云南省人类埃可病毒19型的分子流行病学分析

目的分析云南省1997-2013年急性迟缓型麻痹(AFP)病例和健康儿童中分离到的人类埃可病毒19型(echovirus 19,E19)VP1区基因特征及其分子流行病学特点。方法对分离到的非脊灰肠道病毒(non-polio enterovirus,NPEV)VP1区基因进行逆转录聚合酶链反应(RT-PCR),将PCR产物送生物工程(上海)有限公司用双脱氧链终止法进行双向测序,采用Sequencher4.0软件对序列进行编辑,按Obester的方法对病毒进行定型,对E19病毒与基因库(GenBank)和文献中检索到的国内外不同时期的E19VP1区进行比较。用Neighbor-joining method构建病毒基因进化树,用1 000个Psudoreplicate datasets进行Bootstrap统计学分析。结果云南省1997-2013年从急性迟缓型麻痹(AFP)病例和健康儿童共3 750例中分离到26株E19病毒,与GenBank和文献中检索到的28株进行基因进化分析,54株病毒可分为6个基因型。云南省不同来源(AFP或健康人群)、不同时间分离到的E19主要分布在2、4、5型的基因进化分支,E19具有基因、地理和时间分布多样性特点。结论利用基因测序的方法持续对云南省AFP病例和健康儿童进行肠道病毒监测,增加和更新病毒基因序列,对今后开展肠道病毒的分子流行病学研究,确认病毒传播来源具有重要意义。

Molecular epidemiological analysis of ECHO19 virus isolated from Yunnan province of China

Objective To explore the VP1 gene characteristics and molecular epidemiology of echovirus 19 （E19） isolated from acute flaccid paralysis （AFP） cases and healthy children from Yunnan province of China from 1997 to 2013. Methods RT-PCR was done to amplify the VP1 gene from isolated non-polio enterovir- uses （NPEVs）, PCR products were sent to Sangon Biotech Co. Ltd. （Shanghai, China） for sequencing by fluorescent labeled Sanger method. The sequences were edited by Sequencher 4.0 software, and the virus sero- types were determined according to Obesterrs method. El9 sequences and molecular epidemiologic features were analyzed and compared with that downloaded from the GenBank. The phylogenetic tree was constructed by using 1 000 bootstrap values for statistical analysis. Results Altogether there were 26 El9 virus strains were isolated from 3 750 AFP cases and healthy children in Yunnan province of China from 1997 to 2013. Phylogenetic analysis with other 28 strains in GenBank showed that these 54 E19 strains were grouped into 6 genotypes. E19 viruses isolated from different sources （AFP or healthy children） and different times were mainly clustered into genotypes 2, 4 and 5 with multiple geographical and time characteristics. Conclu- sions Continuous molecular surveillance for enteroviruses from AFP cases and healthy children by sequencing is a very important tool for conducting molecular epidemiological study and identifying the disease transmission pathways in Yunnan province of China.