Intermittent shedding dynamics of EHV-1 across biological matrices during a natural outbreak linked to international equestrian events.

Equine herpesvirus type 1 (EHV-1) outbreaks associated with international equestrian events have demonstrated the capacity of this virus for rapid transboundary dissemination. However, detailed longitudinal data integrating viral shedding across multiple biological matrices within the same animals during naturally occurring outbreaks remain limited. In this study, ten horses hospitalized during a naturally occurring EHV-1 outbreak linked to an international jumping competition were monitored for 10-12 days, from which 306 nasal, ocular, urine and faecal samples were obtained and analysed by quantitative real-time PCR. Viral detection was strongly matrix-dependent, with nasal swabs showing the highest positivity (76.6%), persistence and viral load, whereas ocular (49.3%), faecal (34.6%) and urine samples (32.8%) displayed lower and more transient detection. Shedding was frequently intermittent within individual horses, with alternating positive and negative results across consecutive sampling days. Mixed-effects models demonstrated a significant decrease in both detection probability and viral load over time. Vaccination status was not significantly associated with nasal viral detection or viral load. However, vaccinated horses showed a significantly lower probability of lymphopenia, independently of viral load. These findings indicate that EHV-1 shedding during outbreaks is dynamic, intermittent, and compartment-dependent rather than continuous, which may complicate outbreak management strategies based on single negative PCR results. Although vaccination did not reduce measurable viral excretion in this cohort, it appeared to modulate systemic haematological responses. The results support the implementation of serial testing and sustained isolation measures during outbreak control, in high-density equestrian settings where rapid international dissemination may occur.