贵州省一例犬伤致人患狂犬病的病原学及病毒基因分析

Etiologic and viral gene analysis on a case of human rabies caused by the wounding of a dog inGuizhou province

目的从病原学角度证实贵州省凯里市一起犬伤人致儿童死亡病例为狂犬病所致，了解狂犬病病毒的基因特征。方法采用dFA实验初步检测犬和患儿脑组织狂犬病病毒抗原，以RT-nestedPCR检测狂犬病病毒核酸，测定狂犬病病毒N基因全长序列，根据同源性比较及系统进化树进行分子流行病学分析。结果dFA与RT-nestedPCR检测显示犬脑和人脑组织标本均为狂犬病病毒抗原和核酸阳性。经测序拼接均得到长度为1353bp的核苷酸序列，同源性分析显示犬脑组织（GZD）和人脑组织（GZH）检出的狂犬病病毒N基因核苷酸与推导的氨基酸同源性均为100％，与我国各省已报道的狂犬病病毒基因1型流行毒株及疫苗株核苷酸和氨基酸序列同源性分别为88．4％～99．6％和98．2％-100％，与我省往年报道的毒株N基因核苷酸和推到的氨基酸同源性最高。此外，在与疫苗株的比较中，与CNT株的核苷酸和氨基酸同源性最高。进化树分析显示犬脑和人脑组织标本狂犬病病毒N基因亲缘进化关很近，同属于基因1型狂犬病病毒。结论从病原学和病毒分子生物学证实了贵州省一起犬伤人致儿童死亡病例为狂犬病所致，其病原为狂犬病病毒基因1型，与疫苗毒株中的CNT毒株的亲缘进化关系最近，该起病例可能为我省境内传播，因此，应加强我省狂犬病疫区的防制工作。

Objective To confirm the death of a child injured by a dog was due to rabies and to understand the molecular biologic features of rabies virus in Kaili, Guizhou province. Methods Brain tissue samples of patient and dog were collected to detect the rabies virus by direct immunofluorescence assay （DFA） and RT-nested PCR assay. Homology and phylogenetic tree were analyzed based on the whole nueleotide and deduced amino acid sequence of N gene of rabies virus followed by molecular epidemiological analysis. Results Both the human and dog brain tissue samples were confirmed positive by DFA and RT-nested PCR assay. The homology analysis of N gene sequences among GZH, GZD and other epidemic and vaccine rabies strains isolated from other provinces and other countries indicated that the detected samples shared the highest homology with the strain detected in Anlong prefecture in Guizhou in the year of 2006, and the homology between GZH and GZD was as high as 100%. Besides, among the vaccine strains, GZH and GZD showed the highest homology with strain CNT. Phylogenetic analysis indicated that the two samples were very close and belonged to genetype 1 lyssavirus, with the closest relationship between samples reported in Guizhou and Beijing. Conclusion It was confirmed on the viral molecular level that both the human and dog in Kaili were suffered from rabies, and the pathogens were genetype 1 lyssavirus. The prevalent strains in Kaili city was probably imported from other prefectures of Guizhou province, suggesting that prevention and control measures on rabies in Guizhou province should be strengthened.