Shotgun metagenomic diagnosis of unidentified pathogens in hepatic necrosis samples from Samgye chickens

Chicken infectious anaemia virus (CIAV), infectious bursal disease virus (IBDV), and Eimeria spp. are major immunosuppressive pathogens in chickens that predispose host to secondary infections, including Clostridium septicum-associated hepatic necrosis. In this case, shotgun metagenomic sequencing was applied to identify C. septicum that could not be isolated by traditional bacterial culture in Samgye chickens. Six 35-day-old Samgye chicken carcasses were submitted for disease diagnosis; histopathological examination, and bacterial and viral isolation/identification were performed. Pooled liver samples were subjected to shotgun metagenomic sequencing to identify microbial composition, virulence factors, and antimicrobial resistance genes. Samgye chickens exhibited dorsal dermatitis, hepatic necrosis, and splenomegaly. Histopathology revealed hepatic necrosis with bacterial colonies and lymphoid depletion. PCR detected CIAV, antigenic variant IBDV, chicken astrovirus, and Eimeria, whereas bacterial culture yielded no growth. Shotgun metagenomic analysis identified C. septicum as the predominant bacterium, and CIAV as the dominant viral pathogen. The α-toxin and the antimicrobial resistance tetA(P) genes were detected in liver samples. This is the first report of concurrent CIAV, antigenic variant IBDV, Eimeria spp., and C. septicum infection from Samgye chickens in South Korea, suggesting that immunosuppressive infections may predispose chickens to C. septicum-associated hepatic necrosis and highlighting the diagnostic utility of shotgun metagenomic sequencing.