手足口病患儿不同病程肛拭子中EV71的分离与鉴定

Separation and Identification of EV71 from Anus Swabs of Children with Different Courses of Hand-foot-and-mouth Disease

目的对4例疑似手足口病患儿的肛拭子标本进行EV71的分离与鉴定。方法采集患儿不同病程肛拭子17份接种Hela细胞,盲传3代,观察细胞病变;提取RNA,逆转录,设计三对肠道病毒特异性引物PCR扩增细胞培养物上清中的EV71,分离鉴定。结果具体样本均能引起Hela细胞产生CPE;细胞培养前肛拭子EV71的RT-PCR阳性检出率为52.9%(9/17),培养后细胞上清的阳性检出率为88.2%(15/17);4例患儿全部诊断为肠道病毒EV71感染引起的手足口病。结论病毒分离和核酸检测的联合应用能为肠道病毒EV71的临床诊断提供有力依据;通过病毒分离能提高EV71的核酸检测的阳性率。

Objective To separate and identify the EV71 virus from anus swabs of children suspected with hand - foot and mouth disease. Methods The anus swabs from children with different courses of hand- foot- and mouth disease were centrifugated for inoculation with Hela cells and then blindly transferred to the third generation for observing the cytopathic effect. Then, reverse transcription was conducted after extracting the total RNA, three pairs of PCR primers were designed to amplify the nucleic acid; the cases were diagnosed by the clinical symptoms and an optimal therapy model was set up. Results Most samples caused the Hela cell cytopathic effect. The positive isolating rate was 52.9 % （9/17） before the viral isolation, while the positive isolating rate was 88.2 % （15/17） after the viral isolation. Four cases were diagnosed as hand foot - and - mouth disease which was caused by enterovirus71 infection. Conclus[ons The unification of nucleic acid detection by RT - PCR and viral isolation can offer a powerful way to detect the EV71 infection. Viral isolation can enhance the posi- tive detection rate of EV71 by RT- PCR.