1887

Journal of General Virology header logo

Volume 78, Issue 9

Epitope mapping of the nucleocapsid protein of European and North American isolates of porcine reproductive and respiratory syndrome virus Free

Abstract

Two major genotypes of porcine reproductive and respiratory syndrome virus (PRRSV) have been described, which correspond to the European and North American isolates. PRRSV nucleocapsid (N) protein has been identified as the most immunodominant viral protein. The N genes from two PRRSV isolates, Olot/91 (European) and Québec 807/94 (North American), were cloned and expressed in: (i) baculovirus under the control of the polyhedrin promoter and (ii) using the pET3x system. The N protein from both isolates was expressed much more efficiently in as a fusion protein than in baculovirus. The antigenicity of the protein was similar in both systems and it was recognized by a collection of 48 PRRSV-positive pig sera. The antigenic structure of the PRRSV N protein was investigated using seven monoclonal antibodies (MAbs) and overlapping fragments of the protein expressed in . Four MAbs recognized two discontinuous epitopes that were present in the partially folded protein, or at least a large fragment comprising the first 78 residues. The other three MAbs revealed the presence of a common antigenic site localized in the central region of the protein (amino acids 50–66). This region is well conserved among different isolates of European and North American origin and is the most hydrophilic region of the protein. However, this epitope, although recognized by the MAbs and many pig sera, is not useful for diagnostic purposes. Moreover, none of the N protein fragments were able to mimic the antigenicity of the entire protein.

  • Published Online:
© Society for General Microbiology 1997
Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-78-9-2269
1997-09-01
2024-04-19
Loading full text...

Full text loading...

/deliver/fulltext/jgv/78/9/9292014.html?itemId=/content/journal/jgv/10.1099/0022-1317-78-9-2269&mimeType=html&fmt=ahah

