2014年上海地区肠道病毒71型全长基因序列和毒力基因位点分析  

Analysis on complete genome and virulence determinants of human enterovirus 71 in 2014 in Shanghai area

作　　者：张晓玲 俞慧菊[3] 宋志刚[2] 王蔚[2] Zhang Xiaoling;Yu Huiju;Song Zhigang;Wang Wei(Shanghai Public Health Clinical Center, Fudan University, Shanghai 201508, China. Pathogen detection and biosafety, Shanghai Public Health Clinical Center Affiliated to Fudan University. Department of Pediatrics, Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine)

机构地区：[1]复旦大学附属公共卫生临床中心公共卫生临床中心院系,上海201508 [2]上海市公共卫生临床中心病原体检测和生物安全部 [3]上海交通大学医学院附属新华医院儿科

出　　处：《国际病毒学杂志》2018年第2期77-82,共6页International Journal of Virology

基　　金：“十二五”国家科技重大专项（2012ZX10004-211）;上海市公共卫生临床中心萌芽计划（2015MY01）

摘　　要：目的分析2014年上海地区手足口病患者中肠道病毒71型（enterovirus71，EV71）的全长基因组序列，了解上海地区EV71流行株的序列特征，推测与致病力相关的毒力基因位点。方法收集上海交通大学附属新华医院2014年1月至12月临床诊断为手足口病的436例患者的临床资料和咽拭子标本。采用实时荧光PCR方法检测通用肠道病毒和EV71型，选取部分核酸阳性标本进行全基因片段扩增和测序，构建全长和3’非编码区（untranslated region，UTR）基因序列进化树，并与EV71的BrCr原型株、柯萨奇病毒A组4型（coxsackievirus A4，CA4）、CA14、CA16原型株和其他肠道病毒A组原型株进行各编码区核苷酸同源性比较。另外，结合GenBank数据库中代表株，比较轻症、重症和死亡组的氨基端序列，寻找差异位点。结果436例手足口病患者中，EV71核酸检测阳性123例（28．21％）。对13个EV71检测阳性标本病毒的全长基因序列和3’UTR序列进行系统进化树分析，发现2014年上海地区EV71流行株为C4a亚型，3’UTR序列与CA4、CA14和CA16原型株序列相近。结合GenBank数据库中代表株，比较EV71轻症、重症和死亡患者的氨基端序列，发现159个蛋白氨基酸序列差异位点，其中Q230L（VP2）、11054V（2B）、V1508I（3A）、V1662I（3C）等4个位点差异有统计学意义（r值分别为8．82、6．09、6．09、6．09，P值分别为0．012、0．048、0．048、0．048）。结论2014年上海地区的EV71C4a亚型流行株3’UTR区与CA4、CAl4和CAl6型存在重组，位于VP2结构区的230位可能是与致病力相关的毒力基因位点。Objective In order to understand the genetic characteristics of enterovirus 71 （EV71） epidemic strains and to speculate the virulent determinants associated with pathogenicity in Shanghai area, we analyzed the complete genome of EV71 from mild and severe hand, foot, and mouth disease （HFMD） patients in Shanghai in 2014. Methods The clinical data and throat swab specimens of 436 patients who were clinically diagnosed as hand-foot-mouth disease from January to December 2014 in Xinhua Hospital of Shanghai Jiao Tong University were collected and were detected by real-time fluorescent PCR for the common enterovirus and EV71. EV71 positive samples were selected for amplification and sequencing of full-length genome. The phylogenetic trees of the full-length sequence and 3＇ untranslated region （3＇- UTR） gene were constructed and compared with EV71 BrCr prototype strain, coxsackievirus A4 （CA4）, CA 14, CA 16 prototype strains and other enterovirus A prototype strains of each coding region nucleotide homology comparison. Combined with the representative strains in the GenBank database, the sequences of the amino-terminus in the mild, severe and death groups were compared to find sites of differences. Results A total of 436 HFMD cases were collected, of which 123 cases （28.21%） were positive for EV71. Phylogenetic analysis based on the complete genome and 3＇-UTR of EV71 was carried out. Phylogenetic analysis of the full-length and 3＇-UTR sequences of 13 EV71 strains revealed that EV71 strains in Shanghai were C4a subtype, the sequence of 3＇-UTR was similar to CA4, CA14 and CA16. Together with EV71 representative strains in the GenBank, we compared the amino acid sequence from mild, severe and fatal cases, 159 different sites of amino acids were found. By Chi-square analysis we found four sites： Q230L （VP2）, I1054V （2B）, V1508I （3A） and V1662I （3C） had statistical significantly difference （ xz values were 8.82, 6.09, 6.09, 6.09, and P values were 0.012, 0.048, 0.048, 0.048,

关 键 词：手足口病 肠道病毒属 基因型 重组 遗传 

分 类 号：R725.1[医药卫生—儿科] R440[医药卫生—临床医学]