云南省狂犬病病人和犬类感染狂犬病病毒及M基因序列分析

Survey on rabies viruses in rabies patients and dog population from endemic sites of Yunnan Province,China and the viral M gene sequence analysis

目的调查云南省部分地区狂犬病病人和疫点犬群携带狂犬病病毒（rabies virus,RABV）状况及RABV膜基质蛋白（matrixprotein,M）基因序列分析,为狂犬病防控提供科学依据。方法 2008-2009年在云南省采集犬脑组织标本606份,狂犬病病人唾液8份,脑脊液1份,用直接免疫荧光试验检测RABV抗原,用RT-PCR检测RABV核酸,对阳性标本进行M基因序列测定和分析。结果所有标本经检测,RABV抗原和/或核酸阳性16份,其中疫点扑杀的貌似健康犬脑组织3.10%（10/323）,狂犬病病人唾液5份、脑脊液1份。狗肉餐馆屠宰犬的脑组织283份均为阴性。云南16株RABV的M基因核苷酸和氨基酸同源性分别为88.5%～100%和85.2%～99.5%。它们与中国人用疫苗株aG核苷酸和氨基酸同源性分别为83.9%～85.7%和82.3%～93.6%;与人用疫苗株CTN181核苷酸和氨基酸同源性分别为99%～99.7%和98.5%～99%。系统进化分析表明,云南16株RABV均属基因Ⅰ型并可分为进化Ⅰ和Ⅱ群并分别与泰国等东南亚国家和相邻省份流行株具有较近的亲缘关系。结论云南省狂犬病流行与周边省份和东南亚地区的狂犬病传播扩散有一定关系;狂犬病疫点部分貌似健康犬携带RABV并具有传染源意义;云南狂犬病病毒株M基因与我国人用狂犬病疫苗CTN株的同源性和亲缘关系较近,但与aG株同源性存在。

We conducted investigation on the rabies virus（RABV）infected from rabies patients and dog population in endemic sites,Yunnan Province,China and the viral matrixprotein（M）gene sequence analysis,providing references for control and prevention of rabies.A total of 606 brain tissue samples of dogs collected from endemic sites,as well as 8saliva and 1cerebrospinal fluid samples of rabies patients collected from Yunnan Province in 2008-2009.RABV antigen and nucleic acid were detected by direct immunofluorescence assay and RT-PCR respectively,and positive samples were sequenced for the M gene.According to the nucleotide and amino acid of target gene,the genetic homology and the phylogeny were analyzed.Above specimens tested,16 specimens were RABV antigen and/or nucleic acid positive.Among them,3.10%（10/323）were from apparently healthy dog brain specimens of endemic sites,5saliva and 1cerebrospinal fluid were from human rabies cases.In addition,283 dog brain specimens collected from dog meat restaurant were negative.The M gene sequence of the 16 strains have been acquired,the rate of their nucleotide homology and deduced amino acid homology were 88.5%-100% and 85.2%-99.5% respectively.Compared with aG and CTN181strains（vaccine strains for humans in China）nucleotide（deduced amino acid）homology were 83.9%-85.7%（82.3%-93.6%）and 99%-99.7%（98.5%-99%）respectively.The phylogenic analysis indicated that the 16 isolates from Yunnan Province divided into I and II phylogenic subgroups.These 2phylogenic subgroups of RABV from Yunnan Province have a closer genetic relationship with the street strains isolated from Southeast Asia and neighboring provinces of China.Rabies epidemic in Yunnan Province might be attributed to rabies spread from Southeast Asia and neighboring provinces.Some apparently healthy dogs can infected RABV.The M gene nucleotide and amino acid of Yunnan strains of rabies virus have a higher homology and a closer genetic relationship with CTN strain of human rabies vaccine used in China,but they with aG strains of human rabies vaccine used in China homologous exist obvious difference.