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Title: The molecular pathogenesis of ovine herpesvirus 2
Author: Rosbottom, Jane
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2003
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Malignant catarrhal fever (MCF) is a lymphoproliferative and inflammatory disease of cattle, deer and other ruminants. It is caused mainly by one of two herpesviruses, alcelaphine herpesvirus 1 (A1HV-1) and ovine herpesvirus 2 (OvHV-2). A1HV-1 can be grown in tissue culture; its complete genome has been sequenced and the virus characterised as a gammaherpesvirus. OvHV-2 cannot be grown in tissue culture and only a small amount of OvHV-2 genome sequence is known. Lymphoblastoid cell lines can be propagated from the tissues of cattle, deer and experimentally infected rabbits with MCF. Some of these cell lines transmit disease. The initial aim of this project was to further characterise these cell lines and to use them as a source for cloning the OvHV-2 viral genome. g the OvHV-2 viral genome. The OvHV-2 infected cell lines were found to contain on average 50 copies of viral genome per cell. Gardella gel analysis of OvHV-2 infected rabbit lines showed evidence of substantial levels of viral replication, whilst analysis of an OvHV-2 infected bovine cell line showed the virus genome to be predominantly in the latent form. Analysis of RNA extracted from OvHV-2 infected rabbit cell lines demonstrated evidence of early (ORF 57) and late (ORF 75) gene expression. This prompted examination of cell lysates from OvHV-2 infected rabbit cells, resulting in the first visualisation of an OvHV-2 capsid. A cosmid library was constructed using genomic DNA extracted from the OvHV-2 infected cattle cell line BJ/1035. Screening the cosmid library led to the generation of overlapping cosmids spanning most of the viral genome. The virus sequence was further extended using splinkerette PGR. Analysis of the viral sequence revealed that the OvHV-2 genome is similar to that of A1HV-1 and the unique "A"genes of A1HV-1 are conserved in OvHV-2. OvHV-2 also contains ORFs not present in the A1HV-1 genome, including an IL-10 homologue. The sequence derived is expected to be an invaluable resource for further study of OvHV-2 pathogenesis. The OvHV-2 ORFs 07 and 08 were contained in the cosmid sequence. It was hypothesised that the two sequences were spliced together to form a membrane glycoprotein of similar function to gp350/220 of EBV. Evidence of splicing of the 07/08 sequence was found in 07/08 transfected tissue cultured cells, however this iv splicing appeared incomplete and it was hypothesised that viral factors might be required for correct splicing. Immunofluorescence analysis of OvHV-2 infected rabbit cells using anti-08 immune sera showed evidence that a high proportion of OvHV-2 infected cells expressed 08. These studies provided a starting point for further investigation of this potentially important region of the OvHV-2 genome.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available