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Pathogenesis of enzootic nasal tumor virus

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Title: Pathogenesis of enzootic nasal tumor virus
Author: Walsh, Scott
Department: Department of Pathobiology
Program: Pathobiology
Advisor: Wootton, Sarah
Abstract: Enzootic nasal tumor virus (ENTV) is a betaretrovirus of sheep (ENTV-1) and goats (ENTV-2) associated with neoplastic transformation of epithelial cells of the ethmoid turbinate. Confirmation of the role of ENTV in the pathogenesis of enzootic nasal adenocarcinoma (ENA) has yet to be resolved due to the lack of an infectious molecular clone and inability of the virus to propagate in cell culture. Very little is known about the prevalence of ENA, particularly in North America, and only one full length sequence was available for each of ENTV-1 and ENTV-2 at the initiation of this study. Serological screening has not been used for diagnostic purposes since reports have noted a lack of antibodies to the virus capsid in animals with ENA. In this thesis, I hypothesized that ENTV-1 is the causative agent of ENA and that envelope mediated entry is a determining factor in the tissue tropism of ENA. Ten full-length ENTV-1 provirus sequences were determined from naturally occurring ENA tumors of sheep from North America (ENTV-1NA). Overall, there was an unusually high degree of amino acid conservation among the isolates compared to other RNA viruses suggesting that ENTV-1 is under stabilizing selection and the bias towards synonymous mutations we observed in the viral coding sequences support this hypothesis. Induction of ENA in a healthy lamb was shown after inoculation with filtered tumor homogenate, thereby demonstrating the transmissible nature of ENA. Detection of ENTV-1 antigens and virions with betaretrovirion morphology in tissue homogenates from the experimentally induced ENA supports ENTV-1 as the causative agent of ENA. A molecular clone of ENTV-1 was generated but virus generated from the molecular clone was defective in protease processing unless complemented with JSRV Gag-Pro-Pol polyprotein and was unable to induce tumors in vivo. A highly specific and sensitive RT-PCR assay for detection of ENTV-1 genomic RNA in sheep nasal swab samples was developed based on the previously determined sequences. Antibodies reactive with the ENTV-1 envelope and capsid antigens were detected in sheep affected with ENA as well as in sheep kept in proximity to diseased animals, but results of serology based diagnostic tests were inconsistent. Truncation of the cytoplasmic tail of the ENTV-1 envelope protein was shown to dramatically increase transduction of pseudotyped virions from two separate retroviral vector systems. In vivo transduction with lentivirus vectors pseudotyped with the truncated ENTV-1 envelope protein demonstrated that the envelope protein is not the restricting viral factor for determining the tissue tropism of ENA. In summary, I have shown that ENA is transmissible in sheep and that ENTV-1 is the causative agent of ENA but that induction of ENA is not the most common outcome of infection with ENTV-1.
URI: http://hdl.handle.net/10214/7808
Date: 2014-01


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