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Inclusion body hepatitis (IBH), hydropericardium syndrome, and gizzard erosion associated with fowl adenovirus (FAdV) infection have caused notable economic losses worldwide. In 2020, severe IBH was observed in a layer chicken farm in Hebei Province, China. Liver samples were collected from layer chickens with severe IBH and virus isolation was performed in LMH cells. DNA sequence and bioinformatics analyses were conducted to determine the phylogenetic relationship and the pathogenicity assay was conducted in specific-pathogen-free (SPF) chickens. HeB20 strain was isolated and identified as FAdV-8b, and the complete genome was successfully sequenced (GenBank No. OK188966). Although widespread recombination in clinical strains has been reported within FAdVs, HeB20 showed some novel characteristics, and did not show any recombination, highlighting that recombinant and non-recombinant FAdV-8b coexist in the clinic poultry industry. Finally, pathogenicity animal model of HeB20 was developed and showed severe IBH and 10% mortality. Collectively, a new FAdV-8b strain (HeB20) was isolated and responsible for the severe IBH in layer chickens. Complete genome of HeB20 was sequenced and valuable for future epidemiological investigations. HeB20 was capable of inducing severe IBH and 10% mortality in SPF chickens; this animal model provides a powerful tool for the future vaccine development. [ABSTRACT FROM AUTHOR]