CAJ | 학술논문

目的对深圳地区手足口病(hand,foot and mouth disease,HFMD)肠道病毒71型分离株(enterovirus 71,EV71)的VP4基因遗传进化进行分析.方法 2009年采集深圳市儿童医院就诊的HFMD患者的粪便标本491份,选取其中8份经细胞培养鉴定为阳性的EV71毒株用RT-PCR扩增VP4基因,利用MEGA4.0软件进行遗传进化分析.结果 2009年深圳地区8株EV71 VP4基因全长207 bp,编码69个氨基酸；8株EV71 VP4基因核苷酸同源性为94.2％～98.1％,与深圳2001至2004年分离的17株EV71毒株核苷酸同源性在89.9％～98.1％,与GenBank中其他EV71病毒株VP4的核苷酸同源性为79.2％～100％；其中与亚洲流行株阜阳株(EU703812)核苷酸的同源性最高(100％),其次是C4代表株及2004年深圳株(AY895144)为94.2％～97.1％.除了印度株和其中1株EV71的VP4编码的氨基酸在第54位(ACA)不同之外,其余7株EV71 VP4编码的氨基酸序列之间以及与GenBank报道的其他序列同源性达100％.8株EV71 VP4基因核苷酸与C4代表株相比有17处不同,除1处以外其余全在简并密码位点上；轻重症病例毒株之间VP4基因序列未见明显改变.进化树显示8株EV71均属于C4基因亚型.结论 2009年深圳市流行的EV71属于C4基因亚型,流行的EV71 VP4基因非常保守,不属于变异区段,绝大部分核苷酸的变异属无义突变,VP4基因编码的氨基酸变异几乎为0.
목적 대심수지구수족구병(hand,foot and mouth disease,HFMD)장도병독71형분리주(enterovirus 71,EV71)적VP4기인유전진화진행분석.방법 2009년채집심수시인동의원취진적HFMD환자적분편표본491빈,선취기중8빈경세포배양감정위양성적EV71독주용RT-PCR확증VP4기인,이용MEGA4.0연건진행유전진화분석.결과 2009년심수지구8주EV71 VP4기인전장207 bp,편마69개안기산；8주EV71 VP4기인핵감산동원성위94.2％～98.1％,여심수2001지2004년분리적17주EV71독주핵감산동원성재89.9％～98.1％,여GenBank중기타EV71병독주VP4적핵감산동원성위79.2％～100％；기중여아주류행주부양주(EU703812)핵감산적동원성최고(100％),기차시C4대표주급2004년심수주(AY895144)위94.2％～97.1％.제료인도주화기중1주EV71적VP4편마적안기산재제54위(ACA)불동지외,기여7주EV71 VP4편마적안기산서렬지간이급여GenBank보도적기타서렬동원성체100％.8주EV71 VP4기인핵감산여C4대표주상비유17처불동,제1처이외기여전재간병밀마위점상；경중증병례독주지간VP4기인서렬미견명현개변.진화수현시8주EV71균속우C4기인아형.결론 2009년심수시류행적EV71속우C4기인아형,류행적EV71 VP4기인비상보수,불속우변이구단,절대부분핵감산적변이속무의돌변,VP4기인편마적안기산변이궤호위0.
Objective To analyze the genetic evolution for the common causative agent of hand,foot and mouth disease(HFMD) that VP4 of human enterovirus 71 in Shenzhen district.Method 491 stool specimen were collected from children with hand,foot and mouth diease in Shenzhen Children's Hospital 2009.After cell culture,VP4 gene of eight EV71 strains were amplified by reverse-transcriptase PCR(RT-PCR),phylogenetic analysis of the VP4 gene was constructed by using MEGA 4.0.Result The VP4 gene of eight EV71 strains encoding 69 amino acids with full length 207 bp.The nucleotide homology of VP4 gene among eight EV71 strains was 94.2％-98.1％,compared with VP4 gene of EV71 strains retrieved from Shenzhen 2001 to 2004 and GenBank was 89.1％-98.1％ and 79.2％-100％ respectively.Asian epidemic strain Fuyang had the highest nucleotide homology,representative strain C4 and Shenzhen strain (AY895144) with 94.2％-98.1％ secondly.Except for the 54th amino acid of VP4 gene of India reported strain and one of the eight EV71 strains,the homology of the rest amino acids between the eight EV71 strains and those in GenBank was 100％.Compared with representative strain C4,there were seventeen differences in nucleotide sequences of VP4 of the eight EV71 strains.All of the different nucleotides were located at the degenerate password sites except one.There was no significant difference in VP4 gene between the severe and the mild cases of strains.The eight Shenzhen EV71 strains were classified as sub-genotype C4 in the phylogenetic tree.Conclusions The epidemic of EV71 in Shenzhen 2009 was sub-genotype C4.VP4 gene of EV71 was very conservative which dose not belong to the variation section.The variation of most of nucleotide was invalid variation.The amino acids encoded by VP4 gene which variation was almost zero.