1887

Abstract

The aim of this study was to describe the generation of a PCV2 (porcine circovirus type 2) infectious clone (pIC-PCV2) and its infectivity under and conditions. The constructed pIC-PCV2 contained the whole PCV2 genome from a German isolate together with a partial duplication of 467 bp. PK-15 cells were transfected with pIC-PCV2 and an indirect immune fluorescence assay (IFA) was performed 7 days post-transfection. The PCV2 Cap gene was expressed in approximately 20 % of the cultured cells, and only the recombination product, and not pIC-PCV2, was subsequently detected by PCR and Southern blot. This result indicated that infection by pIC-PCV2 delivered genomic PCV2 DNA specifically into susceptible cells and led to the expression of a functional virus genome. Eighteen 30- to 40-day-old conventional pigs were distributed into three groups. Group 1 pigs (=6) were inoculated intranasally (i.n.) with a Spanish isolate of PCV2 propagated in cell culture; pigs from group 2 (=6) were inoculated with pIC-PCV2 intramuscularly (i.m.), and the last group of pigs (=6) was inoculated with pIC-PCV2 intraperitoneally (i.p.). All pigs remained clinically healthy during the whole experimental period (35 days). Pigs that received pIC-PCV2 i.p. and i.m., as well as those PCV2 i.n. inoculated, became infected based on an hybridization (ISH), PCR, TaqMan PCR and serological results. The results of this study confirm that cloned PCV2 genomic DNA is infectious both and , and is able to cause PMWS-like lesions in i.p. and i.m. experimentally inoculated pigs.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.79836-0
2004-05-01
2019-11-19
Loading full text...

Full text loading...

/deliver/fulltext/jgv/85/5/vir851259.html?itemId=/content/journal/jgv/10.1099/vir.0.79836-0&mimeType=html&fmt=ahah

