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Induction of robust immunity response in mice by dual-expression-system-based recombinant baculovirus expressing the capsid protein of porcine circovirus type 2.

Ye Y, Cheng X, Zhang J, Tong T, Lin W, Liao M, Fan H - Virol. J. (2013)

Bottom Line: Porcine circovirus type 2 (PCV2) is associated with post-weaning multisystemic wasting syndrome (PMWS), an emerging swine disease that causes progressive weight loss, dyspnea, tachypnea, anemia, jaundice, and diarrhea in piglets.In this study, we successfully constructed a dual-expression-system-based recombinant baculovirus BV-GD-ORF2, which can display the PCV2 capsid (Cap) protein and VSV-G protein on the viral envelope and also expressing Cap protein on transduced mammalian cells, thereby functioning as both a subunit and a DNA vaccine.The vaccination of mice with recombinant baculovirus BV-GD-ORF2 successfully induced robust Cap-protein-specific humoral and cellular immune responses.

View Article: PubMed Central - HTML - PubMed

Affiliation: Key Laboratory of Animal Vaccine Development, Ministry of Agriculture, Guangzhou 510642, China. fanhy@scau.edu.cn.

ABSTRACT

Background: Porcine circovirus type 2 (PCV2) is associated with post-weaning multisystemic wasting syndrome (PMWS), an emerging swine disease that causes progressive weight loss, dyspnea, tachypnea, anemia, jaundice, and diarrhea in piglets. Although baculovirus is an enveloped virus that infects insects in nature, it has emerged as a vaccine vector, and we used it to develop a novel candidate vaccine for a preventive or therapeutic strategy to control PCV2 infections.

Methods: Immunoblotting analysis of recombinant baculovirus and immunofluorescent staining of baculovirus-infected cells were followed using anti-ORF2 monoclonal antibodies. The BALB/c mice were immunized intramuscularly with this baculovirus. The titers of antibodies were mensurated with a Cap-protein-specific enzyme-linked immunosorbent assay (ELISA) and a serum neutralization assay. The IFN-γ response in splenocytes harvested from immunized mice was measured by ELISA. Student's t-test was used to compare immune responses of different groups.

Results: In this study, we successfully constructed a dual-expression-system-based recombinant baculovirus BV-GD-ORF2, which can display the PCV2 capsid (Cap) protein and VSV-G protein on the viral envelope and also expressing Cap protein on transduced mammalian cells, thereby functioning as both a subunit and a DNA vaccine. After infection, the Cap protein was expressed and displayed on the viral surface, as demonstrated with an indirect fluorescence assay and immunoblotting. The vaccination of mice with recombinant baculovirus BV-GD-ORF2 successfully induced robust Cap-protein-specific humoral and cellular immune responses.

Conclusions: Our findings collectively demonstrate that the recombinant baculovirus BV-GD-ORF2 is a potential vaccine against PCV2 infections.

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Related in: MedlinePlus

Immunoblotting analysis of recombinant AcMNPV particles using anti-ORF2 monoclonal antibodies. Sf9 cells infected with BV-GD-ORF2 (lanes 1); Sf9cells infected with AcMNPV-WT (lanes 2); complete BV-GD-ORF2 virions (lanes 3); and purified nucleocapsids from BV-GD-ORF2 virions treated with Triton X-100 (lanes 4). All were treated with lysis buffer and subjected toimmunoblotting. Expression of the ORF2–gp64fusion protein was examined.
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Figure 3: Immunoblotting analysis of recombinant AcMNPV particles using anti-ORF2 monoclonal antibodies. Sf9 cells infected with BV-GD-ORF2 (lanes 1); Sf9cells infected with AcMNPV-WT (lanes 2); complete BV-GD-ORF2 virions (lanes 3); and purified nucleocapsids from BV-GD-ORF2 virions treated with Triton X-100 (lanes 4). All were treated with lysis buffer and subjected toimmunoblotting. Expression of the ORF2–gp64fusion protein was examined.

Mentions: To investigate whether the Cap protein was displayed on the membrane of BV-GD-ORF2, the purified viral particles were analyzed with immunoblotting with an anti-ORF2 monoclonal antibody. A protein of an approximately 75 kDa protein (of the predicted size) was presented in the lanes loaded with the recombinant baculovirus BV-GD-ORF2 or with BV-GD-ORF2-infected Sf9 cells, but not in lanes containing pellets of viral nucleocapsids only or in those lanes loaded with AcMNPV wild-type (AcMNPV-WT) infected Sf9 cells (Figure 3). Our observations indicated that the presence of both VSV-G and PCV2 Cap protein does not significantly reduce the displaying efficiency of Cap protein on baculovirus membrane.


