Phylogenetic analysis of FAdV isolates
A phylogenetic tree was constructed based on the partial hexon gene of FAdV sequence alignment of the 46 isolates (Figure 6). The FAdV isolates were distinctly divided into five general clusters: FAdV-A, FAdV-B, FAdV-C, FAdV-D, and FAdV-E. The phylogenic tree’s topology indicated that two different FAdV genotypes with different genomic evolutions circulate in Kurdistan. The FAdV/Kurdistan/2013 and FAdV/Kurdistan/2020 belonged to FAdV-E close to USA isolates. On the other hand, the FAdV/Kurdistan/2015 belonged to FAdV-D closer to the Chinese FAdV isolate.
DISCUSSION
Iraqi Kurdistan contains more than 1300 poultry farms, and intensive farming and deficient control strategies have led to the spread of new viral infections. Thus, epidemiologic research is crucial to monitor disease outbreaks and develop vaccines. FAdV outbreaks have become a considerable concern for poultry farmers globally (Cizmecigil et al., 2020). IBH causes mortality resulting in production and economic losses. A higher number of FAdV clinical cases have been reported in recent years, and multiple FAdV strains have been isolated from sick birds in many countries (Mittal, Jindal, Tiwari, & Khokhar, 2014; Schachner et al., 2018). There is no published data about FAdV in Kurdistan chickens, reinforcing the requirement for molecular surveys on this emerging infectious agent in poultry and studying its fundamental role in clinical diseases. This study describes an IBH outbreak in Kurdistan/Iraq broiler flocks.
In this study, sudden high acute mortality of 8% to 15% started at two days to four weeks of age in affected broiler flocks. IBH can infect chickens of all ages; however, young chicks were more susceptible during the first two weeks, even when immunologically intact. There is an apparent age effect with avian adenoviruses. As the host’s age increases, the multiplication of the viruses within the host is restricted, and mortality decreases (Rahimi & Haghighi, 2015). There was no accumulation of straw-colored fluid in the pericardial sac, indicating no relation with HHS. This outcome is probably because of the different viral strains and serotypes of FAdV causing infection in this study, which agrees with previous studies (El-Tholoth & Abou El-Azm, 2019; Mittal et al., 2014). Mortality during IBH outbreaks is generally between 2% and 10% of the flock, but up to 30% has been described in case of coinfection with other immunosuppressive causative agents (El-Tholoth & Abou El-Azm, 2019). Typical necropsy observations such as an enlarged and pale liver, enlarged and hemorrhagic spleen and kidneys, and clinical findings were detected in this study’s IBH cases (Ahamad, Selvaraj, Sasikala, & BabuPrasath, 2016; Nakamura et al., 2011). Also, a pale and enlarged pancreas and greenish diarrhea were observed in some chicks, as reported by others (Ahamad et al., 2016).
The most critical histopathological finding in liver sections of diseased birds is the presence of intra-hepatocytic inclusion bodies. These are large basophilic intranuclear inclusion bodies surrounded by a pale halo (Anjum, Sabri, & Iqbal, 1989). Many hepatocytes were degenerated and appeared swollen with vacuolated cytoplasm. Infrequently, there was necrosis of some scattered hepatocytes. Moreover, there was lymphocytic and heterophilic hepatitis (Nakamura et al., 2011). These inflammatory infiltrates appear clearly around the central vein. Additionally, there was widening and infiltration of sinusoids with lymphocytes, heterophils, and histiocytes.
Histopathological sections of testicles from infected birds revealed degeneration of spermatozoa, spermatids, Sertoli cells, and Leydig cells. In the interstitial tissue, there was intertubular hemorrhage and large vacuoles. Also, there was dilation of the seminiferous tubular lumens containing necrotic debris and devoid of spermatozoa. Lesions in the testicle are reported for the first time in the present study.
In the present study, the isolates’ phylogenetic analysis revealed the circulation of two different genotypes in Kurdistan. Both Kurdistan FAdV had different genomic evolution. The FAdV/Kurdistan/2013 and FAdV/Kurdistan/2020 belonged to FAdV-E close to USA isolates. On the other hand, the FAdV/Kurdistan/2015 was closer to the Chinese FAdV isolate, proposing possible virus spreading from these areas, perhaps through trades and primary breeders or breeder substitutes. Circulation of two genotypes of FAdV in Kurdistan may hurt the clinical disease because some strains of different serotypes can reproduce IBH and hydropericardium syndrome simultaneously (Zhao et al., 2015).
Immunosuppressors such as IBDV, chicken infectious anemia virus (CIAV), and Marek’s disease virus (MDV) are factors that facilitate IBH outbreaks propagation or worsen clinical manifestations of FAdV infections (Morshed, Hosseini, Langeroudi, Fard, & Charkhkar, 2017; Niczyporuk, Woźniakowski, Samorek-Salamonowicz, & Czekaj, 2013). However, several studies showed that IBH might occur as a primary disease (Gomis et al., 2006; Ojkic et al., 2008).
CONCLUSION
This study is the first molecular characterization and histopathological examination of FAdVs in Sulaymaniyah broiler farms. FAdVs could be emerging infectious agents in Kurdistan poultry flocks, causing severe disease in young chicks. More research is needed to evaluate fowl adenoviral serotypes’ prevalence and pathogenicity in poultry flocks in Iraq. Identification of FAdV serotypes is essential in epidemiological studies of the disease outbreaks, development of preventative measures, and adoption of vaccination strategies. There is also a need for future development in molecular methods to identify adenovirus strains’ origins in IBH.