Molecular diagnosis and adaptation of highly virulent infectious bursal disease virus on chicken embryo fibroblast cell

oleh: Yogender Singh, Kalpana Yadav, Vinod Kumar Singh, Rakesh Kumar

Format: Article
Diterbitkan: Veterinary World 2014-05-01

Deskripsi

Aim: To collect Infectious bursal disease virus (IBDV) infected bursae based on postmortem findings of clinical samples followed by molecular confirmation and adaptation on chicken embryo fibroblast cell. Material and Methods: Enlarged bursae were processed to make 10% suspension in phosphate buffer saline and were used for viral RNA isolation to carryout VP2 gene fragment amplification using RT-PCR technique. Suspension was also used for adaptation of IBDV on chicken embryo fibroblast cells prepared from 11 day old chicken embryos. Infective titer of virus was calculated using Reed and Muench method. Results: Fifteen dead birds suspected of IBD infection were collected from different local poultry farms of Jabalpur (Madhya Pradesh). After post-mortem, twelve enlarged bursae sample were used for total RNA isolation which was used for amplification of VP2 gene. Out of twelve, eleven were found positive for VP2 gene amplification. Virological characterization of PCR positive sample was done on chicken embryo fibroblast cell culture and various cytopathic effects like rounding of cells, cellular detachment and vacuolation were shown by five IBD field isolates after 48 hours on 4th passage. TCID50 per ml of the adapted virus on 4th passage at 48 hours after infection was 1.46 x 107 . Conclusion: VP2 gene amplification using RT-PCR technique is a specific target for IBDV detection. Passaging of highly virulent IBDV field isolates in cell culture leads to attenuation of virus which can be exploited as a cell culture adapted vaccine candidate.