References

  1. Allan, G. M., McNeilly, F., Cassidy, J. P., Reilly, G. A., Adair, B., Ellis, W. A. & McNulty, M. S. ( 1995; ). Pathogenesis of porcine circovirus; experimental infections of colostrum deprived piglets and examination of pig foetal material. Vet Microbiol 44, 49–64.[CrossRef]
    [Google Scholar]
  2. Allan, G. M., Kennedy, S., McNeilly, F., Foster, J. C., Ellis, J. A., Krakowka, S. J., Meehan, B. M. & Adair, B. M. ( 1999; ). Experimental reproduction of severe wasting disease by co-infection of pigs with porcine circovirus and porcine parvovirus. J Comp Pathol 121, 1–11.[CrossRef]
    [Google Scholar]
  3. Allan, G. M., McNeilly, F., Ellis, J., Krakowka, S., Meehan, B., McNair, I., Walker, I. & Kennedy, S. ( 2000a; ). Experimental infection of colostrum deprived piglets with porcine circovirus 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) potentiates PCV2 replication. Arch Virol 145, 2421–2429.[CrossRef]
    [Google Scholar]
  4. Allan, G. M., McNeilly, F., Meehan, B. M., Ellis, J. A., Connor, T. J., McNair, I., Krakowka, S. & Kennedy, S. ( 2000b; ). A sequential study of experimental infection of pigs with porcine circovirus and porcine parvovirus: immunostaining of cryostat sections and virus isolation. J Vet Med B Infect Dis Vet Public Health 47, 81–94.[CrossRef]
    [Google Scholar]
  5. Allan, G. M., McNeilly, F., Meehan, B. & 7 other authors ( 2002; ). Reproduction of PMWS with a 1993 Swedish isolate of PCV-2. Vet Rec 150, 255–256.
    [Google Scholar]
  6. Balasch, M., Segales, J., Rosell, C., Domingo, M., Mankertz, A., Urniza, A. & Plana-Duran, J. ( 1999; ). Experimental inoculation of conventional pigs with tissue homogenates from pigs with post-weaning multisystemic wasting syndrome. J Comp Pathol 121, 139–148.[CrossRef]
    [Google Scholar]
  7. Bolin, S. R., Stoffregen, W. C., Nayar, G. P. & Hamel, A. L. ( 2001; ). Postweaning multisystemic wasting syndrome induced after experimental inoculation of cesarean-derived, colostrum-deprived piglets with type 2 porcine circovirus. J Vet Diagn Invest 13, 185–194.[CrossRef]
    [Google Scholar]
  8. Boyer, J. C. & Haenni, A. L. ( 1994; ). Infectious transcripts and cDNA clones of RNA viruses. Virology 198, 415–426.[CrossRef]
    [Google Scholar]
  9. Choi, C. & Chae, C. ( 2000; ). Distribution of porcine parvovirus in porcine circovirus 2-infected pigs with postweaning multisystemic wasting syndrome as shown by in-situ hybridization. J Comp Pathol 123, 302–305.[CrossRef]
    [Google Scholar]
  10. Clark, E. G. ( 1997; ). Post-weaning multisystemic wasting syndrome. Proc Am Assoc Swine Pract 28, 499–501.
    [Google Scholar]
  11. Cox, G. J., Zamb, T. J. & Babiuk, L. A. ( 1993; ). Bovine herpesvirus 1: immune responses in mice and cattle injected with plasmid DNA. J Virol 67, 5664–5667.
    [Google Scholar]
  12. Ellis, J., Krakowka, S., Lairmore, M. & 11 other authors ( 1999; ). Reproduction of lesions of postweaning multisystemic wasting syndrome in gnotobiotic piglets. J Vet Diagn Invest 11, 3–14.[CrossRef]
    [Google Scholar]
  13. Fenaux, M., Halbur, P. G., Haqshenas, G., Royer, R., Thomas, P., Nawagitgul, P., Gill, M., Toth, T. E. & Meng, X. J. ( 2002; ). Cloned genomic DNA of type 2 porcine circovirus is infectious when injected directly into the liver and lymph nodes of pigs: characterization of clinical disease, virus distribution, and pathologic lesions. J Virol 76, 541–551.[CrossRef]
    [Google Scholar]
  14. Fenaux, M., Opriessnig, T., Halbur, P. G. & Meng, X. J. ( 2003; ). Immunogenicity and pathogenicity of chimeric infectious DNA clones of pathogenic porcine circovirus type 2 (PCV2) and nonpathogenic PCV1 in weanling pigs. J Virol 77, 11232–11243.[CrossRef]
    [Google Scholar]
  15. Gerdts, V., Jons, A., Makoschey, B., Visser, N. & Mettenleiter, T. C. ( 1997; ). Protection of pigs against Aujeszky's disease by DNA vaccination. J Gen Virol 78, 2139–2146.
    [Google Scholar]
