Archives of Virology

, Volume 159, Issue 8, pp 2033–2041 | Cite as

An influenza A virus hemagglutinin (HA) epitope inserted in and expressed from several loci of the infectious bursal disease virus genome induces HA-specific antibodies

  • Yung-Yi C. Mosley
  • Ching Ching Wu
  • Tsang Long Lin
Original Article


The N-terminus of infectious bursal disease virus (IBDV) VP5 has been shown to be capable of tolerating the insertion of small epitopes. The objective of the present study was to determine if IBDV genomic sites, including the 5’ end of vp5, could carry an influenza A virus hemagglutinin (HA) epitope. HA-expressing IBDVs were generated when the HA epitope was fused to the N-terminus of VP5 (HA5-IBDV) or VP4 (HA4-IBDV) or the C-terminus of VP1 (1HA-IBDV). Viral titers obtained after co-transfection with cDNA from the ha-containing segment and the complementary genomic segment were 1.3 × 104, 3.7 × 103 and 3.8 × 104 pfu/ml for HA5-IBDV, HA4-IBDV and 1HA-IBDV, respectively. The HA tag expression remained stable after 10 passages when the tag gene was inserted into the vp4 and vp1 genes. HA-IBDVs did not cause pathogenicity in specific-pathogen-free (SPF) chickens. However, only HA4-IBDV and 1HA-IBDV induced HA-specific antibodies, which were measured by ELISA with a maximum optical density (OD) value of 0.701 and 0.769, respectively, at 24 days after infection. Thus, IBDV can potentially be employed as a bivalent viral vector when the epitope is fused with VP4 or VP1.


Antibody Titer Infectious Bursal Disease Virus Infectious Pancreatic Necrosis Virus Infectious Bursal Disease Virus Strain Infectious Bursal Disease Virus Infection 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



We thank Debby Sherman for her assistance with electron microscopy. We also thank Bong-Suk Kim, Yi-Ning Chen, Chih-Chun Lee, Chun-Yu Tung, Wan-Jung Chen and Ching Giap Tan for their help with the animal study.

Supplementary material

705_2014_2036_MOESM1_ESM.doc (1.2 mb)
Supplementary material 1 (DOC 1223 kb)


