表达O型口蹄疫病毒VP1中和表位的重组猪圆环病毒2型毒株的构建及其生物学特性

为构建以猪圆环病毒2型(PCV2)为载体,表达O型口蹄疫病毒(FMDV)中和表位的重组病毒,将FMDV-VP1蛋白中和抗原表位(第141～160位氨基酸)插入PCV2-ORF2编码的Cap蛋白C-末端,利用感染性克隆技术拯救出1株表达FMDV-VP1的中和抗原表位PCV2重组毒株(命名为recPCV2-CL-VP1);采用免疫过氧化物酶单层试验、捕获ELISA和免疫电镜技术鉴定该重组病毒。结果表明,在重组病毒感染细胞中检出PCV2特异性抗原及VP1表位抗原;重组病毒的形态学特征与亲本病毒相似,可采用PCR和ELISA法相鉴别;拯救毒株的繁殖能力较亲本毒差,经细胞连续传10代,体外培养增殖性能稳定。证实成功构建了1株表达FMDV-VP1中和抗原表位的PCV2重组病毒,为进一步研制基因工程疫苗奠定了基础。

Construction and biological characterization of recombinant porcine circovirus type 2 expressing VP1 neutralization epitope of type O foot-and-mouth disease virus

In order to construct a recombinant porcine circovirus type 2(PCV2) expressing neutralization epitope of type O foot-and-mouth disease virus(FMDV),the neutralization epitope region(141-160 aa) of FMDV VP1 protein was inserted into the C-terminus of the Cap protein encoded by PCV2-ORF2.The cloned virus strain termed as recPCV2-CL-VP1 was rescued by transfecting an infectious clone into PK-15 cells.The recombinant virus recPCV2-CL-VP1 was detected by immunoperoxidase monolayer assay(IPMA),capture enzyme linked immunosorbent assay(ELISA) and immuno-electron microscope.Antigen specific to PCV2 and the VP1 epitope was detected in the recombinant virus by capture ELISA,and there was no detectable difference in the antigenicity of the recombinant virus compared with the parental virus of PCV2-CL strain by IPMA.However,the recombinant virus recPCV2-CL-VP1 can be differentiated from the parental virus by PCR and capture ELISA.The recombinant virus was similar to parental virus in morphological features,and the virus propagation was lower than that of the parental virus.The cloned virus could propagate stably in PK-15 cells for ten passages.In conclusion,the recombinant recPCV2-CL-VP1strain expressing FMDV-VP1 neutralization epitope was successfully established,providing foundation for the further development of engineering vaccine against both PCV2 and FMDV.