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Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus
Dissertation (MSc)--University of Pretoria, 2008.
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University of Pretoria
2013
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| _version_ | 1867613446616383488 |
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| access_status_str | Open Access |
| author2 | Koekemoer, J.J.O. |
| author_browse | Koekemoer, J.J.O. |
| author_facet | Koekemoer, J.J.O. |
| collection | Thesis |
| dc_rights_str_mv | © 2008 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, 2008. |
| format | Thesis |
| id | oai:repository.up.ac.za:2263/24622 |
| institution | University of Pretoria (South Africa) |
| last_indexed | 2026-06-10T12:36:17.013Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UPSpace — University of Pretoria Institutional Repository |
| publishDate | 2013 |
| publishDateRange | 2013 |
| publishDateSort | 2013 |
| 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:2263/24622 Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus Koekemoer, J.J.O. upetd@up.ac.za Modibedi, Lesego Gladys Bluetongue virus African horse sickness virus UCTD AHSV Dissertation (MSc)--University of Pretoria, 2008. African horsesickness is an infectious but non-contagious disease caused by an orbivirus belonging to the Reoviridae family. The disease is classified as notifiable by the OlE because of the potential severe economic consequences that can result from outbreaks. Bluetongue, an arthropod-transmitted disease of wild and domestic ruminants, is caused by the bluetongue virus, which is the prototype species of the genus Orbivirus. Bluetongue is also a notifiable disease because it can spread very rapidly in naive populations of susceptible livestock. Strict restrictions have been issued for the trade in animals and animal products from infected areas. In the present study, a duplex one-step real-time RT-PCR using the fluorogenic dye SYBR® Green I was developed for the specific detection and identification of AHSV and BTV in one reaction, using melting temperatures (Tm) to discriminate between the viruses. Two primers pairs were designed to bind to areas of homology within genome segment 7 (VP7) of AHSV and genome segment 5 (NS1) of BTV respectively. The duplex real-time RT -PCR based test utilizes single tube RT -PCR amplification in which AHSV and BTV primers were used simultaneously. The RT-PCR primers amplified 232 bp of genome segment 7 from all nine serotypes of AHSV and 79 bp of genome segment 5 from all 22 BTV serotypes that were tested. When both viruses were present, two melting peaks were simultaneously generated at 76.30°C and 80.04°C representative of BTV and AHSV amplification products respectively. Serogroup-specific products were amplified from dsRNA of field isolates of AHSV and BTV. dsRNA from EHDV and EEV failed to demonstrate either the 232 bp specific AHSV PCR product or the 79 bp specific BTV product. These results indicate that the duplex real-time RT-PCR could be a useful technique for detection of AHSV and BTV from isolated viral dsRNA. Veterinary Tropical Diseases unrestricted 2013-09-06T18:05:31Z 2009-05-18 2013-09-06T18:05:31Z 2009-04-29 2011-05-18 2009-05-13 Dissertation Modibedi, LG 2008, Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus, MSc dissertation, University of Pretoria, Pretoria, viewed yymmdd < http://hdl.handle.net/2263/24622 > E1341/ag http://hdl.handle.net/2263/24622 http://upetd.up.ac.za/thesis/available/etd-05132009-163340/ © 2008 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 | Bluetongue virus African horse sickness virus UCTD AHSV Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus |
| title | Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus |
| title_full | Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus |
| title_fullStr | Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus |
| title_full_unstemmed | Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus |
| title_short | Development of a single real-time RT-PCR method for the group-specific identification of African horsesickness virus and bluetongue virus |
| title_sort | development of a single real time rt pcr method for the group specific identification of african horsesickness virus and bluetongue virus |
| topic | Bluetongue virus African horse sickness virus UCTD AHSV |
| url | http://hdl.handle.net/2263/24622 http://upetd.up.ac.za/thesis/available/etd-05132009-163340/ |