  1. 1.
    Allan WH, Faragher JT, Cullen GA (1972) Immunosuppression by the infectious bursal agent in chickens immunised against Newcastle disease. Vet Rec 90:511–512PubMedCrossRefGoogle Scholar
  2. 2.
    Azad AA, Jagadish MN, Brown MA, Hudson PJ (1987) Deletion mapping and expression in Escherichia coli of the large genomic segment of a birnavirus. Virology 161:145–152PubMedCrossRefGoogle Scholar
  3. 3.
    Balamurugan V, Kataria JM (2006) Economically important non-oncogenic immunosuppressive viral diseases of chicken—current status. Vet Res Commun 30:541–566PubMedCrossRefGoogle Scholar
  4. 4.
    Berg TP (2000) Acute infectious bursal disease in poultry: a review. Avian Pathol 29:175–194PubMedCrossRefGoogle Scholar
  5. 5.
    Birghan C, Mundt E, Gorbalenya AE (2000) A non-canonical lon proteinase lacking the ATPase domain employs the ser-Lys catalytic dyad to exercise broad control over the life cycle of a double-stranded RNA virus. EMBO J 19:114–123PubMedCentralPubMedCrossRefGoogle Scholar
  6. 6.
    Castón JR, Martínez-Torrecuadrada JL, Maraver A, Lombardo E, Rodríguez JF, Casal JI, Carrascosa JL (2001) C terminus of infectious bursal disease virus major capsid protein VP2 is involved in definition of the T number for capsid assembly. J Virol 75:10815–10828PubMedCentralPubMedCrossRefGoogle Scholar
  7. 7.
    Coulibaly F, Chevalier C, Gutsche I, Pous J, Navaza J, Bressanelli S, Delmas B, Rey FA (2005) The birnavirus crystal structure reveals structural relationships among icosahedral viruses. Cell 120:761–772PubMedCrossRefGoogle Scholar
  8. 8.
    Faragher JT, Allan WH, Cullen GA (1972) Immunosuppressive effect of the infectious bursal agent in the chicken. Nat New Biol 237:118–119PubMedCrossRefGoogle Scholar
  9. 9.
    Fernández-Arias A, Risco C, Martínez S, Albar JP, Rodríguez JF (1998) Expression of ORF A1 of infectious bursal disease virus results in the formation of virus-like particles. J Gen Virol 79(Pt 5):1047–1054PubMedGoogle Scholar
  10. 10.
    Field J, Nikawa J, Broek D, MacDonald B, Rodgers L, Wilson IA, Lerner RA, Wigler M (1988) Purification of a RAS-responsive adenylyl cyclase complex from Saccharomyces cerevisiae by use of an epitope addition method. Mol Cell Biol 8:2159–2165PubMedCentralPubMedGoogle Scholar
  11. 11.
    Graham SC, Sarin LP, Bahar MW, Myers RA, Stuart DI, Bamford DH, Grimes JM (2011) The N-terminus of the RNA polymerase from infectious pancreatic necrosis virus is the determinant of genome attachment. PLoS Pathog 7:e1002085PubMedCentralPubMedCrossRefGoogle Scholar
  12. 12.
    Green N, Alexander H, Olson A, Alexander S, Shinnick TM, Sutcliffe JG, Lerner RA (1982) Immunogenic structure of the influenza virus hemagglutinin. Cell 28:477–487PubMedCrossRefGoogle Scholar
  13. 13.
    Heine HG, Hyatt AD, Boyle DB (1994) Modification of infectious bursal disease virus antigen VP2 for cell surface location fails to enhance immunogenicity. Virus Res 32:313–328PubMedCrossRefGoogle Scholar
  14. 14.
    Ho SN, Hunt HD, Horton RM, Pullen JK, Pease LR (1989) Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene 77:51–59PubMedCrossRefGoogle Scholar
  15. 15.
    Irigoyen N, Garriga D, Navarro A, Verdaguer N, Rodriguez JF, Caston JR (2009) Autoproteolytic activity derived from the infectious bursal disease virus capsid protein. J Biol Chem 284:8064–8072PubMedCentralPubMedCrossRefGoogle Scholar
  16. 16.
    Jagadish MN, Staton VJ, Hudson PJ, Azad AA (1988) Birnavirus precursor polyprotein is processed in Escherichia coli by its own virus-encoded polypeptide. J Virol 62:1084–1087PubMedCentralPubMedGoogle Scholar
  17. 17.
    Lejal N, Da Costa B, Huet JC, Delmas B (2000) Role of Ser-652 and Lys-692 in the protease activity of infectious bursal disease virus VP4 and identification of its substrate cleavage sites. J Gen Virol 81:983–992PubMedGoogle Scholar
  18. 18.
    Li Z, Wang Y, Xue Y, Li X, Cao H, Zheng SJ (2012) Critical role for voltage-dependent anion channel 2 in infectious bursal disease virus-induced Apoptosis in host cells via interaction with VP5. J Virol 86:1328–1338PubMedCentralPubMedCrossRefGoogle Scholar
  19. 19.