  16. Haagmans, B. L., van Rooij, E. M., Dubelaar, M., Kimman, T. G., Horzinek, M. C., Schijns, V. E. & Bianchi, A. T. ( 1999; ). Vaccination of pigs against pseudorabies virus with plasmid DNA encoding glycoprotein D. Vaccine 17, 1264–1271.[CrossRef]
    [Google Scholar]
  17. Harding, J. C. S. & Clark, E. G. ( 1997; ). Recognizing and diagnosing postweaning multisystemic wasting syndrome (PMWS). Swine Health Prod 5, 201–203.
    [Google Scholar]
  18. Harms, P. A., Sorden, S. D., Halbur, P. G., Bolin, S. R., Lager, K. M., Morozov, I. & Paul, P. S. ( 2001; ). Experimental reproduction of severe disease in CD/CD pigs concurrently infected with type 2 porcine circovirus and porcine reproductive and respiratory syndrome virus. Vet Pathol 38, 528–539.[CrossRef]
    [Google Scholar]
  19. Hirai, T., Nunoya, T., Ihara, T., Kusanagi, K. & Shibuya, K. ( 2001; ). Dual infection with PCV-2 and porcine epidemic diarrhoea virus in neonatal piglets. Vet Rec 148, 482–484.[CrossRef]
    [Google Scholar]
  20. Kennedy, S., Moffett, D., McNeilly, F., Meehan, B., Ellis, J., Krakowka, S. & Allan, G. M. ( 2000; ). Reproduction of lesions of postweaning multisystemic wasting syndrome by infection of conventional pigs with porcine circovirus type 2 alone or in combination with porcine parvovirus. J Comp Pathol 122, 9–24.[CrossRef]
    [Google Scholar]
  21. Kiupel, M., Stevenson, G. W., Kanitz, C. L., Anothayanontha, L., Latimer, K. S. & Mittal, S. K. ( 1999; ). Cellular localization of porcine circovirus in postweaning pigs with chronic wasting disease. Eur J Vet Pathol 5, 77–82.
    [Google Scholar]
  22. Krakowka, S., Ellis, J. A., Meehan, B., Kennedy, S., McNeilly, F. & Allan, G. ( 2000; ). Viral wasting syndrome of swine: experimental reproduction of postweaning multisystemic wasting syndrome in gnotobiotic swine by coinfection with porcine circovirus 2 and porcine parvovirus. Vet Pathol 37, 254–263.[CrossRef]
    [Google Scholar]
  23. Krakowka, S., Ellis, J. A., McNeilly, F., Ringler, S., Rings, D. M. & 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 (PCV-2). Vet Pathol 38, 31–42.[CrossRef]
    [Google Scholar]
  24. Larochelle, R., Magar, R. & D'Allaire, S. ( 2002; ). Genetic characterization and phylogenetic analysis of porcine circovirus type 2 (PCV2) strains from cases presenting various clinical conditions. Virus Res 90, 101–112.[CrossRef]
    [Google Scholar]
  25. Magar, R., Larochelle, R., Thibault, S. & Lamontagne, L. ( 2000; ). Experimental transmission of porcine circovirus type 2 (PCV2) in weaned pigs: a sequential study. J Comp Pathol 123, 258–269.[CrossRef]
    [Google Scholar]
  26. Mankertz, A. & Hillenbrand, B. ( 2001; ). Replication of porcine circovirus type 1 requires two proteins encoded by the viral rep gene. Virology 279, 429–438.[CrossRef]
    [Google Scholar]
  27. Mankertz, A., Persson, F., Mankertz, J., Blaess, G. & Buhk, H. J. ( 1997; ). Mapping and characterization of the origin of DNA replication of porcine circovirus. J Virol 71, 2562–2566.
    [Google Scholar]
  28. Mankertz, A., Domingo, M., Folch, J. M., LeCann, P., Jestin, A., Segales, J., Chmielewicz, B., Plana-Duran, J. & Soike, D. ( 2000; ). Characterisation of PCV-2 isolates from Spain, Germany and France. Virus Res 66, 65–77.[CrossRef]
    [Google Scholar]
  29. Mankertz, A., Mueller, B., Steinfeldt, T., Schmitt, C. & Finsterbusch, T. ( 2003; ). New reporter gene-based replication assay reveals exchangeability of replication factors of porcine circovirus types 1 and 2. J Virol 77, 9885–9893.[CrossRef]
    [Google Scholar]
  30. Marco, A. J., Domingo, M., Ruberte, J., Carretero, A., Briones, V. & Dominguez, L. ( 1992; ). Lymphatic drainage of Listeria monocytogenes and Indian ink inoculated in the peritoneal cavity of the mouse. Lab Anim 26, 200–205.[CrossRef]
    [Google Scholar]
  31. Olvera, A., Sibila, M., Calsamiglia, M., Segales, J. & Domingo, M. ( 2004; ). Comparison of PCV2 load in serum quantified by a newly developed real time PCR in PMWS and PDNS naturally affected pigs. J Virol Meth 117, 75–80.[CrossRef]
    [Google Scholar]
