Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus
Bluetongue virus is the type species of the genus Orbivirus, family Reoviridae. Bluetongue viruses (BTV) are transmitted between their vertebrate hosts primarily by biting midges (Culicoides spp.) in which they also replicate. Consequently BTV distribution is dependent on the activity, geographic di...
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| Format: | Article |
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Public Library of Science
2016
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| Online Access: | https://eprints.nottingham.ac.uk/38905/ |
| _version_ | 1848795716947804160 |
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| author | Richard, Yolande Maan, Sushila Maan, Narender Singh Belaganahalli, Manjunatha N. Potgieter, Abraham C. Kumar, Vinay Batra, Kanisht Wright, Isabel M. Kirkland, Peter D. Mertens, Peter P.C. |
| author_facet | Richard, Yolande Maan, Sushila Maan, Narender Singh Belaganahalli, Manjunatha N. Potgieter, Abraham C. Kumar, Vinay Batra, Kanisht Wright, Isabel M. Kirkland, Peter D. Mertens, Peter P.C. |
| author_sort | Richard, Yolande |
| building | Nottingham Research Data Repository |
| collection | Online Access |
| description | Bluetongue virus is the type species of the genus Orbivirus, family Reoviridae. Bluetongue viruses (BTV) are transmitted between their vertebrate hosts primarily by biting midges (Culicoides spp.) in which they also replicate. Consequently BTV distribution is dependent on the activity, geographic distribution, and seasonal abundance of Culicoides spp. The virus can also be transmitted vertically in vertebrate hosts, and some strains/serotypes can be transmitted horizontally in the absence of insect vectors. The BTV genome is composed of ten linear segments of double-stranded (ds) RNA, numbered in order of decreasing size (Seg-1 to Seg-10). Genome segment 2 (Seg-2) encodes outer-capsid protein VP2, the most variable BTV protein and the primary target for neutralising antibodies. Consequently VP2 (and Seg-2) determine the identity of the twenty seven serotypes and two additional putative BTV serotypes that have been recognised so far. Current BTV vaccines are serotype specific and typing of outbreak strains is required in order to deploy appropriate vaccines. We report development and evaluation of multiple ‘TaqMan’ fluorescence-probe based quantitative real-time type-specific RT-PCR assays targeting Seg-2 of the 27+1 BTV types. The assays were evaluated using orbivirus isolates from the ‘Orbivirus Reference Collection’ (ORC) held at The Pirbright Institute. The assays are BTV-type specific and can be used for rapid, sensitive and reliable detection / identification (typing) of BTV RNA from samples of infected blood, tissues, homogenised Culicoides, or tissue culture supernatants. None of the assays amplified cDNAs from closely related but heterologous orbiviruses, or from uninfected host animals or cell cultures. |
| first_indexed | 2025-11-14T19:36:31Z |
| format | Article |
| id | nottingham-38905 |
| institution | University of Nottingham Malaysia Campus |
| institution_category | Local University |
| last_indexed | 2025-11-14T19:36:31Z |
| publishDate | 2016 |
| publisher | Public Library of Science |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | nottingham-389052020-05-04T18:09:58Z https://eprints.nottingham.ac.uk/38905/ Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus Richard, Yolande Maan, Sushila Maan, Narender Singh Belaganahalli, Manjunatha N. Potgieter, Abraham C. Kumar, Vinay Batra, Kanisht Wright, Isabel M. Kirkland, Peter D. Mertens, Peter P.C. Bluetongue virus is the type species of the genus Orbivirus, family Reoviridae. Bluetongue viruses (BTV) are transmitted between their vertebrate hosts primarily by biting midges (Culicoides spp.) in which they also replicate. Consequently BTV distribution is dependent on the activity, geographic distribution, and seasonal abundance of Culicoides spp. The virus can also be transmitted vertically in vertebrate hosts, and some strains/serotypes can be transmitted horizontally in the absence of insect vectors. The BTV genome is composed of ten linear segments of double-stranded (ds) RNA, numbered in order of decreasing size (Seg-1 to Seg-10). Genome segment 2 (Seg-2) encodes outer-capsid protein VP2, the most variable BTV protein and the primary target for neutralising antibodies. Consequently VP2 (and Seg-2) determine the identity of the twenty seven serotypes and two additional putative BTV serotypes that have been recognised so far. Current BTV vaccines are serotype specific and typing of outbreak strains is required in order to deploy appropriate vaccines. We report development and evaluation of multiple ‘TaqMan’ fluorescence-probe based quantitative real-time type-specific RT-PCR assays targeting Seg-2 of the 27+1 BTV types. The assays were evaluated using orbivirus isolates from the ‘Orbivirus Reference Collection’ (ORC) held at The Pirbright Institute. The assays are BTV-type specific and can be used for rapid, sensitive and reliable detection / identification (typing) of BTV RNA from samples of infected blood, tissues, homogenised Culicoides, or tissue culture supernatants. None of the assays amplified cDNAs from closely related but heterologous orbiviruses, or from uninfected host animals or cell cultures. Public Library of Science 2016-09-23 Article PeerReviewed Richard, Yolande, Maan, Sushila, Maan, Narender Singh, Belaganahalli, Manjunatha N., Potgieter, Abraham C., Kumar, Vinay, Batra, Kanisht, Wright, Isabel M., Kirkland, Peter D. and Mertens, Peter P.C. (2016) Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus. PLoS ONE, 11 (9). e0163014/1-e0163014/19. ISSN 1932-6203 http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0163014 doi:10.1371/journal.pone.0163014 doi:10.1371/journal.pone.0163014 |
| spellingShingle | Richard, Yolande Maan, Sushila Maan, Narender Singh Belaganahalli, Manjunatha N. Potgieter, Abraham C. Kumar, Vinay Batra, Kanisht Wright, Isabel M. Kirkland, Peter D. Mertens, Peter P.C. Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus |
| title | Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus |
| title_full | Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus |
| title_fullStr | Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus |
| title_full_unstemmed | Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus |
| title_short | Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus |
| title_sort | development and evaluation of real time rt-pcr assays for detection and typing of bluetongue virus |
| url | https://eprints.nottingham.ac.uk/38905/ https://eprints.nottingham.ac.uk/38905/ https://eprints.nottingham.ac.uk/38905/ |