Induction of robust immunity response in mice by dual-expression-system-based recombinant baculovirus expressing the capsid protein of porcine circovirus type 2.

Ye Y, Cheng X, Zhang J, Tong T, Lin W, Liao M, Fan H - Virol. J. (2013)

Immunoblotting analysis of recombinant AcMNPV particles using anti-ORF2 monoclonal antibodies. Sf9 cells infected with BV-GD-ORF2 (lanes 1); Sf9cells infected with AcMNPV-WT (lanes 2); complete BV-GD-ORF2 virions (lanes 3); and purified nucleocapsids from BV-GD-ORF2 virions treated with Triton X-100 (lanes 4). All were treated with lysis buffer and subjected toimmunoblotting. Expression of the ORF2–gp64fusion protein was examined.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231451&req=5

Figure 3: Immunoblotting analysis of recombinant AcMNPV particles using anti-ORF2 monoclonal antibodies. Sf9 cells infected with BV-GD-ORF2 (lanes 1); Sf9cells infected with AcMNPV-WT (lanes 2); complete BV-GD-ORF2 virions (lanes 3); and purified nucleocapsids from BV-GD-ORF2 virions treated with Triton X-100 (lanes 4). All were treated with lysis buffer and subjected toimmunoblotting. Expression of the ORF2–gp64fusion protein was examined.
Mentions: To investigate whether the Cap protein was displayed on the membrane of BV-GD-ORF2, the purified viral particles were analyzed with immunoblotting with an anti-ORF2 monoclonal antibody. A protein of an approximately 75 kDa protein (of the predicted size) was presented in the lanes loaded with the recombinant baculovirus BV-GD-ORF2 or with BV-GD-ORF2-infected Sf9 cells, but not in lanes containing pellets of viral nucleocapsids only or in those lanes loaded with AcMNPV wild-type (AcMNPV-WT) infected Sf9 cells (Figure 3). Our observations indicated that the presence of both VSV-G and PCV2 Cap protein does not significantly reduce the displaying efficiency of Cap protein on baculovirus membrane.

Bottom Line: Porcine circovirus type 2 (PCV2) is associated with post-weaning multisystemic wasting syndrome (PMWS), an emerging swine disease that causes progressive weight loss, dyspnea, tachypnea, anemia, jaundice, and diarrhea in piglets.In this study, we successfully constructed a dual-expression-system-based recombinant baculovirus BV-GD-ORF2, which can display the PCV2 capsid (Cap) protein and VSV-G protein on the viral envelope and also expressing Cap protein on transduced mammalian cells, thereby functioning as both a subunit and a DNA vaccine.The vaccination of mice with recombinant baculovirus BV-GD-ORF2 successfully induced robust Cap-protein-specific humoral and cellular immune responses.

View Article: PubMed Central - HTML - PubMed

Affiliation: Key Laboratory of Animal Vaccine Development, Ministry of Agriculture, Guangzhou 510642, China. fanhy@scau.edu.cn.

ABSTRACT

Background: Porcine circovirus type 2 (PCV2) is associated with post-weaning multisystemic wasting syndrome (PMWS), an emerging swine disease that causes progressive weight loss, dyspnea, tachypnea, anemia, jaundice, and diarrhea in piglets. Although baculovirus is an enveloped virus that infects insects in nature, it has emerged as a vaccine vector, and we used it to develop a novel candidate vaccine for a preventive or therapeutic strategy to control PCV2 infections.

Methods: Immunoblotting analysis of recombinant baculovirus and immunofluorescent staining of baculovirus-infected cells were followed using anti-ORF2 monoclonal antibodies. The BALB/c mice were immunized intramuscularly with this baculovirus. The titers of antibodies were mensurated with a Cap-protein-specific enzyme-linked immunosorbent assay (ELISA) and a serum neutralization assay. The IFN-γ response in splenocytes harvested from immunized mice was measured by ELISA. Student's t-test was used to compare immune responses of different groups.

Results: In this study, we successfully constructed a dual-expression-system-based recombinant baculovirus BV-GD-ORF2, which can display the PCV2 capsid (Cap) protein and VSV-G protein on the viral envelope and also expressing Cap protein on transduced mammalian cells, thereby functioning as both a subunit and a DNA vaccine. After infection, the Cap protein was expressed and displayed on the viral surface, as demonstrated with an indirect fluorescence assay and immunoblotting. The vaccination of mice with recombinant baculovirus BV-GD-ORF2 successfully induced robust Cap-protein-specific humoral and cellular immune responses.

Conclusions: Our findings collectively demonstrate that the recombinant baculovirus BV-GD-ORF2 is a potential vaccine against PCV2 infections.

Show MeSH
Related in: MedlinePlus