    Lombardo E, Maraver A, Espinosa I, Fernandez-Arias A, Rodriguez JF (2000) VP5, the nonstructural polypeptide of infectious bursal disease virus, accumulates within the host plasma membrane and induces cell lysis. Virology 277:345–357PubMedCrossRefGoogle Scholar
  20. 20.
    Luque D, Saugar I, Rodríguez JF, Verdaguer N, Garriga D, Martín CS, Velázquez-Muriel JA, Trus BL, Carrascosa JL, Castón JR (2007) Infectious bursal disease virus capsid assembly and maturation by structural rearrangements of a transient molecular switch. J Virol 81:6869–6878PubMedCentralPubMedCrossRefGoogle Scholar
  21. 21.
    Luque D, Saugar I, Rejas MT, Carrascosa JL, Rodríguez JF, Castón JR (2009) Infectious Bursal disease virus: ribonucleoprotein complexes of a double-stranded RNA virus. J Mol Biol 386:891–901PubMedCrossRefGoogle Scholar
  22. 22.
    Martin Caballero J, Garzón A, González-Cintado L, Kowalczyk W, Jimenez Torres I, Calderita G, Rodriguez M, Gondar V, Bernal JJ, Ardavín C, Andreu D, Zürcher T, von Kobbe C (2012) Chimeric infectious bursal disease virus-like particles as potent vaccines for eradication of established HPV-16 E7-dependent tumors. PLoS One 7:e52976PubMedCentralPubMedCrossRefGoogle Scholar
  23. 23.
    Mosley YY, Wu CC, Lin TL (2013) Infectious bursal disease virus rescued efficiently with 3’ authentic RNA sequence induces humoral immunity without bursal atrophy. Vaccine 31:704–710PubMedCrossRefGoogle Scholar
  24. 24.
    Mundt E, Beyer J, Müller H (1995) Identification of a novel viral protein in infectious bursal disease virus-infected cells. J Gen Virol 76(Pt 2):437–443PubMedCrossRefGoogle Scholar
  25. 25.
    Müller H, Islam MR, Raue R (2003) Research on infectious bursal disease—the past, the present and the future. Vet Microbiol 97:153–165PubMedCrossRefGoogle Scholar
  26. 26.
    Pan J, Lin L, Tao YJ (2009) Self-guanylylation of birnavirus VP1 does not require an intact polymerase activity site. Virology 395:87–96PubMedCentralPubMedCrossRefGoogle Scholar
  27. 27.
    Peters MA, Lin TL, Wu CC (2005) Real-time RT-PCR differentiation and quantitation of infectious bursal disease virus strains using dual-labeled fluorescent probes. J Virol Methods 127:87–95PubMedCrossRefGoogle Scholar
  28. 28.
    Rémond M, Da Costa B, Riffault S, Parida S, Breard E, Lebreton F, Zientara S, Delmas B (2009) Infectious bursal disease subviral particles displaying the foot-and-mouth disease virus major antigenic site. Vaccine 27:93–98PubMedCrossRefGoogle Scholar
  29. 29.
    Saugar I, Luque D, Oña A, Rodríguez JF, Carrascosa JL, Trus BL, Castón JR (2005) Structural polymorphism of the major capsid protein of a double-stranded RNA virus: an amphipathic alpha helix as a molecular switch. Structure 13:1007–1017PubMedCrossRefGoogle Scholar
  30. 30.
    Sánchez AB, Rodriguez JF (1999) Proteolytic processing in infectious bursal disease virus: identification of the polyprotein cleavage sites by site-directed mutagenesis. Virology 262:190–199PubMedCrossRefGoogle Scholar
  31. 31.
    Upadhyay C, Ammayappan A, Patel D, Kovesdi I, Vakharia VN (2011) Recombinant infectious bursal disease virus carrying hepatitis C virus epitopes. J Virol 85:1408–1414PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Wilson IA, Niman HL, Houghten RA, Cherenson AR, Connolly ML, Lerner RA (1984) The structure of an antigenic determinant in a protein. Cell 37:767–778PubMedCrossRefGoogle Scholar
  33. 33.
    Winterfield RW, Hoerr FJ, Fadly AM (1978) Vaccination against infectious bronchitis and the immunosuppressive effects of infectious bursal disease. Poult Sci 57:386–391PubMedCrossRefGoogle Scholar
  34. 34.
    Xu HT, Si WD, Dobos P (2004) Mapping the site of guanylylation on VP1, the protein primer for infectious pancreatic necrosis virus RNA synthesis. Virology 322:199–210PubMedCrossRefGoogle Scholar
  35. 35.
    Yao K, Vakharia VN (2001) Induction of apoptosis in vitro by the 17-kDa nonstructural protein of infectious bursal disease virus: possible role in viral pathogenesis. Virology 285:50–58PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Wien 2014

Authors and Affiliations

  • Yung-Yi C. Mosley
    • 1
  • Ching Ching Wu
    • 1
  • Tsang Long Lin
    • 1
  1. 1.Department of Comparative Pathobiology, College of Veterinary MedicinePurdue UniversityWest LafayetteUSA

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