H5亚型禽流感重组鸡痘病毒活载体疫苗的构建及其遗传稳定性与免疫效力

以鹅源H5亚型禽流感病毒(AIV)基因组为模板，用RTPCR扩增血凝素（Hemagglutinin, HA）基因，克隆入鸡痘病毒表达载体pFG1175，转染鸡痘病毒感染的鸡胚成纤维细胞，通过蓝斑筛选和间接免疫荧光检测，获得表达HA基因的重组鸡痘病毒（Recombinant fowlpox virus, rFPVHA）。rFPVHA经鸡胚成纤维细胞连续传15代后，报告基因LacZ和HA基因可稳定表达。用103PFU和105PFU的rFPVHA免疫无特定病原体的(Specific pathogen free, SPF)鸡，免疫后22d 血凝抑制(Hemagglutinin inhibition,HI)抗体监测阳性率分别为0%和20%，但均抵御了H5亚型毒株的致死性攻击，保护率为100%。结果表明，构建了表达HA基因的重组鸡痘病毒，该重组病毒具有良好遗传稳定性，免疫鸡可提供完全保护，显示出了一定的应用前景。

Construction, Genetic Stability and Protective Efficacy of Recombinant Fowlpox Virus Expressing Hemagglutinin Gene of H5N1 Subtype Avian Influenza Virus

Construction, genetic stability and protective efficacy of a recombinant fowlpox virus expressing hemagglutinin(HA) gene (rFPV-HA) of H5 subtype avian influenza virus were conducted. The results indicated that the recombinant fowlpox virus was obtained and purified by blue plaque selection. HA expressed by rFPV-HA was identified by indirect immunofluorescence array. The rFPV-HA could express reporter gene LacZ and HA gene after fifteen successive passages in chicken embryo fibroblast. The rFPV-HA induced completely protection of chickens against lethal challenge with homologous avian influenza virus. The results show that rFPV-HA has good genetic stability and antigenicity and should be a useful tool in HPAI control program by preventing illness and death in chickens when the disease spreads widely.