| Summary: | 博士 === 國立中興大學 === 微生物暨公共衛生學研究所 === 105 === Both pseudorabies virus (PRV) and porcine circovirus type 2 (PCV2) are economically important swine pathogens. The only structural protein of viral capsid, Cap has become the major target for development of PCV2 subunit vaccines. The gE gene or tk gene deficient PRV maker vaccines are wildly used in pseudorabies (PR) eradication program. To address the potential for bivalent vaccines applications, the recombinant PRVs defective in the glycoprotein E (gE) or thymidine kinase (tk) gene and containing the PCV2 cap gene were constructed. The recombinant gE-/PCV2cap+ PRV and gE-tk-/PCV2cap+ PRV were generated by homologous recombination between the transfer plasmid and the parental viral DNA. Both the recombinant viruses were shown to express the PRV viral protein and PCV2 Cap protein. By transmission electron microscopy (TEM) and immunogold labelling analysis, the recombinant PCV2 Cap could self-assemble into virus-like particles (VLPs), which are morphologically- and antigenically similar to the original PCV2 virions. In addition, the viral replication titers of gE-/PCV2cap+ PRV and gE-tk-/PCV2cap+ PRV were reached 108 and 107 pfu/ml, respectively. To further investigate the immunogenicity of the viruses, the recombinant PRVs and the recombinant PCV2 VLPs were purified and administrated as vaccines to mice and guinea pigs. Mice and guinea pigs immunized with PCV2 VLPs purified from gE-/PCV2cap+ PRV-infected cells could mount specific antibody responses to PCV2 Cap. PCV2 neutralizing antibody titers could be detected around 1:8~1:16 at 2 weeks after boost immunization in the immunized guinea pigs. The virulence of gE-tk-/PCV2cap+ PRV was determined in mouse and guinea pigs, and the results indicated complete attenuation of gE-tk-/PCV2cap+ PRV, suggesting its potential use as an live attenuated vaccine. Furthermore, both the recombinant PRVs could stimulate the protective PRV neutralizing antibodies in immunized guinea pigs. Taken together, the recombinant PRVs not only retained the immunogenicity of PRV but also a new tool for manufacturing PCV2 VLPs.
|
|---|
