Pathogenicity and amino acid sequences of hemagglutinin cleavage site and neuraminidase stalk of differently passaged H9N2-avian influenza virus in broilers

Abstract

Low pathogenic Avian Influenza (AI) virus has the ability to evolve to high pathogenic viruses resulting in significant economic losses in the poultry sector. This study aims at assessing the impact of H9N2 viral passaging in broilers and its relatedness to pathogenicity and amino acid (a.a) sequences of the hemagglutinin (HA) cleavage site and neuraminidase (NA) stalk. The original H9N2 AI virus (P0) was used to challenge ten-21 days old broilers. Individual recovery of H9N2 virus from homogenates of trachea, lungs and airsacs was attempted in 9 days old chicken embryos, as a conclusion of the first passage (P1). Tracheal isolates of H9N2 were passaged for a second (P2) and a third (P3) time in broilers, followed by a similar embryonic recovery procedure. The a.a. sequence of a part of HA1 cleavage site and Neuraminidase stalk were compared among the differently passaged viruses; an assessement of the relatedness of the determined a.a. sequences to the pathogenicity in broilers, based on frequency of mortality, morbidity signs, gross and microscopic lesions at 3 days post challenge with the P1, P2, and P3-H9N2, is concluded. An increase in certain morbidity signs and specific lesions was observed in P2- and P3-H9N2 challenged broilers compared to birds challenged with P1-H9N2. A conserved R-S-S-R amino acid sequence at the HA1 cleavage site was observed in the differently passaged H9N2, associated with a variability in the NA stalk-a.a sequences. The passaging of the low pathogenic H9N2 virus in broilers leads to a trend of increase in pathogenicity, manifested in higher frequency of morbidity signs, and of specific gross and microscopic lesions of the examined organs. This passaging was associated with a conserved a.a. sequence of the hemaglutinin cleavage site and a variability in the sequence of the neuraminidase stalk. A detailed study of the potential of the detected variability in the neuraminidase stalk of H9N2 in induction of a higher pathogenicity in broilers will be the subject of future investigations.

Share and Cite:

Shaib, H. , Cochet, N. , Ribeiro, T. , Nour, A. , Nemer, G. , Saade, M. and Barbour, E. (2011) Pathogenicity and amino acid sequences of hemagglutinin cleavage site and neuraminidase stalk of differently passaged H9N2-avian influenza virus in broilers. Advances in Bioscience and Biotechnology, 2, 198-206. doi: 10.4236/abb.2011.24030.

