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Introduction: Fowl adenoviruses (FAdVs) are responsible for three economically significant diseases in broiler chicken flocks: Hepatitis-hydropericardium syndrome (HHS), inclusion body hepatitis (IBH), and adenoviral gizzard erosion. IBH has been reported in broiler flocks in northeastern Iran for several years; however, it was not considered a major threat due to its relatively minimal impact on flock performance and mortality compared to other diseases in the region. Following the diagnosis of the first HHS case in the country in 2021 in Khorasan Razavi Province, the dynamics of avian adenovirus infections in the area underwent significant changes.
Objectives: This study aimed to document the scenario of adenovirus infections in Khorasan Razavi Province from the first reported case in 2009 until 2021. Additionally, it sought to establish the serological baselines for FAdVs in broiler and layer breeder flocks following the implementation of vaccination programs in the province.
Materials and Methods: Using PCR or real-time PCR techniques, the FAdVs detection was conducted on tissue samples collected from flocks suspected of having IBH and HHS in the province between 2009 and 2021. In positive cases, nucleotide sequencing of the hexon gene loop 1 region was performed to identify the virus serotype. The serological status of breeder flocks was assessed before and after FAdV vaccination using a commercial ELISA kit.
Results: In March 2021, a commercial broiler flock in Bajestan County exhibited a 50% mortality rate, prompting postmortem examination, histopathology, molecular testing, and phylogenetic analysis to identify possible disease agents. Postmortem examination revealed typical gross lesions of HHS, including hydropericardium with an unusual accumulation of jelly-like, straw-colored fluid in the pericardial sac (without right ventricular failure); petechial or ecchymotic hemorrhages on the myocardium, myocardial valves, and endocardium; and discolored and mottled liver with small white foci and hemorrhages. Histopathological analysis revealed hepatocyte necrosis and basophilic inclusion bodies in the liver. Molecular tests for FAdVs, H5 avian influenza virus (AIV), Newcastle disease virus (NDV), infectious bronchitis virus (IBV), H9N2, chicken infectious anemia virus (CIAV), infectious bursal disease virus (IBDV), Marek’s disease virus (MDV), Ornithobacterium rhinotracheale (ORT), Mycoplasma gallisepticum (MG), and Mycoplasma synoviae (MS) returned positive results for FAdVs, CIAV, IBD vaccine virus, and IBV serotypes 793B and variant I. Phylogenetic analysis of the hexon gene loop 1 region revealed a FAdV serotype 4 (FAdV-4), which was identical to the Pakistani isolate PARC-1/98. Molecular detection of FAdVs in 46 flocks suspected of IBH from 2009 to 2020 in Khorasan Razavi confirmed the presence of FAdVs in 18 cases. Nucleotide sequencing of PCR products revealed that all samples belonged to serotypes 2 and 11, with no evidence of FAdV-4 during this period. Following the vertical spread of HHS in the second wave and horizontal transmission in the third wave across the province, 175 flocks suspected of disease were tested in 2021, with 104 (59%) testing positive for FAdV-4. Cross-sectional ELISA testing of broiler and layer breeder flocks before vaccination indicated varying levels of exposure to FAdVs. Post-vaccination serological analysis revealed varying degrees of vaccination success, along with field FAdVs challenges. The arithmetic mean titers (AMT) of ELISA results reached up to 12,000 in unchallenged vaccinated flocks with a coefficient of variation (CV) of less than 35%. In contrast, challenged vaccinated flocks exhibited serum titers ranging from 13,000 to 18,000, with a CV generally below 25%. The CV was a critical factor in evaluating vaccination success. Serum titers analysis of four broiler flocks affected by HHS at the slaughterhouse showed AMT, %CV, and vaccination index (VI) values of 9,290, 36.5, and 383, respectively. A direct correlation was observed between mortality rates and both AMT and CV.
Conclusion: The scenario of adenovirus infections in Khorasan Razavi Province began with the occurrence of IBH in 2009 and persisted for 11 years without a significant economic impact on the poultry industry. This period of relative stability was abruptly disrupted by the emergence of HHS in 2021, which has become a serious concern in the region. Although breeder flocks were successfully vaccinated, they were unexpectedly challenged by field FAdVs, which warrants further investigation. The ELISA assay proved to be an effective tool for evaluating vaccination success and identifying field challenges. The combined use of molecular tests and ELISA assays can significantly aid in interpreting results and implementing preventive and control measures for the disease.
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