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Immunogenicity of Empty Capsids of Porcine Circovius Type 2 Produced in Insect Cells


Porcine circovirus type 2 (PCV2), a single-stranded DNA virus, is associated with postweaning multisystemic wasting syndrome (PMWS). ORF2 protein (capsid) of PCV2 was recently demonstrated to be a major immunogenable to induce protection in pigs with a prime–boost protocol. In this study, the ORF2 gene of PCV2 was expressed in insect cells. The product self-assembled into particles that were structurally and antigenically indistinguishable from regular PCV2 capsids. To evaluated the immunogenicity of these virus-like particles, PCV2-free piglets were vaccinated with the crude lysate from recombinant baculovirus (Ac.ORF2)-infected insect cells, at doses of 0.1 ml (106 cells), 0.5 ml (5 × 106 cells) or 1.0 ml (107 cells). The immune response was monitored by an indirect enzyme-linked immunosorbent assay (ELISA) for PCV2 antibody and lymphocyte proliferation assay. The ELISA results indicated that primary immune response was elicited with 0.5 ml or 1.0 ml of crude lysate from Ac.ORF2. After boost immunization, relatively higher levels of PCV2 antibody were elicited in 0.5-ml or 1.0-ml vaccinated groups, compared to the 0.1-ml group. In addition, higher PCV2 specific lymphocyte proliferation response was developed in piglets vaccinated with 0.5 ml or 1.0 ml of crude lysate, especially in those vaccinated with with 1.0 ml of crude lysate. Thus, the expressed ORF2 protein has significant potential as a subunit vaccine against PCV2 infection.

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Autographa california nuclear ployhedrosis virus


recombinant AcNPV expressing ORF2 protein


cytotoxic lymphocyte


enzyme-linked immunosorbent assay


horseradish peroxidase


major histocompatibility complex


peripheral blood mononuclear cell


polymerase chain reaction


porcine circovirus


plaque-forming units


post infection


post-weaning multisystemic wasting syndrome


phosphotungstic acid


Spodoptera frugiperda


stimulation index


virus-like particle

wt. AcNPV:

wild-type Autographa california nuclear polyhedrosis virus


  1. Allan, G.M., Meehan, B.M., Todd, D., Kennedy, S., McNeilly, F., Ellis, J., Clark, E.G., Harding, J., Espuna, E., Botner, A. and Charreyre, C., 1998a. Novel porcine circoviruses from pigs with wasting disease syndromes. Veterinary Record, 142, 467–468

  2. Allan, G.M., McNeilly, F., Kennedy, S., Daft, B., Clark, E.G., Ellis, J.A., Haines, D.M., Meehan, B.M. and Adair, B.M., 1998b. Isolation of porcine circovirus-like viruses from pigs with a wasting disease in the USA and Europe. Journal of Veterinary Diagnostic Investigation, 10, 3–10

  3. Allan, G.M. and Ellis, J.A., 2000. Porcine circovirus: a review. Journal of Veterinary Diagnostic Investigation, 12, 3–14

  4. Allan, G.M., McNeilly, F., Ellis, J., Krakowka, S., Meehan, B., McNair, I. and Walker, I., Kennedy, S., 2000. Experimental infection of colostrum-deprived piglets with porcine circovirus 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) potentiates PCV2 replication. Archives of Virology, 145, 2421–2429

  5. Bevan, M.J., 1976a. Cross-priming for a secondary cytotoxic response to minor H antigens with H-2 congenic cells which do not cross-react in the cytotoxic assay. Journal of Experimental Medicine, 143(5), 1283–1288

  6. Bevan, M.J., 1976b. Minor H antigens introduced on H-2 different stimulating cells cross-react at the cytotoxic T-cell level during in vivo priming. Journal of Immunology, 117(6), 2233–2238

  7. Blanchard, P., Mahe, D., Cariolet, R., Keranflec'h, A., Baudouard, M.A., Cordioli, P., Albina, E. and Jestin, A., 2003. Protection of swine against post-weaning multisystemic wasting syndrome (PMWS) by porcine circovirus type 2 (PCV2) proteins. Vaccine, 21, 4565–4575

  8. Boisgérault, F., Moron, G. and Leclerc, C., 2002. Virus-like particles: a new family of delivery systems. Expert Review of Vaccines, 1, 101–109

  9. Clark, E.G., 1997. Post-weaning multisystemic wasting syndrome. In: Proceedings of the American Associations of Swine Practitioners, 499–501

  10. De Arriba, M.L., Carvajal, A., Pozo, J. and Rubio, P., 2002. Isotype-specific antibody-secreting cells in systemic and mucosal associated lymphoid tissues and antibody responses in serum of conventional pigs inoculated with PEDV. Veterinary Immunology and Immunopathology, 84, 1–16

  11. Ellis, J., Hassard, L., Clark, E., Harding, J., Allan, G., Willson, P., Strokappe, J., Martin, K., McNeilly, F., Meehan, B., Todd, D. and Haines, D., 1998. Isolation of circovirus from lesions of pigs with postweaning multisystemic wasting syndrome. Canadian Veterinary Journal, 39, 44–51

  12. Ellis, J.A., Bratanich, A., Clark, E.G., Allan, G., Meehan, B., Haines, D.M., Harding, J., Werb, K.H., Krakowka, S, Konoby, C., and Hassand, L., 2000. Coinfection by porcine circoviruses and porcine parvovirus in pigs with naturally acquired postweaning multisystemic wasting syndrome. Journal of Veterinary Diagnostic Investigation, 12, 21–27

