学创新》2014年第25期68-71,共4页Medical Innovation of China

摘　　要：目的:建立逆转录(RT)-环介导等温扩增方法(LAMP),以快速检测手足口病最重要的两种病原体:肠道病毒71型(EV71)和柯萨奇病毒A组16型(CA16)。方法:收集手足口病患儿咽拭子标本93份,针对EV71和CA16病毒的VP1基因特异性序列8个区域各设计6条LAMP引物,分别于63℃(EV71)、65℃(CA16A)扩增1 h,日光下观察结果,与实时荧光定量PCR比较检测特异性和敏感性。结果:EV71、CA16的LAMP最低检测限均为500拷贝/管,与对照病毒无交叉反应。93份标本中,RT-PCR方法检测显示EV71阳性44例,CA16阳性36例;RT-LAMP方法检测显示EV71阳性46例,CA16阳性36例,两种方法间比较差异均无统计学意义(P>0.05)。结论:应用RT-LAMP检测手足口病病原体EV71、CA16快速、灵敏、经济、特异性高,适合在基层医疗机构推广应用。Objective：To rapidly detect pathogens of hand-foot-and-mouth disease （HFMD） by reverse transcription loop-mediated isothermal amplification （RT-LAMP）,the two most important pathogens：enterovirus 71 （EV71） and coxsackievirus group A type 16 （CA16）.Method：A total of 93 swab specimens were collected from HFMD children.Six primers which recognized 8 distinct regions on the VP1 gene of enterovirus 71 （EV71） and coxsackievirus A16 （CA16） were designed for RT-LAMP assay.The target genes were amplified for 1 hour at isothermal temperatures of 63 ℃ for EV71 and 65 ℃ for CA16,respectively.The sensitivity and specificity of RT-LAMP were compared with fluorescence quantitative PCR using chi-square test measures.Result：The lower limits of EV71 and CA16 detection were both 500 copies/tube by RT-LAMP assay.No cross reaction was shown among other viruses.Of all the 93 swab specimens, 46 cases were EV71 positive and 36 cases were CA16 positive by RT-LAMP,while 44 cases were EV71 positive and 36 cases were CA16 positive by quantitative PCR.There were no significant differences between the two measures（P〉0.05）. Conclusion：RT-LAMP may be a more rapid,sensitive,specific and economical measure for detection of HFMD pathogens,especially in primary health service.

关 键 词：逆转录-环介导等温扩增技术 肠道病毒71型 柯萨奇病毒A组16型 手足口病 

分 类 号：R725.1[医药卫生—儿科]