摘要
目的 了解2008-2009年北京分离的肠道病毒71型（EV71）基因组3＇末端的基因特征（未包括多聚腺苷尾）,探讨是否与病毒的致病性有关.方法 于2008-2009年手足口病流行期间采集来首都儿科研究所附属儿童医院就诊的普通手足口病患儿和合并重症神经系统并发症患儿的咽/鼻拭子或疱疹液标本.将标本接种Vero细胞进行肠道病毒分离,同时设计肠道病毒通用引物、EV71和柯萨奇病毒A组16型（CA16）特异性引物通过巢式RT-PCR对分离到的肠道病毒进行型别鉴定.对引起不同临床类型感染的10株EV71分离株的基因组3＇末端进行序列测定和分析.结果 10株EV71的3＇末端均包含1386 nt的3D、终止密码子TGA和81 nt的3＇UTR,由3D核苷酸序列所推导的3D蛋白含462个氨基酸.10株EV71的3D和3＇UTR核苷酸同源性分别为95.8%～99.6%和96.3%～100%,重症来源的4株毒株中有3株在3Dpol第140位和第263位同时出现了相同的氨基酸变异（R140K和I263V）.10株EV71的3D与C4亚型代表株具有最高的核苷酸和氨基酸同源性,分别为92.7%～94.2%和96.8%～97.6%;在3＇UTR与CA16/G10具有最高的核苷酸同源性（88.9%～91.4%）.3D区的遗传进化分析表明10株EV71在亲缘关系上与C4亚型最密切,与CA16/G10较密切,而与EV71其他基因型和亚型较疏远.结论 基因组3＇末端在EV71进化中可能有一定的作用.
Objective To investigate the sequences of the 3＇ end of genomes from enterovirus 71 isolated from pediatric patients with different symptoms in Beijing during 2008-2009.Methods Clinical specimens were collected from pediatric patients suffering from hand,foot and mouth disease（HFMD）and/or with neurological complications who visited the affiliated Children＇s Hospital during the epidemic seasons of 2008 and 2009 in Beijing.The samples were inoculated into the Vero cell line,and the virus isolates were further identified by nested reverse transcription-nested PCR（RT-nPCR）assay using universal enterovirus primers,type specific primers for EV71 and CA16.The 3＇ end of genomes（including 3D and 3＇ UTR regions）from 10 EV71 strains derived from various clinical presentations were amplified and sequenced.Results Analysis of the nucleotide sequences of the amplified fragments showed that the 3＇ end of genomes of 10 EV71 isolates include the 3D region of 1386 nucleotides（nt）encoding the 3D polymerase（3Dpol,462 amino acids）,the terminator codon TGA and 3＇ UTR of 81 nt.Nucleotide sequence identities among 3D regions from these 10 EV71 isolates were in the range of 95.8%-99.6%,while the nucleotide sequence identities for 3＇ UTR were 96.3%-100%.The majority of nucleotide changes were located at the third codon positions which caused silent mutations,thus the amino acid sequence changes of 3Dpol among those 10 EV71isolates were scanty.The residues 140 and 263 which were R and 1 were substituted by K and V,respectively in 3 of 4 neurovirulent strains,whereas only 1 of 6 strains from mild cases had these 2 amino acid changes.The sequences of the 3D and 3＇ UTR regions of 10 EV71 isolates were compared to the representative strains of known genotypes from the GenBank.The nucleotide and amino acid sequences of 10 EV71 isolates in the 3D region exhibited highest homology to the subgenotype C4 of EV71（92.7%-94.2%and 96.8%-97.6%.respectively）.However,3＇ UTR of 10 EV71 isolates shared the highest n
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2010年第5期392-398,共7页
Chinese Journal of Microbiology and Immunology
基金
国家科技支撑计划（2008BA170B00）
