鸽副黏病毒灭活疫苗候选株aYQ的生物学特性研究

摘要

评价鸽副黏病毒 (PPMV) 致弱毒株aYQ的生物学特性, 为研制鸽副黏病毒灭活疫苗奠定基础.将PPMV aYQ株经SPF鸡胚连续传代, 建立种子批.测定各代次毒种的血凝效价 (HA) 和鸡胚半数感染量 (EID50) , 并进行纯净性检验;测定毒种鸡胚最小致死量的平均死亡时间 (MDT) 、1日龄鸡脑内接种致病指数 (ICPI) 和6周龄鸡静脉接种致病指数 (IVPI) ;测定毒种对鸡胚和鸽的毒力、遗传稳定性和抗原相关性.PPMV aYQ株经SPF鸡胚传代至F10, HA效价为1:256～1:512, 病毒含量为10-8.5 EID50/0.1mL～10-8.9 EID50/0.1mL, 毒种各代次均无细菌、霉菌、支原体以及外源病毒污染;PPMV aYQ株的ICPI和IVPI结果均为0, MDT测定结果大于120h;以103倍稀释的PPMV aYQ株病毒液接种10日龄SPF鸡胚96h无死亡, 无眼观病变.以106EID50/羽的剂量人工感染鸽, 未见异常症状;PPMV aYQ株各代次F基因均未发生基因突变, F蛋白裂解位点氨基酸序列均为-112GRQGRL 117-;交叉血凝抑制试验和交叉中和试验结果表明, PPMV aYQ株与NDV La Sota株间存在明显的抗原性差异.鸽副黏病毒弱毒株aYQ能够在SPF鸡胚上稳定传代, 且生物学特性稳定, 免疫原性良好, 可以作为鸽源副黏病毒灭活疫苗的毒种.%The objective of this study was to evaluate the biological characteristics of the aYQ strain of pigeon paramyxovirus and to lay the foundation for the development of the pigeon-paramyxovirus inactivated vaccine.PPMV aYQ strain were passaged continuously in SPF chicken embryos to establish a seed batch.HA and EID50 were determined and the purity of each strain was tested;The virulence of the aYQ strain was evaluated by MDT, ICPI and IVPI.Pathogenicity of aYQ strain to chicken embryos and pigeons, genetic stability and antigen relevance were determined.PPMV aYQ strain was passaged to 10 generations in SPF chicken embryos in the absence of bacteria, mold, mycoplasma and exogenous viruses.The HA titers of various generation viruses were 1:256-1:512and the virus contents were 10-8.5-10-8.9 EID50/0.1mL.Both ICPI and IVPI of PPMV aYQ strain were 0, and the MDT was more than 120h;The 10-day-old SPF chicken embryos inoculated with 103-fold diluted PPMV aYQ strain survived at 96 hpost inoculation.Pigeons which were infected artificially at a dose of 106EID50 with PPMV aYQ strain showed no apparent clinical symptoms;The results of sequence analysis showed that there was no gene mutation in F gene of PPMV aYQ strain, and the deduced amino acid sequences were-112 GRQGRL 117-.The results of cross-hemagglutination inhibition test and cross-neutralization test showed that there was obvious antigenic difference between PPMV aYQ strain and NDV La Sota strain.The aYQ strain can be passaged in SPF chicken embryos with stable genetic and biological characteristics.It indicated that the PPMV aYQ strain possess satisfied immunogenicity, and can be used as the preferred pigeon paramyxovirus inactivated vaccine candidate strain.