Molecular detection and descriptive epidemiology of fowl adenovirus infection in broiler chickens

Abstract

Fowl adenoviruses represent substantial viral pathogens distributed across poultry sectors globally, being etiologically linked with severe disease complexes that induce devastating economic losses in broiler production. This study was conducted to establish definitive molecular evidence of fowl adenovirus infection in commercial broiler chickens within the Diyala Governorate using an optimized real-time polymerase chain reaction framework and to mathematically model its regional distribution boundaries. For this purpose, a total number of 96 clinical liver tissue specimens were aseptically collected from 32-day-old broiler chickens displaying clinical indications of hepatic dysfunction across various private operations. Following tissue processing and lysis, viral genomic DNA was isolated using standard silica-membrane extraction matrices. Molecular screening was executed via a qualitative real-time PCR (qRT-PCR) platform utilizing specific oligonucleotide primers (L1-F: 5′-GACGGCATTTGGGGAGGAATT-3′ and L1-R: 5′-GCTGAGGATTGAGCTTCACCC-3′) targeting the hypervariable loop L1 region of the viral hexon gene, followed by a continuous post-amplification dissociation gradient analysis from 58°C to 95°C. Out of the total number of examined specimens, 20 samples returned positive diagnostic outcomes, establishing a regional viral infection prevalence rate of 20.8%. Descriptive statistical variance modeling successfully projected the true expected prevalence limits within a stable 95% confidence interval ranging from 12.7% to 28.9%, maintaining an estimated standard error of 4.14. Furthermore, a non-parametric Chi-square goodness-of-fit test demonstrated an intense, highly significant deviation from theoretical uniform frequencies (χ² = 32.66, df = 1, P < 0.001), determining a powerful localized infectious pressure across the regional poultry ecosystem despite the statistical predominance of negative samples (79.2%). The recorded kinetic amplification plots manifested rapid exponential fluorescence signal emergence between cycles 15 and 22, while melt curve evaluation demonstrated sharp, singular dissociation peaks centered at approximately 85°C, validating absolute amplicon specificity and confirming the total absence of primer-dimer artifacts or non-specific carryover products. These empirical data determine that fowl adenovirus circulation is firmly established at a substantial frequency within the regional poultry networks, confirming that real-time PCR remains an exceptionally rapid, high-fidelity, and dependable diagnostic screening tool for early epidemiological mapping.