南京市2012年-2013年肠道病毒71型分离株VP1基因特性分析

Analysis of the VP1 region characteristics of enterovirus 71 isolated strains in Nanjing from 2012 to 2013

目的了解南京地区肠道病毒71型流行株的遗传学背景。方法采集2012年-2013年手足口病患者咽拭子样本,经特异性逆转录-聚合酶链式反应（RT-PCR）鉴定EV71病毒核酸阳性者进行细胞接种分离病毒。选取阳性分离株扩增其VP1完整编码区全长基因并进行序列测定,利用生物信息学软件对序列加以分析,构建序列遗传亲缘关系树。结果共分离得到14株EV71毒株,经VP1全长基因测序分析发现,所分离病毒VP1区核苷酸与EV71病毒A型代表株同源性为79.6%～81.3%,与B型代表株的同源性为81.7%～84.7%,而与C型代表株的同源性明显较高,为86.0%～95.2%,其中与C4代表株（AY895144）的同源性最高,为93.8%～95.2%。进化树分析发现本地流行株与上海、安徽等地区流行株的亲缘较近,而与台湾地区流行株亲缘较远。相应的氨基酸位点分析也表明符合C基因型的氨基酸模式。结论南京地区2012年-2013年肠道病毒71型的流行株主要为C4亚型,与上海和安徽地区流行株亲缘较近,可能具有同一来源。

Objective To investigate the genetic background of enterovirus 71 epidemic strains in Nanjing area. Methods Pharyngeal swab samples of patients suffering hand- foot- and- mouth disease（ HFMD） from 2012 to 2013 were collected and the specific reverse transcription polymerase chain reaction（ RT- PCR） was performed to identify whether there was an EV71 virus infection or not. The samples which were EV71 nucleic acid positive were then carried out to a virus isolation process. The VP1 region of EV71 was amplified on those successfully isolated virus strains and subsequently sequenced. All these sequences were analyzed by using bioinformatics software and finally constructed the genetic relationship tree. Results Fourteen EV71 virus strains were isolated,and the nucleic acid sequence homology analysis showed that these isolated strains were 79. 6%- 81. 3%homologous with A reference strains,81. 7%- 84. 7% homologous with B reference strains,and 86. 0%- 95. 2% homologous with C reference strains,especially close to C4 reference strains（ the homology was 93. 8%- 95. 2%）. According to the genetic relationship tree,we found the local epidemic strains were close to strains from Shanghai,Anhui and other regions in genetic relationship,but far from Taiwan strains. Amino acid analysis also showed accordance with amino acid pattern of C genotype.Conclusion The epidemic strains of enterovirus 71 in Nanjing from 2012 to 2013 belonged to subgenotype C4,which were in close kinship with strains in Shanghai and Anhui areas and might have the same source.