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目的对东莞市2014年手足口病(HFMD)疑似病例样本进行病原学检测,并对引起HFMD的肠道病毒71型(EV71)和柯萨奇病毒A16型(CA16)进行基因特征分析,为东莞市HFMD的防控提供基础资料。方法采用Real timeRT-PCR(实时荧光PCR)方法对HFMD疑似病例样本进行检测,核酸阳性样本上送广东省疾病预防控制中心,进行病毒分离及鉴定,基因扩增并测定VP1基因序列。从Gen Bank中选取EV71和CA16不同基因型参考毒株,利用DNA-STAR软件进行核苷酸、氨基酸序列分析和同源性比较,并用Mega4.0.2软件构建亲缘性进化树。结果共检测HFMD疑似病例样本693份,核酸阳性结果614份,总阳性率为88.60%。其中EV71阳性结果241份,占39.25%,CA16阳性194份,占31.60%,其他肠道病毒(EV)179份,占29.15%。共获得11株EV71分离株和8株CA16分离株的VP1基因全长序列。11株EV71分离株的核苷酸和氨基酸序列同源性分别为94.3%~100.0%和98.3%~100.0%,8株CA16分离株的核苷酸和氨基酸序列同源性分别为89.9%~100.0%和99.3%~100.0%。系统进化树分析显示,11株EV71全部属于C4a基因亚型;8株CA16分离株中有6株属于B1b基因亚型,2株属于B1a基因亚型。结论 2014年引起东莞市HFMD的EV71型病毒流行株为C4a基因亚型,CA16型病毒流行株B1a和B1b亚型并存,并兼有一定比例的其他肠道病毒。
Objective To analyze the etiology of suspected cases of hand-foot-mouth disease (HFMD) in Dongguan City in 2014 and analyze the genetic characteristics of EV71 and C16 of Coxsackie virus type A16 (CA16) City HFMD prevention and control to provide the basic information. Methods Real-time PCR-based real-time PCR (RT-PCR) was used to detect suspected cases of HFMD. The positive samples of nucleic acid were sent to Guangdong Provincial Center for Disease Control and Prevention for virus isolation and identification, gene amplification and determination of the VP1 gene sequence. The genomic reference strains EV71 and CA16 were selected from GenBank, nucleotide and amino acid sequence analysis and homology comparison were made by DNA-STAR software, and phylogenetic tree was constructed by Mega4.0.2 software. Results A total of 693 suspected cases of HFMD were detected, and 614 were positive for nucleic acid. The total positive rate was 88.60%. Among them, 241 were positive for EV71, accounting for 39.25%, 194 were positive for CA16, accounting for 31.60%, and 179 were other enterovirus (EV), accounting for 29.15%. The total length of the VP1 gene of 11 EV71 isolates and 8 CA16 isolates was obtained. The nucleotide and amino acid sequence homologies of 11 EV71 isolates were 94.3% -100.0% and 98.3% -100.0%, respectively. The nucleotide and amino acid sequence homologies of 8 strains of CA16 were 89.9% -100.0 % And 99.3% ~ 100.0%. Phylogenetic tree analysis showed that 11 EV71 belonged to C4a genotypes; 6 of 8 CA16 isolates belonged to B1b genotype and 2 belonged to B1a genotype. CONCLUSION: The EV71 strain of epidemic strain causing HFMD in 2014 was C4a subtype. The subtype of CA16 strain B1a and B1b co-existed with other enteroviruses.