宁夏地区人畜BDVp24基因系统发生树构建和序列分析

Construction of phylogenetic tree and sequence analysis on BDV p24 gene in both livestock and host in Ningxia

目的了解宁夏及其周边地区博尔纳病病毒（BDV）的感染状况和基因特征及其种系发生来源。方法采用巢式反转录实时荧光定量聚合酶链反应（FQ—nRT-PCR），检测119例病毒性脑炎（VE）患者和其密切接触的患病或健康牛205头、绵羊978只及脑血管病患者46例、多发硬化患者13例的外周血单核细胞BDVp24片段，检出的阳性序列连同前期检出的宁夏绵羊、VE患者和抑郁症患者阳性BDVp24基因序列与GenBank中5个国家7个动物种属29例BDVp24基因序列进行比对，分析其核苷酸和氨基酸序列的同源性，重新构建基因系统发生树。结果23例阳性检测标本连同3例前期检测序列基因聚合分析显示核苷酸之间的同源相似度为96．5％-100．0％，氨基酸的同源相似度为95_3％～100．O％。与德国马源性标准株HE80同源相似度较高。构建基因的系统发生树发现宁夏绵羊、VE患者和抑郁症患者同德国马和绵羊、奥地利马都各自形成了独立的支系，其余样本形成混合支系，其中宁夏的vE患者、牛、绵羊同德国马、日本绵羊形成德国-中国宁夏-日本混合支系。结论宁夏及其周边地区人和动物BDV的感染，存在区域源性BDV独立株和国外传人基因保守株的多源状态。

Objective To gain detailed insights into the Boma disease virus infection and its genetic characteristics and phylogeny in Ningxia, China. Methods BDV p24 segment were detected by fluorescence quantitative nested RT-PCR from peripheral blood mononuclear cells of 119 patients with viral encephalitis, 205 cattle, 978 sheep, 46 patients with cerebrovascular diseases and 13 patients with multiple sclerosis. Data from phylogenetic analysis on BDV p24 positive samples together with those from the positive gene sequences of animals and patients in the previous studies in Ningxia, were compared to the 29 sequences provided by the GenBank from 7 animals in 5 countries. Both the sequence homologous similarity of the nucleotide and amino acid sequences were analyzed and gene phylogenetic tree was reconstructed. Results Data from 23 collections of positive test samples, together with the ones from early detection of gene sequence analysis, showed that the homologous similarity sequences of both nucleotide and amino acid was between 95.3% and 100.0% and highly homophylic with HE80 that detected from ill horses in Germany. One part of nucleotide sequences formed the ＇Ningxia independent branch＇ while the other one belonged to the ＇Germany - Ningxia - Japan mixed branch＇. There was a high identity within the branch. Conclusion A Ningxia independent BDV strain from geographical origin might exist while the epidemic strains were imported with multiple sources.