表达H9亚型禽流感病毒血凝素基因的重组鸡痘病毒及其免疫效力

以RTPCR法扩增获得H9亚型禽流感病毒(AIV)分离株(A/Chicken/China/F/1998)的血凝素(HA)基因，将其定向插入鸡痘病毒转移载体1175的痘苗病毒启动子P75的下游，得到重组转移载体1175HA。以脂质体转染法将1175HA转染至已感染鸡痘病毒282E4疫苗株(wtFPV)的鸡胚成纤维细胞(CEF)中，通过在含Xgal的营养琼脂上连续挑选蓝色病毒蚀斑获得并纯化rFPVHA。以间接免疫荧光法证实感染rFPVHA的CEF表达了HA。rFPVHA在免疫7日龄SPF鸡7天后即能诱生可检出的血凝抑制(HI)抗体，14天后诱生的HI抗体到达高峰，且诱生的HI抗体保持较高水平达55天。在7日龄SPF鸡及含抗FPV母源抗体的商品鸡上进行的免疫效力试验表明，rFPVＨＶ能显著抑制静脉攻毒后免疫鸡从泄殖腔的排毒，效果与AIV全病毒灭活苗相当。

Recombinant Fowlpox Virus Expressing HA From Subtype H9N2 of Avian Influenza Virus and Its Protective Immunity Against Homologous Challenge in Chickens

The hemagglutinin (HA) gene from the AIV,A/Chicken/China/F/1998(H9N2),was amplified with the RT PCR technique and directionally inserted into transferring vector 1175,resulted in recombinant transferring vector 1175HA.In order to generate recombinant fowlpox virus expressing HA(rFPV HA),the recombinant transferring vector 1175HA was used to transfect the chicken embryo fibroblasts(CEF)pre infected with wide type fowlpox virus.Then,by selection of blue plaques on the CEF overlaid with agar containing X gal,rFPV HA was obtained and purified.The expression of HA by rFPV HA was detected in the recombinant virus infected CEF by indirect immunofluorescence.Experiments on chickens demonstrated that rFPV HA could induce detectable HI antibodies 7 days post vaccination and those HI antibodies of relatively high titers could persist 55 days.rFPV HA also had the same protective efficacies to suppress SPF chickens or commercial broiler chickens with antibodies against FPV from shedding challenged virus from intestine as inactivated vaccine in oil emulsion.