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Comparative gene expression of different African horse sickness virus strains
Dissertation (MSc)--University of Pretoria, 2017.
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| Format: | Thesis |
| Language: | English |
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University of Pretoria
2018
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| _version_ | 1867613660740845568 |
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| access_status_str | Open Access |
| author2 | Van Staden, Vida |
| author_browse | Van Staden, Vida |
| author_facet | Van Staden, Vida |
| collection | Thesis |
| dc_rights_str_mv | © 2018 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. |
| description | Dissertation (MSc)--University of Pretoria, 2017. |
| format | Thesis |
| id | oai:repository.up.ac.za:2263/65884 |
| institution | University of Pretoria (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:39:41.079Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UPSpace — University of Pretoria Institutional Repository |
| publishDate | 2018 |
| publishDateRange | 2018 |
| publishDateSort | 2018 |
| publisher | University of Pretoria |
| publisherStr | University of Pretoria |
| record_format | dspace |
| source_str | UPSpace — University of Pretoria Institutional Repository |
| spelling | oai:repository.up.ac.za:2263/65884 Comparative gene expression of different African horse sickness virus strains Van Staden, Vida u04281438@tuks.co.za Theron, Jacques De Wet, Carolina Elizabeth UCTD Dissertation (MSc)--University of Pretoria, 2017. Currently orbiviruses are recognised as causing worldwide emerging and re-emerging disease outbreaks including African horsesickness (AHS), a devastating disease of equids. AHS is endemic to sub-Saharan Africa, and of great economic importance in South Africa. AHS is a non-contagious arboviral disease with a mortality rate of up to 90% in susceptible horses. The disease is caused by African horsesickness virus (AHSV) and transmitted through species of the biting midge Culicoides. AHS can present with different disease severity and varying mortality rates, and to date no known correlation exist between specific serotypes and specific clinical forms. AHS disease pathogenesis is complex, and the result of an interaction of both viral and host factors. When using a tissue culture model system there are less variables, and one should theoretically be able to pinpoint viral genes or proteins that contribute to cellular cytopathic effect (CPE) with greater accuracy and ease. AHSV is a member of the Orbivirus genus, within the Reoviridae family, and is one of nine genera with a characteristic segmented double stranded RNA genome. This genome organisation allows the formation of genetic reassortants, which have assisted in revealing the functions of specific viral proteins. In a previous study eight AHSV strains, representing three different serotypes and five reassortant, were compared and differences observed with respect to virus release, effects on membrane permeability, cytotoxicity and cell viability, timing and extent of CPE, and induction of apoptosis. As the molecular basis for these differences were not fully understood, the aim of this study was to monitor different stages of the AHSV lifecycle of these strains to propose a cause for the diverse cytopathogenesis phenotypes observed. To accomplish this, systematic comparisons of gene expression studies on the three parental and five reassortant AHSV strains were done in both a mammalian and insect tissue culture system. This study set out to monitor different AHSV replication cycle stages, and focused mainly on i) transcription, ii) translation, iii) genome segment packaging and dsRNA synthesis, and iv) virus release since little was known about the discrepancies in replication kinetics between the different strains. In this investigation, varying degrees of CPE were observed in cultured mammalian cells upon infection with the different AHSV strains. Results suggested that a reassortant combination of genome segments 5 (NS1) and 10 (NS3) influenced the time of onset and severity of CPE observed in vitro in mammalian tissue cultures. Monitoring protein synthesis with Western blot assays, suggested probable interaction between the proteins encoded by AHSV-2 segments 5 and 10. When heterologous relative to the backbone genome, this reassortment combination produced a more severe cytopathic phenotype compared to the AHSV-2 parent. It may be hypothesised that the interaction between AHSV NS1 and NS3, when originating from the same parental strain, may play a role in the rate of onset of CPE and consequently influence the severity of cytopathicity in vitro, at least with respect to AHSV-2, -3 and -4. However, this was only observed when monitoring VP7 but not NS1 and NS2 protein synthesis levels. Moreover, similar findings were not observed when monitoring virus dsRNA synthesis levels or virus yield. Results further indicated no significant differences between the parental and reassortant strains when monitoring protein synthesis levels, dsRNA synthesis levels and virus yield in insect cell cultures. In our study the replication kinetics for the different reassortment progeny indicated changes in early replication stages only in infected mammalian cells. Although reassortant strains with an AHSV-2 NS1/NS3 recombination achieved changes early during virus replication, late virus replication stages remained unchanged. Results suggested that no specific step during virus replication was more efficient, as indicated by the parameters monitoring replication kinetics. We have also shown that replication kinetics in insect cells appear to be unchanged, at least during the replication steps monitored. This suggested that the cause for superior in vitro cytopathogenic strains in mammalian cells, but not in insect cells, grants merit for further investigation of host cell interactions and responses. Genetics MSc Unrestricted 2018-07-25T09:00:45Z 2018-07-25T09:00:45Z 2018/04/26 2017 Dissertation De Wet, CE 2017, Comparative gene expression of different African horse sickness virus strains, MSc Dissertation, University of Pretoria, Pretoria, viewed yymmdd <http://hdl.handle.net/2263/65884> A2018 http://hdl.handle.net/2263/65884 en © 2018 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. application/pdf University of Pretoria |
| spellingShingle | UCTD Comparative gene expression of different African horse sickness virus strains |
| title | Comparative gene expression of different African horse sickness virus strains |
| title_full | Comparative gene expression of different African horse sickness virus strains |
| title_fullStr | Comparative gene expression of different African horse sickness virus strains |
| title_full_unstemmed | Comparative gene expression of different African horse sickness virus strains |
| title_short | Comparative gene expression of different African horse sickness virus strains |
| title_sort | comparative gene expression of different african horse sickness virus strains |
| topic | UCTD |
| url | http://hdl.handle.net/2263/65884 |