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目的为探讨2012-2013年贵州省儿童手足口病非CVA16、非EV71肠道病毒株的毒株型别及分子进化特征,从而为制订防治手足口病的措施提供科学依据。方法收集2012~2013年贵州省儿童手足口病疑似患者肛拭子200例标本,应用实时荧光逆转录-聚合酶链反应法筛选,获得非EV71、非CVA16型肠道病毒株。提取所得病毒核酸,采用RT-PCR法扩增病毒VP1区序列,PCR阳性扩增产物进行测序,对测得序列进行型别鉴定,并与国内外各型代表株进行核苷酸、氨基酸同源性比较,构建系统进化树。结果本研究共分离到6株非CVA16、非EV71毒株,分别为:CVA5 2株,CVA2、CVA4、CVB2、E16各1株。分别归属Cox A、Cox B、埃可肠道病毒。同源性分析显示,6株毒株与其代表株核苷酸序列均存在不同程度的差异。系统进化树表明,CVA4、CVB2、E16与国内分离株存在一定亲缘关系,而CVA2、CVA5与国内外分离株亲缘关系较远。结论贵州省儿童手足口病的病原以非CVA16、非EV71肠道病毒株为主要类别。
Objective To explore the strain types and molecular evolutionary characteristics of non-CVA16 and non-EV71 enterovirus strains of hand, foot-mouth disease and childhood in 2012-2013 in Guizhou Province, and to provide a scientific basis for the development of measures to prevent hand-foot-mouth disease. Methods 200 samples of suspected swine anus swab from children with hand, foot and mouth disease in Guizhou province from 2012 to 2013 were collected and screened by real-time fluorescence reverse transcription-polymerase chain reaction to obtain non-EV71 and non-CVA16 enterovirus strains. The obtained viral nucleic acid was extracted, and the VP1 region of the virus was amplified by RT-PCR and the positive PCR products were sequenced. The sequences of the VP1 region were sequenced, and the nucleotide and amino acid homology Compare sex and construct phylogenetic tree. Results Six non-CVA16 and non-EV71 strains were isolated in this study. They were 2 strains of CVA5, 1 strain of CVA2, CVA4, CVB2 and E16 respectively. Respectively belong to Cox A, Cox B, E coli enterovirus. Homology analysis showed that the nucleotide sequences of the 6 strains and their representative strains all had different degrees of difference. Phylogenetic tree showed that CVA4, CVB2 and E16 had some genetic relationship with domestic isolates, whereas CVA2 and CVA5 had distant genetic relationship with domestic and foreign isolates. Conclusion The pathogens of hand-foot-mouth disease in children in Guizhou Province are mainly non-CVA16 and non-EV71 enterovirus strains.