Evaluation of avian adenovirus inactivation methods used in the production of influenza vaccines

Inactivation of influenza virus and other potential contaminants like avian adenoviruses coming from embryonated chicken eggs is a critical step in the production of inactivated influenza vaccines. Inactivation must lead to a guaranteed reduction in contaminant titers by at least 4 lg (PFU)/ml. The aim of this study was to identify an optimum cell line for adenovirus propagation and to estimate a reduction in adenovirus titers in vaccine intermediates after inactivation. In a series of experiments, we identified the optimum conditions and the optimum cell line for the propagation of avian adenovirus (strains CELO and Fontes). The most commonly used inactivation methods were analyzed, including inactivation by β-propiolactone and UV light. Viral titers were measured by plaque assays. After 10 h of inactivation with β-propiolactone, CELO titers fell by 4.12 ± 0.06 lg, whereas Fontes titers, by 4.20 ± 0.19 lg, suggesting that β-propiolactone is an effective inactivating agent. Exposure to UV light led to a reduction in CELO titers by 4.69 ± 0.89 lg and a reduction in Fontes titers by 4.44 ± 1.06 lg after 5 min. N-octyl-β-D-glucopyranoside added at the splitting step reduced CELO titers by 0.93 ± 0.15 lg and Fontes titers by 1.04 ± 0.12 lg, whereas tetradecyltrimethylammonium bromide led to a reduction in CELO and Fontes titers by 1.18 ± 0.17 lg and 1.12 ± 0.38 lg, respectively.

Molecular characterization of pathogenic and nonpathogenic fowl aviadenovirus serotype 11 isolates

Fowl aviadenoviruses, many of which are of importance in veterinary medicine, are classified into 5 species. In this study, a pathogenic isolate and a nonpathogenic isolate of fowl aviadenovirus serotype 11 (FAdV-11) of species Fowl aviadenovirus D were characterized. Growth rates were analyzed for the 2 isolates, showing notable differences. The complete genomic sequences of the viruses were fully determined and were analyzed. The genomes of the 2 isolates showed 98.1% sequence identity and revealed 6 nonsynonymous mutations between the Ontario isolates. Two of the 6 mutations were also found in the sequences of recently published pathogenic Chinese fowl aviadenovirus 11 isolates, suggesting potential molecular markers that could be associated with pathogenesis. Deletions were found in the L5 region within the overlapping coding sequences for the 100, 22, and 33 kDa proteins, and these were found in only the nonpathogenic isolates. This molecular pattern was identified in FAdV-9, another nonpathogenic FAdV-D species virus. Furthermore, the tandem repeat regions varied dramatically; the pathogenic isolates contained a reduced number of tandem repeats compared with the nonpathogenic isolates. Lastly, a protein produced early in infection was analyzed using bioinformatics to determine its role in disease. This study highlights several candidate molecular determinants of avian adenovirus genomes related to pathogenicity.

Detection of enteric viruses in pancreas and spleen of broilers with runting-stunting syndrome (RSS)

Enteric disease is a multifactorial problem in chickens, which causes gastrointestinal alterations, elevated feed conversions and impairment. In the last years, several enteric viruses were implicated in enteric disease; case reports have shown their presence alone or in concomitant infections during outbreaks and have suggested that they might be determining factors in the aetiology of enteric disease. This study shows high detection rates of enteric viruses in the pancreas and spleen in samples from an outbreak of enteritis and malabsorption in 16 chicken flocks (n=80 broilers). Avian nephritis virus (ANV) was the most ubiquitous virus, present in 75% of the flocks followed by avian rotavirus group A (ART-A) with 68.75%, and by chicken astrovirus (CAstV) and chicken parvovirus (ChPV) in 43.75% of samples. Viruses were present in the pancreas of positive flocks at extremely high rates: 100% for ART-A, 91.7% for ANV, 100% for CAstV and 57.14% for ChPV. By contrast, only 16.7% and 57.14% of intestine samples were positive for ANV and CAstV, respectively. Avian reovirus (AReo) and avian adenovirus group 1 (FAdV-1) were not detected. These results suggest that high viral detection rates in pancreas samples may be a result of viremia during enteric disease, with subsequent damage of the exocrine pancreas, leading to runting-stunting syndrome (RSS).