摘要
利用SPF鸡胚从吉林省某肉鸽养殖场送检的疑似新城疫病毒(Newcastle disease virus,NDV)感染肉鸽体内分离获得1株病毒,通过电镜负染观察、血清学检查、RT-PCR检测以及病理组织学观察等方法对该株病毒进行系统的鉴定,电镜负染观察可见呈圆形、有囊膜的病毒粒子,且在囊膜表面有呈放射状排列的纤突;血清学检查结果显示,该病毒株可凝集鸡的红细胞,具有血凝性;RT-PCR扩增获得与预期目的基因片段大小一致的条带;病理组织学观察可见脑血管及神经胶质细胞周围间隙增大,呈典型的脑水肿变化。综合分析以上结果,证实该分离株为NDV,命名为NDV-cc-12。参照GenBank中NDV F基因的核苷酸序列,设计合成1对特异性引物,利用RT-PCR成功扩增获得NDV-cc-12分离株F基因并对其进行序列测定。序列分析结果显示,获得的NDV-cc-12分离株F基因片段长为374bp,其裂解位点的氨基酸序列为112RRQKRF 117,符合NDV强毒株裂解位点序列特征。F基因的系统发育进化树结果表明,NDV-cc-12分离株与HLJ4-95毒株的亲缘关系较近,处于同一基因亚型上,属于基因Ⅵ型。
A Newcastle disease virus isolate,named as NDV-cc-12,was obtained from the pigeon in Jilin province by using SPF chick embryos.The virus was identified as NDV based on the results of morphology observation,serologic detection,histopathological observation and RT-PCR method.The round virions with peploses were observed by negative stained,which had spikes on the surface of peploses.The detection result of serology indicated that the isolate had hemagglutination,which could agglutinate the red blood cells of chicken.The fragment obtained by RT-PCR amplification was in accordance with the size of the objective gene.The result of histopathological observation indicated that the interspace around cerebral vessels and gliacytes enlarged,which presented the typical lesion of brain edema.The F gene was amplified by the specific primers,which were designed based on nucleotide sequence of F gene of NDV published in GenBank.The F gene was sequenced and analyzed.The length of F gene was 374 bp,and the amino acid sequence of F gene at the cleavage site shared a velogenic motif 112 RRQKRF 117 with other virulent isolates.The results of the phylogenetic tree analysis indicated that there was a close genetic relationship between NDV-cc-12 isolate and HLJ4-95strain,which located in the same subtype and belonged to genotypeⅥ.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2013年第9期1333-1338,共6页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(31072106)
