2021年云南省红河州手足口病病例标本中分离的2株新型肠道病毒EV-C96 VP1区基因特征分析

Molecular identification of VP1 gene of two novel enterovirus EV-C96 strains from hand,foot and mouth disease cases in Honghe prefecture,Yunnan province,China,2021

目的对2021年云南省红河州手足口病(hand,foot and mouth disease,HFMD)病例标本中分离的2株新型肠道病毒EV-C96的基因特征进行分析,为今后对该病毒的防控提供分子流行病学数据。方法收集2021年1—12月红河州手足口病病例的粪便和咽拭子标本,从处理后的标本中提取病毒RNA,采用实时荧光定量反转录聚合酶链反应(real-time quantitative reverse transcription PCR,real-time qRT-PCR)法进行病毒鉴定。对鉴定出的其他肠道病毒进行基因测序定型。用肠道病毒特异性引物扩增VP1区基因,用DNAStar软件中的Seqman程序对病毒基因进行编辑、整理,得到其VP1区序列。按文献下载EV-C96病毒VP1基因代表株参考序列,用Mega 5.2软件进行基因特征分析。结果2021年红河州共报告手足口病病例6017例,发病率为171.73/10万,其中,经real-time qRT-PCR法确诊的429例手足口病病例中,EV-A71感染57例,占13.29%(57/429),CVA16感染137例,占31.93%(137/429),其他肠道病毒感染235例,占54.78%(235/429),用基因测序法从235例其他肠道病毒感染病例样本中检测到2株EV-C96,占0.85%(2/235)。基因特征分析表明这2株EV-C96病毒属C基因型。结论2021年云南省红河州手足口病病原体以其他肠道病毒为主,其中2株为EV-C96,属C基因型。这是云南省首次从手足口病病例中检测到新型肠道病毒EV-C96。

Objective To analyze the genetic characteristics of two strains of enterovirus C type 96(EV-C96)detected from hand,foot and mouth disease(HFMD)cases in Honghe prefecture,Yunnan province in 2021 and to provide molecular epidemiological data for future prevention and control of the virus.Methods Fecal and pharyngeal swab samples from HFMD cases in Honghe prefecture in 2021 were collected.The virus RNA was extracted directly from the treated specimens.Enteroviruses were identified by real-time quantitative reverse transcription PCR(realtime qRT-PCR).The VP1 genes of other enteroviruses identified by real-time qRT-PCR were amplified and sequenced by specific VP1 gene primers.The sequences were edited and organized with the Seqman program in DNAStar software to obtain its VP1 region sequence.The reference sequences of the VP1 gene of the representative strains of the EV-C96 virus were downloaded from the GenBank,and the genetic characteristics were analyzed by Mega 5.2 software.Results A total of 6017 HFMD cases were reported in Honghe prefecture in 2021,with an incidence rate of 171.73/100000.Among the 429 samples confirmed by real-time qRT-PCR to be HFMD cases,57 were identified as EV-A71,accounting for 13.29%(57/429),and 137 were identified as CVA16,accounting for 31.93%(137/429).There were 235 cases of other enteroviruses,accounting for 54.78%(235/429).Two strains of EV-C96 were detected from 235 cases of non-EV-A71 and non-CVA16 enteroviruses by gene sequencing,accounting for 0.85%(2/235).The two EV-C96 belonged to genotype C.Conclusion In 2021,the pathogens of HFMD in Honghe prefecture,Yunnan province,are mainly other enteroviruses.Among them,only two strains are EV-C96,which belong to genotype C.This is the first report of EV-C96 from HFMD cases in Yunnan,China.