Multiple skin nodules, with histological features of adnexal tumours consistent with trichoepithelioma, were observed on the head and trunk of Syrian hamsters. Skin biopsies from 20 hamsters from five different colonies were affected, and two of the affected hamsters also had lymphoma. Two owners reported that 16 of 70 hamsters and 50 of 100 hamsters in their colonies had similar skin lesions. These tumours have previously been associated in laboratory colonies with hamster polyomavirus (HaPv) infection. Examination of skin tissues by electron microscopy failed to reveal intranuclear virus particles. Using recombinant major capsid protein vP1 of HaPV, vP1-specific antibodies were detected in sera from 12 of 12 affected hamsters and in four of four unaffected in-contact hamsters, by ELISA. The ELISA data were verified by immunoblot analysis. Eleven of 13 serum samples contained antibodies which reacted with at least one recombinant structural HaPv protein (VP2), including samples from three in-contact unaffected hamsters. Nine of the 11 anti-vP2-positive samples also reacted with recombinant VP3 of HaPV, and six readed with vP1. Amplification by PCR and sequencing detected vp1-encoding sequences showing a high degree of homology with HaPV. The findings suggest a possible infection by HaPV or a HaPv-like virus and it is likely that such an infection was enzootic within the affected colonies.