Ahmed N. Flayyih
MSc student, Department of Pathology and Poultry Diseases, College of Veterinary Medicine, University of Mosul, Iraq.
Fanar A. Isihak
Department of Microbiology, College of Veterinary Medicine, University of Mosul, Iraq.
The infectious bursal disease virus (IBDV) remains one of the most important immunosuppression diseases of broiler chickens because of its targeting the bursa of Fabricius and its adverse effect on flock health and performance. This study aimed to characterize field IBDV infection in broiler chickens by combining molecular and immunohistochemical approaches. An investigation based on a cross-section of 70 broiler chickens aged 25–35 days from 10 commercial farms in Nineveh Governorate, Iraq, showing clinical suspicion of infectious bursal disease. Representative bursal samples were examined by conventional RT-PCR, RT-qPCR, sequencing of the VP2 gene, and immunohistochemistry using CD19 and CD57 markers. Conventional RT-PCR successfully amplified the expected VP2 fragment in 20 out of 28 bursal samples tested, with positive results found in 9 out of 10 farms. All samples were also confirmed positive for IBDV RNA by RT-qPCR, which corresponds to an overall positivity rate of 83.3% (n = 24). However, there were significant variations in RNA load within positive samples with Ct values ranging from 21 to 39. Melt curve analysis revealed a single sharp peak for all positive samples, which showed single peak melt curves with a specific melting temperature at 82.89°C. Phylogenetic analysis of four study isolates demonstrated separation into two distinct clusters, with two isolates grouping with Mosul-related reference sequences and two clustering with Egyptian reference strains, indicating genetic heterogeneity among circulating field viruses. Immunohistochemically, bursal tissues showed graded expression of both CD19 and CD57, ranging from negative to high immunoreactivity, reflecting variable immune-cell involvement and bursal immunopathological damage. In conclusion, the combination of RT-qPCR, conventional RT-PCR, VP2-based sequence analysis, and CD19/CD57 immunohistochemistry provided a comprehensive framework for confirming IBDV infection and characterizing the associated molecular and tissue-level changes in field cases.
