Characterization of enzootic nasal tumor virus 1–associated enzootic nasal adenocarcinoma in sheep and development of a tumor-derived organoid model

Enzootic nasal adenocarcinoma (ENA) is a contagious neoplasm of the ethmoid turbinate mucosa in sheep, caused by enzootic nasal tumor virus 1 (ENTV1; family <i>Retroviridae</i>, unclassified <i>Betaretrovirus</i>). We report an outbreak of ENTV1-associated ENA in a sheep flock in South Dakota, USA. Affected animals had dyspnea, unilateral nasal discharge, and progressive weight loss. Postmortem examination revealed unilateral nasal masses that were diagnosed histologically as invasive nasal adenocarcinoma. PCR amplification followed by Sanger sequencing of the <i>gag</i> and <i>env</i> gene regions confirmed the presence of ENTV1. Our proviral genome assembly via next-generation sequencing is only the second ENTV1 sequence submitted to GenBank from the United States. Our isolate clustered within the ENTV1 clade and was closely related to the reported U.S. and Canadian isolates, indicating a shared evolutionary lineage. To further investigate tumor biology, we established 3-dimensional organoids derived from the nasal adenocarcinoma, which maintained the histologic features of the primary tumor and tested positive for ENTV1. These organoids also had an invasive phenotype, demonstrating their potential utility as a novel in vitro model for studying ENA pathogenesis and evaluating therapeutic interventions.