Indian Journal of Animal Research

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Indian Journal of Animal Research, volume 47 issue 4 (august 2013) : 292-300

PROKARYOTIC EXPRESSION OF N GENE OF INFECTIOUS BRONCHITIS VIRUS AND ASSESSMENT OF ITS IMMUNOGENICITY AND ANTIGENICITY

Tripti Jain*, Megha Kadam, Asit Jain, B.C. Sarkhel
1Animal Biotechnology Centre, Nataji Deshmukh Pashu Chikitsa Vigyan Vishwavidyalaya, Jabalpur- 482 002, India

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Cite article:- Jain* Tripti, Kadam Megha, Jain Asit, Sarkhel B.C. (2024). PROKARYOTIC EXPRESSION OF N GENE OF INFECTIOUS BRONCHITIS VIRUS AND ASSESSMENT OF ITS IMMUNOGENICITY AND ANTIGENICITY. Indian Journal of Animal Research. 47(4): 292-300. doi: .

ABSTRACT

Infectious bronchitis (IB) is highly contagious disease of great economic importance to the poultry industry. IB Virus has three major structural proteins spike (S) glycoprotein, membrane (M) glycoprotein and nucleocapsid (N) phosphoprotein. As an potent immunogen of coronavirus, N protein is thought to be an appropriate candidate for development of diagnostics and recombinant vaccine due to its highly conserved and immunogenic nature. Cloning and sequencing of N gene of local isolate of IBV has already been done in M.P., so in present study we assessed its immunogenicity with the aim to further  develop recombinant vaccine by exploiting it. N gene was cloned in pQE30 vector and high-level expression of recombinant N protein was achieved in E. coli. SDS-PAGE analysis showed 2 bands of 52 kDa and 45 kDa. Recombinant N protein was purified using Ni-NTA affinity chromatography. Purified recombinant N protein was used to immunize rabbits. Characterization of polyclonal antiserum raised against the purified recombinant N protein was done by DID, WB, dot blot assay and indirect N-ELISA (using recombinant N protein as coating antigen). Polyclonal antisera reacted against the purified recombinant N protein in all the assays. Significant difference in OD450 was found between preimmune and hyperimmune sera in N-ELISA, which indicated potential immunogenecity of recombinant N protein. Purified recombinant N protein also reacted with chicken anti IBV sera collected from vaccinated poultry flocks in WB, N ELISA and dot blot assays which indicated its antigenicity and potential to develop diagnostics in future.

