一步法实时RT-PCR快速检测手足口病肠道病毒  

The application of TaqMan one-step real-time RT-PCR in rapid diagnosis of hand,foot and mouth disease

作　　者：林裕龙[1] 温坤[1] 潘玉先[1] 王压娣[1] 晋晶[1] 车小燕[1] 

机构地区：[1]南方医科大学珠江医院医学检验中心,广州510282

出　　处：《临床检验杂志》2010年第3期169-171,共3页Chinese Journal of Clinical Laboratory Science

基　　金：十一五国家科技重大专项(2009ZX10004-306)

摘　　要：目的探讨实时荧光RT-PCR在手足口病(HFMD)肠道病毒快速检测中的应用价值。方法以实时荧光RT-PCR检测64例临床初诊为HFMD患儿标本中总肠道病毒(EV)、柯萨奇病毒A组16型(CA16)和肠道病毒71型(EV71),用RD细胞、Vero细胞和Hep-2细胞进行病毒分离培养,以免疫荧光法鉴别分离培养的EV71。结果64例标本中,EV(+)/EV71(+)为23.4%(15/64)、EV(+)/CA16(+)为48.4%(31/64)、EV(+)/EV71(-)/CA16(-)为10.9%(7/64)、EV(-)/EV71(-)/CA16(-)为17.2%(11/64)。实时荧光RT-PCR检测为EV71且病毒分离培养出现细胞病变效应的培养细胞经抗EV71单克隆抗体免疫荧光检测均为阳性,未发生细胞病变效应或非EV71的培养细胞均为阴性。结论实时荧光RT-PCR检测HFMD病原具有特异和快速等优点,与病毒分离培养具有较高的一致性,可用于手足口病的早期诊断和鉴别诊断。Objective To evaluate one step real-time fluorescent RT-PCR in diagnosis of hand,foot and mouth disease（HFMD）.Methods The nucleic acids of enterovirus（EV）,Coxsackie virus A Group 16（CA16）,and enterovirus 71（EV71） in 64 specimens from pediatric patients with suspected HFMD were detected by the real-time RT-PCR.Immunofluorescence assay（IFA） was used to identify EV71 after virus was cultured in RD cell,Vero cell,and Hep-2 cell,respectively.Results Among 64 specimens,EV（＋）/EV71（＋）,EV（＋）/CA16（＋）,EV（＋）/EV71（-）/CA16（-） and EV（-）/EV71（-）/CA16（-） accounted for 23.4%（15）,48.4%（31）,10.9%（7）,and 17.2%（11）,respectively.Five EV71 isolates with cytopathic effect in cell culture were positive by IFA with EV71 monoclonal antibody,while those without cytopathic effect or infected with non-EV71 were negative.Conclusions Real-time fluorescence RT-PCR may detect HFMD quickly and specifically.It is consistent with virus isolation and can be used to early diagnose HFMD.

关 键 词：手足口病 实时荧光RT-PCR 肠道病毒 免疫荧光检测 

分 类 号：R373.2[医药卫生—病原生物学]