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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 isolate and identify EV71 from four swabs suspected to be hand, foot and mouth disease in children. Methods Seventeen Hela cells were inoculated blindly for 3 passages in different stages of infection. RNA was extracted and reverse transcribed. Three pairs of enterovirus specific primers were designed and used to amplify EV71 in cell culture supernatant ,clementines. Results The specific samples all could induce CPE in Hela cells. The positive rate of RT-PCR in rectal swab EV71 was 52.9% (9/17) before culturing and the positive rate of cell supernatant in culture was 88.2% (15 / 17). All four cases were diagnosed as hand, foot and mouth disease caused by enterovirus EV71 infection. Conclusion The combined application of virus isolation and nucleic acid detection can provide a strong basis for clinical diagnosis of enterovirus EV71. The positive rate of nucleic acid detection of EV71 can be increased by virus isolation.