References

  1. Chirnside E. D., Francis P. M., Mumford J. A. 1995; Expression cloning and antigenic analysis of the nucleocapsid protein of equine arteritis virus. Virus Research 39:277–288
     [Google Scholar]
  2. Collins J. E., Benfield D. A., Christianson W. T., Harris J. C., Hennings D. P., Shaw S. M., Goyal S. M., McCullough S., Morrisson R. B., Joo H. S., Gorcyca D., Chladek D. W. 1992; Isolation of swine infertility and respiratory syndrome virus (isolate ATCC VR-2332) in North America and experimental reproduction in gnotobiotic pigs. Journal of Veterinary Diagnostic Investigation 4:117–126
     [Google Scholar]
  3. Dea S., Gagnon C. A., Mardassi H., Milane G. 1996; Antigenic variability among North American and European strains of porcine reproductive and respiratory syndrome virus as defined by monoclonal antibodies to the matrix protein. Journal of Clinical Microbiology 34:1488–1493
     [Google Scholar]
  4. Drew T. W., Meulenberg J. J. M., Sands J. J., Paton D. J. 1995; Production, characterization and reactivity of monoclonal antibodies to porcine reproductive and respiratory syndrome virus. Journal of General Virology 76:1361–1369
     [Google Scholar]
  5. Felgner P. L., Gadek T. R., Holm M., Roman R., Chan H. W., Wenz M., Northrop J. P., Ringold G. M., Danielsen M. 1987; Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure. Proceedings of the National Academy of Sciences, USA 84:7413–7417
     [Google Scholar]
  6. Grodberg J., Dunn J. J. 1988; OmpT encodes the Escherichia coli outer membrane protease that cleaves T7 RNA polymerase during purification. Journal of Bacteriology 170:1245
     [Google Scholar]
  7. Harlow E., Lane D. 1988 Antibodies: A Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  8. Hopp T. P., Woods K. R. 1981; Prediction of protein antigenic determinants from amino acid sequences. Proceedings of the National Academy of Sciences, USA 78:3824–3828
     [Google Scholar]
  9. Kitts P. A., Possee R. D. 1993; A method for producing recombinant baculovirus expression vectors at high frequency. Biotechniques 14:810–817
     [Google Scholar]
  10. Lopez de Turiso J. A., Cortés E., Martinez C., Ruiz de Ybañez R., Simarro I., Vela C., Casal J. I. 1992; Recombinant vaccine for canine parvovirus in dogs. Journal of Virology 66:2748–2753
     [Google Scholar]
  11. Magar R., Larochelle S., Dea S., Gagnon C. A., Nelson E. A., Christopher-Hennings J., Benfield D. A. 1995; Antigenic comparison of Canadian and US isolates of porcine reproductive and respiratory syndrome virus using monoclonal antibodies to the nucleo- capsid protein. Canadian Journal of Veterinary Research 59:232–234
     [Google Scholar]
  12. Mardassi H., Mounir S., Dea S. 1994; Identification of major differences in the nucleocapsid protein genes of a Québec strain and European strains of porcine reproductive and respiratory syndrome virus. Journal of General Virology 75:681–685
     [Google Scholar]
  13. Martinez-Torrecuadrada J. L., Casal J. I. 1995; Identification of a linear neutralization domain in the protein VP2 of African horsesickness virus. Virology 210:391–399
     [Google Scholar]
  14. Martinez-Torrecuadrada J. L., Iwata H., Venteo A., Casal I., Roy P. 1994; Expression and characterization of the two outer capsid proteins of African horsesickness virus : the role of VP2 in virus neutralization. Virology 202:348–359
     [Google Scholar]
  15. Matsuura Y., Possee R. D., Overton H. A., Bishop D. H. L. 1987; Baculovirus expression vectors: the requirements for high level expression of proteins, including glycoproteins. Journal of General Virology 68:1233–1250
     [Google Scholar]
  16. Meng X.-J., Paul P. S., Halbur P. G., Lum M. A. 1995; Phylogenetic analysis ofthe putative M (ORF6) andN (ORF7) genes ofporcine reproductive and respiratory syndrome virus (PRRSV) : implication for the existence of two genotypes of PRRSV in the US and Europe. Archives of Virology 140:745–755
     [Google Scholar]
  17. Meng X. J., Paul P. S., Halbur P. G., Morozov I. 1996; Sequence comparison of open reading frames 2 to 5 of low and high virulence United States isolates of porcine reproductive and respiratory syndrome virus. Journal of General Virology 76:3181–3188
     [Google Scholar]
  18. Meulenberg J. J. M., Petersen Den Besten A. 1996; Identification and characterization of a sixth structural protein of Lelystad virus : the glycoprotein GP2 encoded by ORF2 is incorporated in virus particles. Virology 225:44–51
     [Google Scholar]
  19. Meulenberg J. J., Hulst M. M., De Meijer E. J., Moonen P. L. J. M., Den Besten A. P., De Kluyver E. P., Wensvoort G., Moormann R. J. M. 1993; Lelystad virus, the causative agent of porcine epidemic abortion and respiratory syndrome (PEARS), is related to LDV and EAV. Virology 192:62–72
     [Google Scholar]
  20. Meulenberg J. J. M., Bende R. J., Pol J. M. A., Wensvoort G., Moorman R. J. M. 1995a; Nucleocapsid protein of Lelystad virus. Expression by recombinant baculovirus : immunological properties and suitability for the detection of serum antibodies. Clinical Diagnostic Laboratory Immunology 2:652–656
     [Google Scholar]
  21. Meulenberg J., Den Besten A. P., De Kluyver E. P., Moorman R. J. M., Schaaper W. M. M., Wensvoort G. 1995b; Characterization of proteins encoded by ORFs 2 to 7 of Lelystad virus. Virology 206:155–163
     [Google Scholar]
