Equine herpesvirus 4 infected domestic horses associated with Sintashta spoke-wheeled chariots around 4,000 years ago

Equine viral outbreaks have disrupted the socio-economic life of past human societies up until the late 19th century, and continue to be of major concern to the horse industry today. With a seroprevalence of 60-80%, equine herpesvirus 4 (EHV-4) is the most common horse pathogen on the planet. Yet, its evolutionary history remains understudied. Here, we screen the sequenced data of 264 archaeological horse remains to detect the presence of EHV-4. We recover the first ancient EHV-4 genome with 4.2X average depth-of-coverage from a specimen excavated in the Southeastern Urals and dated to the Early Bronze Age period, approximately 3,900 years ago. The recovery of an EHV-4 virus outside of the upper respiratory tract not only points to an animal particularly infected, but also highlights the importance of post-cranial bones in pathogen characterisation. Bayesian phylogenetic reconstruction provides a minimal time estimate for EHV-4 diversification to around 4,000 years ago, a time when modern domestic horses spread across the Central Asian steppes together with spoke-wheeled Sintashta chariots, or earlier. The analyses also considerably revise the diversification time of the two EHV-4 subclades from the 16th century based solely on modern data to nearly a thousand years ago. Our study paves the way for a robust reconstruction of the history of non-human pathogens and their impact on animal health.