REFERENCES

  1. Boots, A.M.H, Benaissa Trouw, B. J., Hesselink,W., Rijke, E., Schrier, C. and Hensen, E. J. (1992) Induction of anti- viral immune responses by immunization with recombinant-DNA encoded avian coronavirus nucleocapsid protein. Vaccine, 10: 119-124.
  2. Boursnell, M. E. G., Brown, T. D. K. and Foulds, I. J. (1987) Completion of the sequence of the genome of the coronavirus avian infectious bronchitis virus. J.Gen.Virol., 68: 55-77.
  3. Calnek, B.W. (1997) Diseases of poultry, 10th edn. Iowa State University., Press, Ames, IA .
  4. Cavanagh, D. (1983) Coronavirus IBV structural characterization of the spike protein. J. Gen. Virol., 64: 2577-2583.
  5. Chen , H., Cook, B., Attree, S. and Hiscox, A. (2003) Evaluation of a nucleoprotein based enzyme-linked immunosorbent assay for the detection of antibodies against infectious bronchitis virus. Avian Pathology, 32 (5): 519-526.
  6. Collisson, E.W., Pei, J., Dzielawar, J. and Seo, S. H. (2000) Cytotoxic T lymphocytes are critical in the control of infectious bronchitis virus in poultry. Developmental and Comparative Immunology, 24: 187-200.
  7. Cook, J.K.A. (1984) The classification of new serotypes of infectious bronchitis virus isolated from poultry flocks in Britain between 1981 and 1983: Avian Pathology, 13:733-741.
  8. Crowther J. R. (2001) Methods in molecular biology: The ELISA Guidebook.Humana Press, Totowa, New Jersey, Inc. NY 405 p.
  9. Gelb, J. Jr., Perkins, B. E., Rosenberger, J. K. and Alien, P. H. (1981) Serological and cross-protection studies with several infectious bronchitis virus isolates from Delmarva reared broiler chicken respiratory tracts of the chicken inoculated with Infectious bronchitis virus. Avian Dis., 25: 655-666.
  10. Hofstad, M.S. (1981) Cross-immunity in chickens using seven isolates of avian infectious bronchitis virus. Avian Diseases, 25: 650-654.
  11. Hofstad, M. S. (1984) In: Diseases of poultry. 8th edn. Iowa State University Press, Ames, Iowa, USA, 429-443.
  12. Kaul, M., Kadam, M., Malik, Y.P.S. and Tiwari, A.K. (2009) Nucleocapsid gene sequence analysis and characterization ofan Indian isolate of avian infectious bronchitis virus. Int. J. Poult. Sci., 8: 493-499.
  13. Lambrechts, C., Pensaert, M. and Ducatelle, R. (1993) Challenge experiments to evaluate cross-protection induced at the trachea and kidney level by vaccine strains and Belgian nephropathogenic isolates of avian infectious bronchitis virus. Avian Pathol., 22: 577-590.
  14. Lee, C. and Jackwood, M. W. (2001) Origin and evoluation of Georgia 98 (GA98), a new serotype of avian infectious bronchitis virus. Virus Research, 80: 33-39.
  15. Liu S., Chen, J. J., Chen, J., Kong, X., Shao, Y., Han, Z., Fegn, L., Cai, X., Gu, S. and Liu, M. (2005) Isolation of avian infectious bronchitis coronavirus from domestic peafowl (Pavocristatus) and teal (Anas). J. Gen. Virol., 86 (3): 719-725.
  16. Naqi, S.A., Karaca, K., Bauman, B. (1993) A monoclonal antibody-based antigen capture enzyme-linked immunosorbent assay for identification of infectious bronchitis virus serotypes. Avian Pathol. 22(3):555-64.
  17. Ndifuna, A., Waters, A.K., Zhou, M. and Collisson, E. W. (1998) Recombinant nucleocapsid protein is potentially an inexpensive, effective serodiagnostic reagent for infectious bronchitis virus. J. Virol Methods, 70: 37-44.
  18. Sambrook, J. and Russel DW (2001) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Pres.; A8.40-A8.54.
  19. Schalk, A. F. and Hawn, M. C. (1931) An apparently new respiratory disease of baby chicks .J. Am. Vet. Med. Asso., 78 :413-422.
  20. Seo, S. H., Wang, L., Smith, R. and Collison, E. W. (1997) The carboxyl–terminal 120-residue polypeptide of infectious bronchitis virus nucleocapsid induces cytotoxic T-lymphocytes and protects chicken from acute infection. J. Virol., 71 (10): 7889-7894.
  21. Singh, A. (2002) A laboratory manual of veterinary immunology and serology, Inc., NY; pp. 22-24.
  23. Stern, D. F., Burgess, L. and Sefton, B.M. (1982) Structural analysis of virion proteins of the avian coronavirus infectious bronchitis virus. J. Virol., 42: 208-219.
  24. Verma, K. C. (1964) Studies on infectious bronchitis virus. M.V.Sc. Thesis submitted to Agra University.
  25. Wadey, C. N. and Westaway, E. J. (1981) Structural proteins and glycoproteins of infectious bronchitis virus particles labeled during growth in chick embryo cells. Intervirology, 15: 19-27.
  26. Williams, A. K., Wang, L., Sneed, L. W., Collison, E. W. (1992) Comparative analyses of the nucleocapsid genes of several strains of Infectious bronchitis virus and other coronaviruses. Virus Res., 25: 211-222.
  27. Zanella, A., Lavazza, A., Marchi, R., MorenoMartin, A. and Paganell,i F. (2003) Avian infectious bronchitis: characterization of new isolates from Italy. Avian Dis., 47: 180-185.
  28. Zhang, D.Y, Zhou, J. Y., Fang, J., Hu, J. Q., Wu, J. X. and Mu, A. X. (2005) An ELISA for antibodies to infectious bronchitis virus based on nucleocapsid protein produced in Escherichia coli. Vet.Med-Czech, 50 (8): 336-344.
  29. Zhou, M.L., and Collison, E. W. (2000) The amino and carboxyl domains of the infectious bronchitis virus nucleocapsid protein interact with 3’ genomic RNA. Virus Res., 67: 31-39.
  30. Zhou, J. Y., Zhang, D. Y., Ye, J. X., Cheng, L. Q. (2004) Characterization of an avian infectious bronchitis virus isolated in China from chicken with nephritis.Jour.Vet Med B, inf. dis. and vet.pub.health, 51:147-152.
  31. Zimmermann, G.I., Nelson M.J., (1985) Diagnosis of ovine fasciolosis by Dot-ELISA: a rapid serodiagnosis technique. Am J Vet Res., 40: 1513-15.
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