Two divergent BoHV-1 isolates from yaks show distinct biological properties and neutralization profiles.

Bovine herpesvirus type 1 (BoHV-1) is a major cattle pathogen, yet its epidemiology and genetic diversity in yaks are poorly understood. In this study, we isolated and fully sequenced two BoHV-1 strains from yaks, namely BoHV-1/Yak/2025/2K (BoHV-1.1) and BoHV-1/Yak/2025/12 (BoHV-1.2), and confirmed their distinct lineages through phylogenetic analysis. Functional profiling revealed clear phenotypic differences: BoHV-1.1 replicated faster, targeted the lower respiratory tract, and caused more severe lung lesions in rabbits; BoHV-1.2 replicated preferentially in the upper respiratory and reproductive tracts, causing milder respiratory disease but higher genital viral loads. Neutralization assays showed strong homologous neutralization but weak cross-neutralization between subtypes, indicating limited cross-protection. These results provide the first experimental evidence of co-circulating, antigenically and phenotypically distinct BoHV-1 subtypes in plateau yaks, supporting subtype-specific surveillance and evidence-informed vaccine policy in endemic areas.