  32. Quintana, J., Balasch, M., Segalés, J., Calsamiglia, M., Rodríguez-Arrioja, G. M., Plana-Duran, J. & Domingo, M. ( 2002; ). Experimental inoculation of porcine circoviruses type 1 (PCV1) and type 2 (PCV2) in rabbits and mice. Vet Rec 33, 229–237.[CrossRef]
    [Google Scholar]
  33. Rodríguez-Arrioja, G. M., Segalés, J., Balasch, M., Rosell, C., Quintant, J., Folch, J. M., Plana-Duran, J., Mankertz, A. & Domingo, M. ( 2000; ). Serum antibodies to porcine circovirus type 1 and type 2 in pigs with and without PMWS. Vet Rec 146, 762–764.[CrossRef]
    [Google Scholar]
  34. Rosell, C., Segales, J., Plana-Duran, J. & 7 other authors ( 1999; ). Pathological, immunohistochemical, and in-situ hybridization studies of natural cases of postweaning multisystemic wasting syndrome (PMWS) in pigs. J Comp Pathol 120, 59–78.[CrossRef]
    [Google Scholar]
  35. Rosell, C., Segales, J. & Domingo, M. ( 2000; ). Hepatitis and staging of hepatic damage in pigs naturally infected with porcine circovirus type 2. Vet Pathol 37, 687–692.[CrossRef]
    [Google Scholar]
  36. Rovira, A., Balasch, M., Segales, J., Garcia, L., Plana-Duran, J., Rosell, C., Ellerbrok, H., Mankertz, A. & Domingo, M. ( 2002; ). Experimental inoculation of conventional pigs with porcine reproductive and respiratory syndrome virus and porcine circovirus 2. J Virol 76, 3232–3239.[CrossRef]
    [Google Scholar]
  37. Segales, J. & Domingo, M. ( 2002; ). Postweaning multisystemic wasting syndrome (PMWS) in pigs. A review. Vet Q 24, 109–124.[CrossRef]
    [Google Scholar]
  38. Tischer, I., Gelderblom, H., Vettermann, W. & Koch, M. A. ( 1982; ). A very small porcine virus with circular single-stranded DNA. Nature 295, 64–66.[CrossRef]
    [Google Scholar]
  39. Tischer, I., Mields, W., Wolff, D., Vagt, M. & Griem, W. ( 1986; ). Studies on epidemiology and pathogenicity of porcine circovirus. Arch Virol 91, 271–276.[CrossRef]
    [Google Scholar]
  40. Todd, D., McNulty, M. S., Mankertz, A., Lukert, P., Randles, J. W. & Dale, J. L. ( 2000; ). Circoviridae. In Seventh Report of the International Committee on Taxonomy of Viruses, pp. 299–303. Edited by M. H. V. van Regenmortel, C. M. Fauquet, D. H. L. Bishop, E. B. Carstens, M. K. Estes, S. M. Lemon, J. Maniloff, M. A. Mayo, D. J. McGeoch, C. R. Pringle & R. B. Wickner. San Diego: Academic Press.
  41. Ulmer, J. B., Donnelly, J. J. & Liu, M. A. ( 1997; ). Categories of products. Mechanism of action (Advantages/Disadvantages). DNA vaccines: a new category. In Veterinary Vaccinology, pp. 285–292. Edited by P.-P. Pastoret, J. Blancou, P. Vannier & C. Verschueren. Elsevier Science.
  42. Valerie, K. & Povirk, L. F. ( 2003; ). Regulation and mechanisms of mammalian double-strand break repair. Oncogene 22, 5792–5812.[CrossRef]
    [Google Scholar]
  43. van Drunen Littel-van den Hurk, S., Gerdts, V., Loehr, B. I., Pontarollo, R., Rankin, R., Uwiera, R. & Babiuk, L. A. ( 2000; ). Recent advances in the use of DNA vaccines for the treatment of diseases of farmed animals. Adv Drug Deliv Rev 43, 13–28.[CrossRef]
    [Google Scholar]
  44. van Rooij, E. M., Haagmans, B. L., de Visser, Y. E., de Bruin, M. G., Boersma, W. & Bianchi, A. T. ( 1998; ). Effect of vaccination route and composition of DNA vaccine on the induction of protective immunity against pseudorabies infection in pigs. Vet Immunol Immunopathol 66, 113–126.[CrossRef]
    [Google Scholar]
  45. Verbeke, G. & Molenberghs, G. ( 2000; ). Linear Mixed Models for Longitudinal Data. New York: Springer.
  46. West, S. C. ( 2003; ). Molecular views of recombination proteins and their control. Nat Rev Mol Cell Biol 4, 435–445.
    [Google Scholar]
  47. Wolff, J. A., Malone, R. W., Williams, P., Chong, W., Acsadi, G., Jani, A. & Felgner, P. L. ( 1990; ). Direct gene transfer into mouse muscle in vivo. Science 247, 1465–1468.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.79836-0
Loading
/content/journal/jgv/10.1099/vir.0.79836-0
Loading

Data & Media loading...

Most Cited This Month

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error