摘要
目的对2株犬源狂犬病病毒街毒株全基因组测序,了解国内狂犬病病毒的流行和变异情况。方法采用乳鼠脑内接种方法分离2株天津地区狂犬病病毒街毒株,RT-PCR扩增病毒DNA覆盖全基因组12个相互重叠的基因片段,PCR产物平端克隆后进行全基因组核苷酸序列测定,然后对其分子变异和进化特点进行分析。进行两分离株的序列相似性、氨基酸同源性、主要功能位点的变异比较和种系发生分析。结果 TJD04和TJD05两株狂犬病病毒全基因组长均为11 924 nts,全基因组序列的组成和结构均符合弹状病毒科狂犬病病毒属的特征。病毒基因组由5个编码区组成,基因起始位点和终止位点高度保守,氨基酸编码长度未发生变异,5个编码蛋白序列较少发生变化,仅有个别主要功能位点发生变异,且大部分变异均属于无意义沉默突变。多序列相似性比较和种系发生分析显示,两株病毒均属于基因1型,N基因与全基因进化树保持一致,与中国犬源狂犬病病毒BD06同源性最高(99.6%),进化关系近,并具有较为独特的中国地域特点。结论两天津株狂犬病毒可能是自然界中固有的狂犬病病毒街毒株。
Objective In order to ascertain the prevalence and diversity of rabies viruses in China,two wild-type strains of rabies virus were isolated by intracerebral inoculation of mouse pups with homogenate samples.Methods The genomes of two isolates were amplified by RT-PCR with series of specified primers and cloned into pEASY-Blunt vectors.The recombinant plasmids were sequenced and the obtained sequences of virus were processed and assembled with Seqman software from DNAstar.Results Analysis of the diversity of the viral genomes,amino acid residues,and functional sites revealed that the genomes of the two rabies viruses（TJD04 and TJD05） have the same nucleotide length（11924 nts）.The two genomes had similar nucleotides and amino acid residues and were homologous to Rhabdoviridae Lyssavirus according to BLAST and multi-sequence alignment.Both viruses had a genome with 5 ORFs,start and stop sites of genes were highly conserved,and both coded for the same number of amino acids.Compared to reference rabies viruses,the two viruses had little variation in terms of coded protein sequences but did vary in terms of the main functional sites of genes.The phylogenic tree showed that both viruses were type I genes and the viruses were highly homologous to the rabies strain BD06 from China.Both isolates were closely related evolutionarily and differed from other strains of the rabies virus from China（BD06）.Conclusion Data showed that two isolates were probably natural wild-type forms of the rabies virus particular to China.
出处
《中国病原生物学杂志》
CSCD
2011年第7期481-485,489,F0003,共7页
Journal of Pathogen Biology
基金
国家重点基础研究发展计划(973计划)项目(No.2011CB504706)
国家科技重大项目(No.2009ZX10602-14)
关键词
狂犬病病毒
街毒株
全基因组
序列分析
Rabies virus
wild-type
full-length genome
sequencing and analysis
