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Title: Analysis of the in vivo role of the M4 gene of murine gammaherpesvirus-68
Author: Townsley, A. C.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2004
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Murine gammaherpesvirus-68 (MHV-68) represents a tractable animal model of gammaherpesvirus pathogenesis as it is able to infect numerous murid rodent species, including laboratory mice, and replicate productively in a wide range of cell types in vivo, permitting efficient manipulation of the viral genome. MHV-76, a related gammaherpesvirus, is a deletion-mutant of MHV-68 and lacks 4 MHV-68-specific genes (M1-M4) and 8 viral tRNA-like sequences at the 5’-end of the genome. These genes are implicated in latency and/or immune evasion. Consequently, MHV-76 is attenuated during productive infection and the early stages of splenic latency, with respect to MHV-68. The aim of the project was the characterisation of the M4 gene of MHV-68. To elucidate the contribution M4 makes to in vivo pathogenesis, a novel MHV-76 mutant (MHV-76inM4), in which the region of MHV-68 coding for M4 and accompanying putative promoter elements was inserted into the 5’-region of the MHV-76 genome, was created. A revertant virus was subsequently generated (MHV-76.Rev) which restored the 5’-region of the MHV-76inM4 genome to that of MHV-76. Genomic rearrangements were confirmed by Southern analysis and sequencing. The growth of MHV-76inM4 in vitro was indistinguishable from that of MHV-76 and MHV-68. However, viral titres from MHV-76inM4-infected BALB/c mice were significantly increased with respect to MHV-76 at early times in the lung, suggesting an important role for M4 during productive infection. Additionally, at days 17 and 21 post-infection, there was a significant elevation in latent viral load in splenocytes of MHV-76inM4-infected mice compared to MHV-76, as measured by ex vivo reactivation assay and real-time PCR. Like MHV-76, MHV-76inM4 displays no evidence of overt spenomegaly, characteristic of MHV-68 infection at this time. M4 expression in vivo was detectable by RT-PCR during productive infection in the lung and during the establishment of latency in the spleen, but in general was not detectable during long-term latency.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available