Tropical Animal Health and Production

, Volume 43, Issue 2, pp 331–338 | Cite as

Preparation and evaluation of chicken embryo-adapted fowl adenovirus serotype 4 vaccine in broiler chickens

  • Muhammad Khalid MansoorEmail author
  • Iftikhar Hussain
  • Muhammad Arshad
  • Ghulam Muhammad
Original Research


The current study was planned to develop an efficient vaccine against hydropericardium syndrome virus (HSV). Currently, formalin-inactivated liver organ vaccines failed to protect the Pakistan broiler industry from this destructive disease of economic importance. A field isolate of the pathogenic hydropericardium syndrome virus was adapted to chicken embryos after four blind passages. The chicken embryo-adapted virus was further serially passaged (12 times) to get complete attenuation. Groups of broiler chickens free from maternal antibodies against HSV at the age of 14 days were immunized either with 16th passage attenuated HSV vaccine or commercially formalized liver organ vaccine. The antibody response, measured by enzyme-linked immunosorbent assay was significantly higher (P < 0.05) in the group immunized with the 16th passage attenuated HSV vaccine compared to the group immunized with liver organ vaccine at 7, 14, and 21 days post-immunization. At 24 days of age, the broiler chickens in each group were challenged with 103.83 embryo infectious dose50 of pathogenic HSV and were observed for 7 days post-challenge. Vaccination with the 16th passage attenuated HSV gave 94.73% protection as validated on the basis of clinical signs (5.26%), gross lesions in the liver and heart (5.26%), histopathological lesions in the liver (1.5 ± 0.20), and mortality (5.26%). The birds inoculated with liver organ vaccine showed significantly low (p < 0.05; 55%) protection estimated on the basis of clinical signs (40%), gross lesions in the liver and heart (45%), histopathological lesions in the liver (2.7 ± 0.72), and mortality (35%). Birds in the unvaccinated control group showed high morbidity (84%), mortality (70%), gross (85%), and histopathological lesions (3.79 ± 0.14) with only 10% protection. In conclusion, this newly developed HSV vaccine proved to be immunogenic and has potential for controlling HSV infections in chickens.


Hydropericardium syndrome Fowl adenovirus serotype 4 Attenuated vaccine Hexon gene 



Hydropericardium syndrome


Hydropericardium syndrome virus


Fowl adenovirus 4


Chorioallantoic membrane



We acknowledge the financial support from the Pakistan Science Foundation under the project PSF #P-AU/Bio (367) entitled “Preparation of an in-vitro passaged live attenuated hydro-pericardium syndrome virus (local isolates) vaccine” is greatly appreciated. The authors are also thankful to the director of the Centre of Agricultural Biochemistry and Biotechnology for providing facilities for the present study. Technical assistance of Dr. Faisal Saeed Awan is highly acknowledged.


