Shotgun metagenomic sequencing and genetic characterization of four enteric viruses in a dairy farm suffering a diarrhea outbreak of undetermined cause in adult cattle.

Diarrhea is an acute disease of cattle that has been extensively studied in calves; however, it has been far less investigated in adult cattle, despite its significant impact on milk production and animal health. In this study, we investigated an outbreak in a dairy herd in Uruguay for which initial diagnostic testing for the main known pathogens yielded negative results. Therefore, the outbreak was further investigated in depth using shotgun metagenomic sequencing, molecular virology assays and viral isolation. Of the three fecal samples from affected cows that were submitted for diagnostic testing and were negative for the main enteric pathogens, including BCoV, two were positive for bovine torovirus (BToV) by RT-qPCR. Next-generation sequencing-based virome analysis revealed over 3,300 viral operational taxonomic units (vOTUs), predominantly bacteriophages, but also eukaryotic viruses from 11 families. The complete genome of BToV was obtained and a putative novel protoparvovirus species was characterized. In addition, two distinct enteroviruses belonging to the species Enterovirus fitauri and Enterovirus idromi were identified. Notably, Enterovirus idromi had previously been reported exclusively in dromedaries; its detection in cattle expands the known host range of this species. Furthermore, the Enterovirus idromi strain appears to represent an interspecies recombinant enterovirus. These results highlight the importance of emerging approaches such as next-generation sequencing for identifying novel pathogens in cattle diseases and may serve as a basis for evaluating their potential role in disease pathogenesis, as well as for incorporating them into routine initial diagnostic testing.