  22. Murtaugh M. P., Elam M. R., Kakach L. T. 1995; Comparison of the structural protein coding sequences of the VR-2332 and Lelystad virus strains of the PRRS virus. Archives of Virology 140:1451–1460
     [Google Scholar]
  23. Nelson E. A., Christopher-Hennings J., Drew T., Wensvoort G., Collins J. E., Benfield D. A. 1993; Differentiation of United States and European isolates of porcine reproductive and respiratory syndrome virus by monoclonal antibodies. Journal of Clinical Microbiology 31:3184–3189
     [Google Scholar]
  24. Plana-Durán J., Vayreda M., Vilarasa J. L., Bastons M., Rosell R., Martinez M., San Gabriel A., Pujols J., Badiola J. L., Ramos J. A., Domingo M. 1992; Porcine epidemic abortion and respiratory syndrome (mystery swine disease). Isolation in Spain of the causative agent and experimental reproduction of the disease. Veterinary Microbiology 33:203–211
     [Google Scholar]
  25. Plana-Durán J., Climent I., Sarraseca J., Urniza A., Cortés E., Vela C., Casal J. I. 1997; Baculovirus expression of proteins of porcine reproductive and respiratory syndrome virus strain Olot/91. Involvement of ORF3 and ORF5 proteins in protection. Virus Genes 14:19–29
     [Google Scholar]
  26. Rost B. 1996; PHD : predicting one-dimensional protein structure by profile based neural networks. Methods in Enzymology 266:525–539
     [Google Scholar]
  27. Rota P. A., Black R. A., De B. K., Harmon M. W., Kendal A. P. 1990; Expression of influenza A and B virus nucleoprotein antigens in baculovirus. Journal of General Virology 71:1545–1554
     [Google Scholar]
  28. Sanger F., Nicklen S., Coulson A. R. 1977; DNA sequencing with chain terminating inhibitors. Proceedings of the National Academy of Sciences, USA 74:5463–5467
     [Google Scholar]
  29. Sanz A., Garcia J., Cortés E., Sarraseca J., Climent I., Plana J., Casal J. I. 1995; Development of diagnostic assays for antibodies to PRRSV using monoclonal antibodies and a recombinant antigen. In Proceedings of the 2nd Symposium on Aujeszky and PRRS viruses Copenhagen, Denmark:
     [Google Scholar]
  30. Shibata I., Uruno K., Mori M. 1996; Serological property and replication in cell cultures of porcine reproductive and respiratory syndrome virus isolated in Japan. Journal of Veterinary Medical Science 58:805–807
     [Google Scholar]
  31. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. 1990; Use of T7 RNA polymerase to direct expression of cloned genes. Methods in Enzymology 185:60–89
     [Google Scholar]
  32. Suarez P., Zardoya R., Martín M. J., Prieto C., Dopazo J., Solana A., Castro J. M. 1996; Phylogenetic relationships ofEuropean strains of porcine reproductive and respiratory syndrome virus (PRRSV) inferred from DNA sequences ofputative ORF-5 and ORF-7 genes. Virus Research 42:159–165
     [Google Scholar]
  33. Summers M. D., Smith G. E. 1987; A manual of methods for baculovirus vectors and insect cell culture procedures. Texas Agricultural Experimental Station Bulletin1555
     [Google Scholar]
  34. Terpstra C., Wensvoort G., Pol J. M. A. 1991; Experimental reproduction of porcine epidemic abortion and respiratory syndrome (mystery swine disease) by infection with Lelystad virus: Koch’s postulates fulfilled. Veterinary Quarterly 13:131–136
     [Google Scholar]
  35. van Nieuwstadt A. P., Meulenberg J. J. M., van Essen-Zanderbergen A., Petersen Den Besten A., Bende R. J., Moorman R. J. M., Wensvoort G. 1996; Proteins encoded by open reading frames 3 and 4 of the genome of Lelystad virus (Arteriviridae) are structural proteins of the virion. Journal of Virology 70:4767–4772
     [Google Scholar]
  36. Vapalahti O., Lundkvist A, Paukku K., Julkunen I., Lankinen H., Vaheri A. 1996; Antigenic properties and diagnostic potential of Puumala virus nucleocapsid protein expressed in insect cells. Journal of Clinical Microbiology 34:119–125
     [Google Scholar]
  37. Wensvoort G., Terpstra C., Pol J. M. A., Ter Laak E. A., Bloemraad M., De Kluyver E. P., Kragten C., Van Buiten L., Den Besten A. P., Wagenaar F., Broekhuijsen J. M., Moonen P. L. J. M., Zetstra T., De Boer E. A., Tibbem H. J., De Jong M. F., Vant Veld P., Groenland G. J. R., Van Gennep J. A., Voets M. T., Verheijden J. H. M., Braamskamp J. 1991; Mystery swine disease in the Netherlands: the isolation of Lelystad virus. Veterinary Quarterly 13:121–130
     [Google Scholar]
  38. Wensvoort G., De Kluyver E. P., Luijtze E. A., Den Besten A. P., Harris L., Collins J. E., Christianson W. T., Chladek D. 1992; Antigenic comparison of Lelystad virus and swine respiratory syndrome (SIRS) virus. Journal of Veterinary Diagnostic Investigation 4:134–138
     [Google Scholar]
  39. Wieczorek-Krohmer M., Weiland F., Conzelman K., Kohl D., Visser N., Van Woensel P., Thiel H.-J., Weiland E. 1996; Porcine reproductive and respiratory syndrome virus (PRRSV): monoclonal antibodies detect common epitopes on two viral proteins of European and US isolates. Veterinary Microbiology 51:257–266
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-78-9-2269
Loading
Epitope mapping of the nucleocapsid protein of European and North American isolates of porcine reproductive and respiratory syndrome virus
J. Gen. Virol. 78, 2269 (1997); https://doi.org/10.1099/0022-1317-78-9-2269
/content/journal/jgv/10.1099/0022-1317-78-9-2269
/content/journal/jgv/10.1099/0022-1317-78-9-2269
Loading

Data & Media loading...

Most cited Most Cited RSS feed

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