家蚕生物反应器表达传染性法氏囊病病毒多聚蛋白的免疫原性研究

将传染性法氏囊病病毒(IBDV)浙江分离株JD1的多聚蛋白(VP2/4/3)基因克隆到家蚕杆状病毒转移载体pBacPAK8中,重组载体与杆状病毒Bm-BacPAK6的线性化基因组DNA共转染家蚕细胞后,将获得的重组病毒BacPAK-A感染家蚕5龄起幼虫进行虫体内表达.用感染后第5日的蚕血淋巴作抗原制备油佐剂苗免疫14日龄非免疫鸡,安全性试验表明,接种1倍和5倍免疫剂量的试验鸡在临床反应和剖检中均无异常变化;在免疫-攻毒试验中设杆状病毒表达的IBDV VP2蛋白和正常蚕血淋巴免疫对照组,并于28日龄加强免疫,25 d后用IBDV强毒BC6/85攻击.通过临床保护率、病理保护率及血清学试验表明,基于多聚蛋白制备的疫苗更成功地诱导了体液免疫应答,比VP2蛋白对IBDV强毒的攻击具有更高的保护力,更适于作为IBD基因工程亚单位疫苗的抗原成分.

Immunogenicity of infectious bursal disease virus (IBDV) polyprotein expressed in silkworm as the bioreactor

The polyprotein (VP2/4/3) gene of infectious bursal disease virus (JD1 strain) was cloned into BmNPV-based vector pBacPAK8. Recombinant baculovirus BacPAK-A harbouring polyprotein gene was obtained from BmN cell co-transfected with recombinant vector and lineared genomic DNA of baculobvirus Bm-BacPAK6. BacPAK-A was used to infect fifth-instar silkworm larvae for expression of recombinant protein and haemolymph of the silkworm harvested 5-day post inoculation was emulsionized with mineral oil for immunization of 14-day old chickens. The safety trial indicated that 1-time and 5-time dose of vaccine are both safe to chickens. Groups inoculated with silkworm-expressed (IBDV) VP2 and normal silkworm haemolymph, respectively, were set for comparison in immunity-challenge trial. A boost was administered 14 days post primary immunization and a challenge with virulent (IBDV) (BC6/85) 25 days post the boost. Clinical and pathological protection ratio, ratio of bursa weight vs body weight, immunoprecipitation and neutrlization test were analysed. The result indicated that the polyprotein vaccine induced humoral immunity more effectively than that of the VP2 vaccine, as the former provided a more efficient protection against virulent (IBDV) challenge. The recombinant polyprotein could be a promising candidate for genetic engineering subunit vaccine against IBD.