Prevalence and Molecular Characterization of FAdV-4 Isolated in Fujian Province, Southeast China During 2023-2025

Fowl adenovirus (FAdV) serotype 4 (FAdV-4) is a significant pathogen causing hepatitis-hydropericardium syndrome (HHS) in poultry, which can cause severe economic losses to the global poultry industry. Molecular epidemiological investigation of FAdV-4 has not been conducted in Fujian province, southeast China. In this study, we collected 219 samples of liver exhibiting similar pathological features of hepatitis or hydropericardium syndrome (HPS)-like clinicopathology changes from Fujian province during 2023-2025, which were analyzed using PCR assays. Further isolation of the virus was performed, and complete genomic sequences of the isolates were obtained through next-generation sequencing technology. Genetic evolution analysis of the gene sequences was conducted to elucidate the prevalence, molecular characteristics, and genetic variations of FAdV-4 in Fujian province. The results indicated that among the 219 clinical samples, 65 tested positive, yielding a positive rate of 29.68% (65/219). Positive samples from broilers were predominantly detected in the 21-120 days old, while those from layers were primarily concentrated in the 200-500 days old. The positive rate of Muscovy ducks infected with FAdV-4 was 4.57%. Ultimately, seven strains of FAdV-4 were isolated and designated as FJ08, FJ09, FJ20, FJ23, FJ33, FJ35, and FJ45. The preliminary challenge assay showed that all seven isolates were highly virulent in 7 days old SPF chickens, resulting in the death of all chickens within 2-3 days, leading to a mortality rate of 100%. Comparative analysis of whole genome sequences revealed that the isolated FAdV-4 strains from Fujian province clustered within the major genetic lineage as the domestic FAdV-4 reference strains (e.g., JSJ13, HLJFAd15, and HB1510 strains) and exhibited the highest identity with the FAdV-4 BNC strain derived from black-necked cranes in Yunnan province. Notably, the Hexon genes of seven isolated strains were divided into two major lineages based on phylogenetic analysis. The FJ08, FJ09, and FJ23 strains clustered in the major lineage as representative strains of FAdV-4 from both domestic and international sources, whereas FJ20, FJ33, FJ35, and FJ45 strains formed an independent genetic lineage with the FAdV-10 AHFY19 strain. The prevalence analysis revealed that FAdV-4 has been circulating among commercial chickens, indigenous chicken breeds, and Muscovy ducks in Fujian province, southeast China, indicating that the FAdV-4 strains in Fujian province possess the potential for cross-host transmission. Moreover, the age of the infected hosts (layers) has extended to 500 days, and their genomic characteristics exhibit genetic diversity.