  1. Abdul-Aziz, T.A. and Al-Attar, M. A., 1991. New syndrome in Iraqi chicks, Veterinary Record, 129, 272CrossRefPubMedGoogle Scholar
  2. Afzal, M. and Ahmad, I., 1990. Efficacy of an inactivated vaccine against hydropericardium syndrome in broilers,Veterinary Record, 126, 59–60PubMedGoogle Scholar
  3. Ahmad, K., Malik, M. I., Iqbal, K., Ahmad, K. and Naz, S., 1990. Efficacy of formalized liver organ vaccine against angara disease in broiler, Veterinary Archive, 63, 131–138Google Scholar
  4. Akhtar, M., Ahmad, R., Hayat, C. S, Hussain, I. and Ashfaque, M., 2000. Comparative immune response of formalin inactivated and binary ethyleneimine inactivated Angara disease vaccines, Pakistan Journal Biology Science, 3, 1313–1314CrossRefGoogle Scholar
  5. Attrache, J. and Villegas, P., 2001. Genomic identification and characterization of avian adenovirus associated with inclusion body hepatitis, Avian Diseases, 45, 780–787CrossRefPubMedGoogle Scholar
  6. Anjum, A. D., Sabri, M. A. and Iqbal, Z., 1989. Hydropericarditis syndrome in broiler chickens in Pakistan, Veterinary Record, 124, 247–248CrossRefPubMedGoogle Scholar
  7. Cheema, A. H., Afzal, M. and Ahmad, J., 1989. An adenovirus infection of poultry in Pakistan, Review of Science and Technology Office International des Epizooties, 8, 789–795Google Scholar
  8. Cowen, B. S., 1988. Chicken embryo propagation of type I avian adenoviruses, Avian Diseases, 32, 347–352CrossRefPubMedGoogle Scholar
  9. Duncan, D. B., 1955. Multiple range and multiple F tests, Biometrics,11, 1–42CrossRefGoogle Scholar
  10. Fadly, A. M. and Winterfield, R. W., 1975. Antigenic characterization of the inclusion body hepatitis virus, American Journal of Veterinary Research, 36, 532–534PubMedGoogle Scholar
  11. Fadly, A. M. and Nazerian, K., 1984. Efficacy and safety of a cell culture live virus vaccine for hemorrhagic enteritis of turkeys: laboratory studies, Avian Diseases, 28, 183–196CrossRefPubMedGoogle Scholar
  12. Fadly, A. M., Nazerian, K., Nagaraja, K. and Below, G., 1985. Field vaccination against hemorrhagic enteritis of turkeys by a cell culture live virus vaccine, Avian Diseases, 29, 768–777CrossRefPubMedGoogle Scholar
  13. Ganesh, K., Suryanarayana, V., Raghavan, R. and Gowda, R. N., 2001. Nucleotide sequence of L1 and part of P1 of hexon gene of fowl adenovirus associated with hydropericardium hepatitis syndrome differs with the corresponding region of other fowl adenoviruses, Veterinary Microbiology, 78, 1–11CrossRefPubMedGoogle Scholar
  14. Ganesh, K., Suryanarayana, V. and Raghavan, R., 2002. Detection of fowl adenovirus associated with hydropericardium hepatitis syndrome by a polymerase chain reaction, Veterinary Research Communication, 26, 73–80CrossRefGoogle Scholar
  15. Grewal, G. S., Sharma, S. N. and Deka, B. C., 1981. Inclusion body hepatitis in broiler chickens, Indian Journal of Poultry Science, 16, 51–56Google Scholar
  16. Grimes, T. M., King, D. J., Kleven, S. H. and Fletcher, O. J., 1977. Involvement of a type-8 avian adenovirus in the etiology of inclusion body hepatitis, Avian Diseases, 21, 26–33CrossRefGoogle Scholar
  17. Hitchner, S. B., 1980. Virus propagation in embryonating eggs. In: C. H. Domermuth, H. G. Purchase and J. E. Williams (eds), Isolation and Identification of Avian Pathogens. (American Association of Avian Pathologists, College Station, Texas), 120–121Google Scholar
  18. Hussain, I., Anjum, A. A. and Izhar-ul-Haq., 1996. An oil emulsion vaccine against hydropericardium syndrome in broiler chickens, Pakistan Veterinary Journal, 16, 125–128Google Scholar
  19. Hussain, I., Munir, R. and Ahmad, R., 1999. Evaluation and comparison of hydropericardium syndrome vaccines in broiler chicks, Pakistan Veterinary Journal, 19, 88–90Google Scholar
  20. Itakura, C., Yasuba, M. and Goto, M., 1974. Histopathological studies on inclusion body hepatitis in broiler chickens, Japanese Journal of Veterinary Science, 36, 329–340PubMedGoogle Scholar
  21. Kaur, A., Oberoi, M. S. and Singh, A., 1997. Neutralising antibody and challenge response to live and inactivated avian adenovirus-1 in broilers, Tropical Animal Health Production, 29, 141–146CrossRefGoogle Scholar
  22. Kawamura, H., Shimizu, F. and Tsubahara, H., 1964. Avian adenovirus: its properties and serological classification, National Institute of Animal Health Quarterly, 4,183–193PubMedGoogle Scholar
  23. Khan, A. A., Sabri, A. N., Mansoor, M. K. and Hussain, I., 2005. Hydropericardium syndrome in Pakistan: a review, World’s Poultry Science Journal, 8, 647–654CrossRefGoogle Scholar
  24. Kim, J. N., Byun, S. H., Kim, M. J., Kim, J. J., Sung, H. W. and Mo, I. P., 2009. Outbreaks of hydropericardium syndrome and molecular characterization of Korean fowl adenoviral isolates, Avian Disease, 50, 526–530Google Scholar
