Isolation and characterization of Bovine Gammaherpesvirus-4 (BoHV-4) in diarrheic sheep

While commonly detected in cattle, the presence of Bovine gammaherpesvirus 4 (BoHV-4) in small ruminants remains insufficiently examined. This study reports the isolation, identification, and genetic characterization of BoHV-4 from a sheep presenting hemorrhagic diarrhea. A fecal sample collected from an infected adult sheep was submitted to a diagnostic study using a real-time PCR panel for various bacteria, parasites, and viruses, yielding negative results for all the pathogens in the panel. Virus isolation was performed on MDBK, BHK-21, SFT-R cell lines, and primary lamb kidney cell culture (PLK), and cytopathogenic effects were observed in all the cell lines except for PLK. The virus was then identified through chloroform and halogen assays. The presence of BoHV-4 was validated by nested PCR targeting the thymidine kinase (TK) gene, and phylogenetic analysis classified the isolate as Genotype II, showing close genetic similarity with Argentinean strains. Serum neutralization assay demonstrated low cross-reactivity between BoHV-1 polyclonal serum and BoHV-4 isolate. This study reports the first detection of BoHV-4 genotype II in sheep and suggests its potential association with gastrointestinal disease. Further studies are needed to investigate the pathogenic role of BoHV-4 in small ruminants and its impact on various organ systems.