| Pseudorabies, hog cholera and Foot-and-mouth disease, classified as World Organization for Animal Health (OIE) List A disease, are all devastating disease of livestock, lead to the financial losses to animal husbandry. Pseudorabies and Foot-and-mouth disease are also infectious to human beings and follow with extensive interest as zoonosises. Although the conventional vaccines made by chemically inactivated whole viral particle have been efficiently controlled such diseases in many countries, it was found to cause outbreaks by release of incompletely inactivated virus from the vaccine. DNA vaccine is a new generation of vaccine beginning in the 90's, which has shows powerful potency in the prevention and therapy of many diseases. Based on initial achievement in DNA vaccines against the diseases, strategy for constructing a tricistronic DNA vaccine against pseudorabies,hog cholera and Foot-and-mouth disease simultaneously is investigated in our study.1. Three eukaryotic expression plasmids containing cDNA encoding pseudorabies virus (PRV) gC antigen, Classical Swine Fever Virus (CSFV) E2 antigen, gC, E2 and foot-and-mouth disease virus (FMDV ) VP1 antigens simultaneously were obtained by DNA recombination. The 3 plasmids were designated as: (a) pVGC containing sequence from the gC gene including the heparin binding domain (HBD) insertion with signal sequence of tissue plasminogen activator (tPA); (b) pVE2 containing the E2 gene insertion with the signal, but without transmembrane sequence and (c) pVGEV containing gC, E2 and VP1 gene combined the picornavirus FMDV 2A sequence and the internal ribosome entry site (IRES) from encephalomyocarditis virus (ECMV). The pVGEV plasmid was transfected into COS7 cells in vitro by electroporation. The expression of three antigens was detected by PR-PCR.2. We measured the ability of three DNA vaccine plasmids, pVGC, pVE2 and PF-TPA, to induce viral specific IgG measured in sera samples using an enzyme-linked immunosorbent assay (ELISA) and IFN-r secreted by the separated splenocytes from mice was assayed using the commercial ELISA kits when delivered alone or in a mixture containing all nine plasmids. We further examined the possible effect of individual plasmids, by assessing a series of mixtures in which each of the three vaccine plasmids was replaced with a control plasmid. Given alone, each of the vaccine plasmids induced significant antibody titers and IFN-r responses. Significant suppression response was seen when the plasmid PF-TPA was pooled in a three-plasmid cocktail and injected in a single site. Removal of single genes from the mixture frequently reduced the observed suppression. Boosting with VP1 antigen protein overcome the suppressive effect of mixing for the antibody response in animals primed with the mixture, but, even after boosting, IFN-r response was higher in animals primed with single plasmids than in those primed with the three-plasmid mixture. Interactions between components in a multiplasmid DNA vaccine may limit the ability to use plasmid pools alone to induce responses against multiple targets... |