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1] Guo, Y.J., Krauss, S., Senne, D.A., Mo, I.P., Lo, K.S., Xiong X.P., Norwood, M., Shortridge, K.F., Webster R.G. and Guan Y. (2000) Characterization of the pathogenicity of members of the newly established H9N2 influenza virus lineages in Asia. Virology, 267, 279-288.
[2] Barbour, E.K., Sagherian, V.K., Sagherian, N.K., Dankar, S.K., Jaber, L.S., Usayran, N.N. and Farran, M. T. (2006) Avian Influenza outbreak in poultry in the Lebanon and transmission to neighbouring farmers and swine. Veterinaria Italiana, 42(2), 13-21.
[3] Banet-Noach, C., Perk, S., Simanov, L., Grebenyuk, N., Rozenblut, E., Pokamunski, S., Pirak, M., Tendler, Y. and Panshin, A. (2007) H9N2 influenza viruses from Israeli poultry: a five-year outbreak. Avian Diseases, (51), 290-296.
[4] Barbour, E.K., Shaib, H.A. and Rayya, E.G. Reverse transcriptase-polymerase chain reaction-based surveillance of type A influenza viruses in wild and domestic birds of the Lebanon. Veterinaria Italiana, 43(1), 33-41.
[5] Pantin- Jackwood, M.J. and Swayne, D.E. (2009) Pathogenesis and pathobiology of avian influenza virus infection in birds. Revue Scientifique et Technique/ Office Internationale des Epizooties, 28(1), 113-136.
[6] Ito, T., Goto, H., Yamamoto, E., Tanaka, H., Takeuchi, M., Kuwayama, M., Kawaoka, Y. and Otsuki, K. (2001) Generation of a highly pathogenic avian influenza A virus from an avirulent field isolate by passaging in chickens. Journal of Virology, 75, 4439- 4443.
[7] Shaib, H.A., Cochet, N., Ribeiro, T., Abdel Nour, A.M., Nemer, G. and Barbour, E.K. (2010) HImpact of embryonic passaging of H9N2 virus on pathogenicity and stability of HA1- amino acid sequence cleavage siteH. Medical Science Monitor, 16(10), 333-337.
[8] Swayne, D.E. and Halvorson, D.A. (2008) Influenza. In: Saif, Y.M., Barnes, H.J., Fadly, A.M., Glisson, J.R., McDougald, L.R. and Swayne, D.E. Eds., Diseases of Poultry, 12th Ed., Ames. Iowa, Iowa State University Press, 153-184.
[9] Ebrahimi, S.M., Nili, H. and Sohrabi, N. (2010) Histopathological evaluation of A/chicken/Iran/339/02(H9N2), an Iranian field isolate of influenza virus, on Japanese quail (Coturnix coturnix japonica). World Applied Sciences Journal, 9(2), 226-229.
[10] Lee, D.C.W., Mok, C.K.P., Law, A.H.Y., Peiris, M. and Lau, A.S. (2010) Differential replication of avian influenza H9N2 viruses in human alveolar epithelial A549 cells. Virology Journal, 7, 71-75.
[11] Kianizadeh, M., Pourbakhch, S.A., Toroghi, R. and Momayez, R. (2006) Pathogencitiy and haemagglutinin gene sequence analysis of Iranian avian influenza H9N2 viruses isolated during (1998-2001). Iranian Journal of Veterinary Research, 7(3), 37-41.
[12] Liu J., Okazaki, K., Shi W., Wu, Q.M., Mweene, A.S. and Kida, H. (2003) Phylogenetic Analysis of Neuraminidase gene of H9N2 influenza viruses prevalent in chickens in China during 1995-2002. Virus Genes, 27(2), 197-202.
[13] Pazani, J., Marandi, M.V., Ashrafihelan, J., Marjanmehr, S.H. and Ghods, F. (2008) Pathological studies of A/ chicken/ Tehran/ ZMT-173/99( H9N2) influenza virus in commercial broiler chickens of Iran. International Journal of Poultry Science, 7(5), 502-510.
[14] Mosleh, N., Dadras, H. and Mohammadi, A. (2009). Evaluation of H9N2 avian influenza virus dissemination in various organs of experimentally infected broiler chickens using RT-PCR. Iranian Journal of Veterinary Research, 10, 12-20.
[15] Gharaibeh, S. (2008) Pathogenicity of an avian influenza virus serotype H9N2 in chickens. Avian Diseases, 52, 106-110.
[16] Swayne, D.E., Senne, D.A. and Suarez, D.L. 2008. Avian influenza. In: A Laboratory Manual for the Isolation and Identification of Avian Pathogens, 5th ed. Kennett Square, PA: American Association of Avian Pathologists, 128-134.
[17] Banks, J., Speidel, E.C., Harris, P.A. and Alexander, D.J. (2000) HPhylogenetic analysis of influenza A viruses of H9 haemagglutinin subtype. HHAvian Pathology,H 33, 353-360.
[18] Altschul, S., Gish, W., Miller, W., Meyers, E.W. and Lipman, D.J. (1990) Basic local alignment search tool. Journal of Molecular Biology, 215(3), 403-410.
[19] Barbour, E.K., El-Hakim, R.G., Kaadi, M.S., Shaib, H.A., Gerges, D.D. and Nehme, P.A. (2006) Evaluation of the histopathology of the respiratory system in essential oil treated broilers following a challenge with Mycoplasma gallisepticum and/or H9N2 influenza virus. International Journal of Applied Research in Veterinary Medicine, 4(4), 293-300.
[20] Barbour, E.K., El-Hakim, R.G., Kaadi, M.S., Shaib, H., Gerges, D.D., Nehme, P.A. and Harakeh, S. (2006) Pathology of the respiratory system in essential oil-treated broilers following a challenge with mucosal bacterium and/or a haemagglutinating virus. Journal of the American Holistic Veterinary Medical Association, 25, 15-30.
[21] Kishida, N., Eto, M., Sunaga, Y. and Kida, H. (2004) Enhancement of pathogenicity of H9N2 influenza A viruses isolated from chicken in China by co-infection with Staphylococcus aureus and Haemophilus paragallinarum. International Congress Series, 263, 481-485.
[22] Garcia-Sastre, A. (2010) Influenza virus receptor specificity: disease and transmission. American Journal of Pathology, 176, 1584-1585.
[23] Fedson, D.S. (2009) Confronting the next influenza pandemic with anti-inflammatory and immunomodulatory agents: why they are needed and how they might work. Influenza and Other Respiratory Viruses, 3(4), 129–142.
[24] Ewbank, J. (2008) Innate immunity. In: Ewbank, J., Vivier, E., Eds., Totowa, N.J. Humana Press, pp. 458.
[25] Swaggerty, C., Kaiser, P., Rothwell, L., Pevzner, I.Y. and Kogut, M.H. 2006. Heterophil cytokine mRNA profiles from genetically distinct lines of chickens with differential heterophil mediated innate immune responses. Avian Pathology, 35(2), 102-108.
[26] Soltanialvar, M., Shoushtari, H., Bozorgmehrifard, M., Charkhkar S., and Eshratabadi, F. (2010) Molecular characterization of hemagglutinin and neuraminidase genes of H9N2 avian influenza viruses isolated from commercial broiler chicken in Iran. Journal of Biological Sciences, 10, 145-150.
[27] Senne, D.A. (1998) Virus propagation in embryonating eggs. In: A laboratory manual for the isolation and identification of avian pathogens, 4th ed., Swayne, D., Glisson, J.R., Jackwood, M.W., Pearson, J.E., Reed, W.M. Eds., American Association of Avian Pathologists, New Bolton Center, Kennett Square, PA, 235-247.
[28] Bano, S., Naeem, K. and Malik, S.A. (2003) Evaluation of pathogenic potential of avian influenza virus serotype H9N2 in chickens. Avian Diseases, 47, 817-822.

Copyright © 2024 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.