猪流行性腹泻病毒重组S1蛋白的免疫原性评价

Immunogenicity of recombinant S1 of porcine epidemic diarrhea virus

将猪流行性腹泻病毒(PEDV)流行毒株S1基因经密码子优化后克隆到慢病毒表达载体,在293T细胞包装PEDV S1重组慢病毒;重组慢病毒感染293T细胞,经有限稀释法筛选获得高效稳定表达S1蛋白的重组细胞系HEK-293T-S1;生产、纯化S1蛋白,添加佐剂制备成重组S1亚单位疫苗,经肌肉注射免疫实验用巴马小型猪,评价重组S1蛋白的免疫原性。结果表明,疫苗经肌肉注射免疫能诱导唾液、粪便及血清产生S1特异性Ig A;证实巴马小型猪3日龄仔猪对PEDV流行毒株易感,在临床症状及组织病理学变化均符合高毒力PEDV特征,可以作为感染动物模型;在实验用巴马小型猪证实PEDV重组S1蛋白结合佐剂经肌肉注射可诱导粘膜免疫,这为PEDV基因工程疫苗研发奠定了基础。

S1 gene of porcine epidemic diarrhea virus(PEDV)was codon-optimized and cloned into a lentiviral expression vector,and the recombinant lentivirus that carried PEDV S1 gene was packaged and transduced into 293 T cells.A recombinant cell line HEK-293 T-S1 with high and stable expression of S1 protein was obtained by limited dilution and identified by western-blot analysis.The recombinant S1 protein was produced and purified,and its immunogenicity was evaluated in weaned Bama miniature pigs.The results showed that the recombinant S1 protein,combined with adjuvants,induced S1-specific Ig A in saliva,feces and sera from the pigs immunized intramuscularly with this vaccine,and 3-days aged Bama piglet was susceptible to a PEDV wild strain and was suitable as an animal model of PEDV infection.This study demonstrated that a PEDV recombinant S1 protein binding adjuvant administered intramuscularly could induce mucosal immunity of Bama miniature pigs,which set the foundation for PEDV genetically engineered vaccine development.