Experimental transmission of enzootic nasal adenocarcinoma in sheep

Abstract

Enzootic nasal adenocarcinoma (ENA) is a contagious neoplasm of the secretory epithelial cells of the nasal mucosa of sheep and goats. It is associated with the betaretrovirus, enzootic nasal tumor virus (ENTV), but a causative relationship has yet to be demonstrated. In this study, 14-day-old lambs were experimentally infected via nebulization with cell-free tumor filtrates derived from naturally occurring cases of ENA. At 12 weeks post-infection (wpi), one of the five infected lambs developed clinical signs, including continuous nasal discharge and open mouth breathing, and was euthanized. Necropsy revealed the presence of a large bilateral tumor occupying the nasal cavity. At 45 wpi, when the study was terminated, none of the remaining infected sheep showed evidence of tumors either by computed tomography or post-mortem examination. ENTV-1 proviral DNA was detected in the nose, lung, spleen, liver and kidney of the animal with experimentally induced ENA, however there was no evidence of viral protein expression in tissues other than the nose. Density gradient analysis of virus particles purified from the experimentally induced nasal tumor revealed a peak reverse transcriptase (RT) activity at a buoyant density of 1.22 g/mL which was higher than the 1.18 g/mL density of peak RT activity of virus purified from naturally induced ENA. While the 1.22 g/mL fraction contained primarily immature unprocessed virus particles, mature virus particles with a similar morphology to naturally occurring ENA could be identified by electron microscopy. Full-length sequence analysis of the ENTV-1 genome from the experimentally induced tumor revealed very few nucleotide changes relative to the original inoculum with only one conservative amino acid change. Taken together, these results demonstrate that ENTV-1 is associated with transmissible ENA in sheep and that under experimental conditions, lethal tumors are capable of developing in as little as 12 wpi demonstrating the acutely oncogenic nature of this ovine betaretrovirus.

Diagnostic pathology in microbial diseases of sheep or goats

“…The low degree of anisokaryosis, low mitotic rate and apparent absence of metastases define the tumour as a 'low grade adenocarcinoma'. Tumour cells express the envelope protein of the virus(es), which can be detected by immunohistochemistry (Walsh et al, 2013) and retrovirus-like particles can be observed in the tumours by electron microscopy (De las Heras et al, 2003b).…”

Construction of a molecular clone of ovine enzootic nasal tumor virus

“…2b lane 7 and 8 and 2C lane 3). Proteins of approximately 62, 57, 52 and 40 kDa in a banding pattern reminiscent of partially processed Gag [14, 23] were also detected (Fig. 2b, lane 8).…”

“…This was unexpected considering that the ENTV-1NA4 sequence was derived from nasal tumor tissue isolated from a sheep with ENA. Although only fully processed Gag was observed in virions purified from the ENTV-1NA4 tumor or any of the other ENTV-1NA tumors [14], a processing defect could exist outside the context of transformed sheep cells. Since ENTV-1OVC was extracted from a tumor transmitted by cell-free tumor homogenate [14], it was assumed that this sequence represented a replication-competent, infectious ENTV-1.…”

“…Enzootic nasal tumor virus (ENTV)-1 is an ovine betaretrovirus that is associated with enzootic nasal adenocarcinoma (ENA), a nasal tumor of sheep [14]. Experiments showing transmission of ENA to a healthy lamb by inoculation with cell-free ENA tumor homogenate containing ENTV-1 antigens suggested that ENTV-1 is the causative agent of ENA, but did not completely fulfil Koch’s postulates [14].…”

“…Experiments showing transmission of ENA to a healthy lamb by inoculation with cell-free ENA tumor homogenate containing ENTV-1 antigens suggested that ENTV-1 is the causative agent of ENA, but did not completely fulfil Koch’s postulates [14]. A factor limiting these experiments is an inability to produce high titer virus from cell culture since the virus cannot be propagated in vitro.…”