  13. Fenaux M., Oprissing, T., Halbur, P.G. and Meng, X.J., 2003. Immunogenicity and pathogenicity of chimeric infectious DNA clones of pathogenic porcine circovirus type 2 (PCV2) and nonpathogenic PCV1 in weaning pigs. Journal of Virology, 77(2), 11232–11243

  14. Fenaux, M., Oprissing, T., Halbur, P.G., Elvinger, F. and Meng, X.J., 2004. A chimeric procine circovirus (PCV2) with the immunogenic capsid gene of the pathogenic PCV2 type 2 (PCV2) cloned into the genomic backbone of the nonpathogenic PCV1 induces protective immunity against PCV2 infection in pigs. Journal of Virology, 78(12), 6297–6303

  15. Hamel, A.L., Lin, L.L. and Nayar, G.P., 1998. Nucleotide sequence of porcine circovirus associated with postweaning multisystemic wasting syndrome in pigs. Journal of Virology, 72: 5262–5267

  16. Heermann, K.H. and Gerlich, W.H., 1991. In: A. McLachlan (ed.), Molecular Biology of Hepatitis B Virus, (CRC Press, Boca Raton, FL), 109

  17. Kennedy, S., Moffett, D., McNeilly, F., Meehan, B., Ellis, J., Krakowka, S. and Allan, G.M., 2000. Reproduction of lesion of postweaning multisystemic wasting syndrome by infection of conventional pigs with porcine circovirus type 2 alone or in combination with porcine parvovirus. Journal of Comparative Pathology, 122, 9–24

  18. Kim, Y., Kim, J., Kang, K. and Lyoo, Y.S., 2002. Characterization of the recombinant proteins of porcine circovirus type 2 field isolate expressed in the baculovirus system. Journal of Veterinary Science, 3(1), 19–23

  19. Koch, G., van Roozelaar, D.J., Verschueren, C.A., Van der Eb, A.J. and Noteborn, M.H., 1995. Immunogenic and protective properties of chicken anemia virus proteins expressed by baculovirus. Vaccine, 13, 763–770

  20. Krakowka, S., Ellis, J.A., McNeilly, F., Rings, D.M. and Allan, G., 2001. Activation of the immune system is the pivotal event in the production of wasting disease in pigs infected with porcine circovirus-2 (PCV2). Veterinary Pathology, 38, 31–42

  21. Kuntz-Simon, G., Blanchard, P., Cherbonnel, M., Jestin, A. and Jestin, V., 2002. Baculovirus-expressed muscovy duck reovirus sigmaC protein induces serum neutralizing antibodies and protection against challenge. Vaccine, 20, 3113

  22. Liu, Q., Tikoo, S.K. and Babiuk, L.A., 2001. Nuclear localization of the ORF2 protein encoded by porcine circovirus Type 2. Virology, 285, 91–99

  23. Madec, F., Eveno, E., Morvan, P., Hamon, L., Blanchard, P., Cariolet, R., Amenna, N., Marvan, H., Truong, C., Mahe, D., Albina, E. and Jestin, A., 2000. Post-weaning multisystemic wasting syndrome (PMWS) in pigs in France: clinical observations from follow-up studies on affected farms. Livestock Production Science, 63, 223–233

  24. Morozov, I., Sirinarumitr, T., Sorden, S.D., Halbur, P.G., Morgen, M.K., Yoon, K.J., and Paul, P.S., 1998. Detection of a novel strain of porcine circovirus in pigs with postweaning multisystemic wasting syndrome, Journal of Clinical Microbiology, 36, 2535–2541

  25. Nawagitgul, P., Morozov, I., Bolin, S.R., Harms, P.A., Sorden, S.D. and Paul, P.S., 2000. Open reading frame 2 of porcine circovirus type 2 encodes a major capsid protein. Journal of General Virology, 81, 2281–2287

  26. Nawagitgul, P., Harms, P.A., Morozov, I., Thacker, B.J., Sorden, S.D., Lekcharoensuk, C. and Paul, P.S., 2002. Modified indirect porcine circovirus (PCV) type 2-based and recombinant capsid protein (ORF2)-based enzyme-linked immunosorbent assays for detection of antibodies to PCV. Clinical and Diagnostic Laboratory Immunology, 9, 33–40

  27. O'Reilly, D.R., Miller, L.K. and Luckow, U.A., 1992. Baculovirus Expression Vectors: A Laboratory Manual (W.H. Freeman, New York), 109–134

  28. Rovira, A., Balasch, M., Segales, J., Garcia, L., Plana-Duran, J., Rosell, C., Ellerbrok, H., Mankertz, A. and Domingo, M., 2002. Experimental inoculation of conventional pigs with porcine reproductive and respiratory syndrome virus and porcine circovirus 2. Journal of Virology, 76, 3232–3239

  29. Todd, D., McNulty, M.S., Mankertz, A., et al., 2000. Family Circoviridae. In: F.A. Murphy, C.M. Fauquet, D.H.L. Bishop, et al. (eds.), Virus Taxonomy. Classification and Nomenclature of Viruses, Seventh Report of the International Committee of Taxonomy of Viruses (Academic Press, New York), 299–303

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Fan, H., Ju, C., Tong, T. et al. Immunogenicity of Empty Capsids of Porcine Circovius Type 2 Produced in Insect Cells. Vet Res Commun 31, 487–496 (2007). https://doi.org/10.1007/s11259-007-3469-7

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  • ORF2
  • porcine circovirus type 2 (PCV2)
  • postweaning multisystemic wasting syndrome
  • PMWS subunit vaccine