The purpose of this experiment was to study the duration and distribution of equine influenza virus in actively infected ponies over a 3 wk period. Pony foals (6-8 mo old) were infected experimentally by nebulizing equine influenza subtype-2 virus ultrasonically through a face mask. Successful infection was clinically apparent as each of the foals (n = 6) had a febrile response, a deep hacking cough and mucopurulent nasal discharge for 7 to 10 d. The virus was isolated from nasopharyngeal swabs of all the ponies 3 and 5 d after infection and all the ponies seroconverted to the virus. Samples were taken from the nasopharynx, mid-trachea and the mainstem bronchus with cytology brushes through an endoscope as well as from bronchoalveolar lavage fluid. On days 3 to 7 post-infection, ciliacytophtorea (the presence of cilia and ciliated plates separated from columnar epithelial cells) was recognized on routine cytological stain. Indirect immunoperoxidase staining utilizing polyclonal antibodies demonstrated viral antigen in intact and fragmented ciliated epithelial cells and in fragments of ciliated plates. The infected cells and cell fragments were particularly evident on days 3 and 5 post-infection in the nasopharynx, mid-trachea and mainstem bronchus and on days 3 to 7 post-infection in the bronchoalveolar lavage samples. On days 7 and 21 post-infection, viral antigen was identified in vacuoles of alveolar macrophage-like cells collected by bronchoalveolar lavage. It can be concluded from this study that equine influenza virus can infect not only the upper airways but also the bronchial epithelium and that viral antigen can persist up to 21 d post-infection.