  25. Kumar, R., Chandra, R., Shukla, S. K., Agrawal, D. K. and Kumar, M., 1997. Hydropericardium syndrome in India: a preliminary study on the causative agent and control of the disease by inactivated autogenous vaccine, Tropical Animal Health Production, 29, 158–164CrossRefGoogle Scholar
  26. Lal, B., Maiti, N. K., Oberoi, M. S. and Sharma, S. N., 1992. An enzyme linked immunosorbent assay to detect antibodies against fowl adenovirus type-l, Indian Journal of Animal Science, 62, 35–37Google Scholar
  27. Maiti, N. K. and Sarkar, P., 1997. Humoral immune response of chicks to different clinical isolates of avian adenovirus type-1, Comparative Immunology Microbiology and Infectious Diseases, 20, 59–62CrossRefGoogle Scholar
  28. Mansoor, M. K., Hussain, I., Arshad, M., Muhammad, G., Hussain, M. H. and Mehmood, M. S., 2009. Molecular characterization of fowl adenovirus serotype 4 (FAV-4) isolate associated with fowl hydropericardium-hepatitis syndrome in Pakistan, Pakistan Journal of Zoology, 41, 269–276Google Scholar
  29. Manzoor, S. and Hussain, I., 2003. Reverse passive haemagglutination (RPHA) test for the detection and quantification of hydropericardium syndrome virus, Pakistan Journal of Life and Social Sciences, 1, 141–143Google Scholar
  30. McFerran, J. B., McCracken, R. M., Connor, T. J. and Evans, R. T., 1976. Isolation of viruses from clinical outbreaks of inclusion body hepatitis, Avian Pathology, 5, 315–324CrossRefPubMedGoogle Scholar
  31. Naeem, K., Niazi, T., Malik, S. A. and Cheema, A. H., 1995. Immunosuppressive potential and pathogenecity of an avian adenovirus isolate involved in hydropericardium syndrome in broilers, Avian Diseases, 39, 723–728CrossRefPubMedGoogle Scholar
  32. Nakamura, K., Mase, M., Yamaguchi, S. and Yuasa, N., 2000. Induction of hydropericardium in one day old specific pathogen free chicks by adenoviruses from inclusion body hepatitis, Avian Diseases, 44,192–196CrossRefPubMedGoogle Scholar
  33. Norby, E. and Wadell, G., 1969. Immunological relationships between hexon of certain human adenoviruses, Journal of Virology, 4, 663–670Google Scholar
  34. Nazerian, K and Fadly, A. M., 1982. Propagation of virulent and avirulent turkey hemorrhagic enteritis virus in cell culture, Avian Diseases, 26, 816–827CrossRefPubMedGoogle Scholar
  35. Ojkic, D. and Nagy, E., 2003. Antibody response and virus tissue distribution in chickens inoculated with wild-type and recombinant fowl adenoviruses, Vaccine, 22, 42–48CrossRefPubMedGoogle Scholar
  36. Roberts, M. M., White, J. L., Grutter, M. G. and Burnett, R. M., 1986. Three-dimensional structure of the adenovirus major coat protein hexon, Science, 232, 1148–1151CrossRefPubMedGoogle Scholar
  37. Russell, W. C., 2009. Adenoviruses: update on structure and function, Journal of General Virology, 90, 1–20CrossRefPubMedGoogle Scholar
  38. Sahidullah., Rahman, S., Rabbani, M., Shah, M. K., Naseem, S., Khan, S. H. and Muhammad, K., 2008. Development of standard protocols for preparation and evaluation of liver homogenate vaccines against hydropericardium syndrome virus in poultry, Pakistan Veterinary Journal, 28, 163–166Google Scholar
  39. Sambrook, J and Russell, D., 2001. Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NewYorkGoogle Scholar
  40. Schonewille, E., Singh, A.., T. Gobel, Gerner, W. W., Saalmuller, A.. and Hess, M., 2008. Fowl adenovirus (FAdV) serotype 4 causes depletion of B and T cells in lymphoid organs in specific pathogen-free chickens following experimental infection, Veterinary Immunology and Immunopathology, 121, 130–139CrossRefPubMedGoogle Scholar
  41. Schrenzel, M., Oaks, J. L. Rotstein, D. Maalouf, G. Snook, E. Sandfort, C. and Rideout, B., 2005. Characterization of a new species of adenovirus in falcons, Journal of Clinical Microbiology, 43, 3402–3413CrossRefPubMedGoogle Scholar
  42. Sheppard, M., McCoy, R. J. and Werner, W., 1995. Genomic mapping and sequence analysis of the fowl adenovirus serotype 10 hexon gene, Journal of General Virology, 76, 2595–2600CrossRefPubMedGoogle Scholar
  43. Toogood, C. I. A., Crompton, J. and Hay, R. T., 1992. Antipeptide antisera define neutralizing epitopes on the adenovirus hexon, Journal of General Virology, 73, 1429–1435CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2010

Authors and Affiliations

  • Muhammad Khalid Mansoor
    • 1
    Email author
  • Iftikhar Hussain
    • 1
  • Muhammad Arshad
    • 1
  • Ghulam Muhammad
    • 2
  1. 1.Department of Microbiology, Faculty of Veterinary ScienceUniversity of AgricultureFaisalabadPakistan
  2. 2.Department of Clinical Medicine and Surgery, Faculty of Veterinary ScienceUniversity of AgricultureFaisalabadPakistan

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