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The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer)
Thesis (MSc)--Stellenbosch University, 2021.
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| Format: | Thesis |
| Language: | en_ZA |
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Stellenbosch : Stellenbosch University
2020
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| _version_ | 1867614102776446976 |
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| access_status_str | Open Access |
| author | Smith, Katrin |
| author2 | Miller, Michele |
| author_browse | Miller, Michele Smith, Katrin |
| author_facet | Miller, Michele Smith, Katrin |
| author_sort | Smith, Katrin |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MSc)--Stellenbosch University, 2021. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/109399 |
| institution | Stellenbosch University (South Africa) |
| language | en_ZA |
| last_indexed | 2026-06-10T12:46:42.610Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2020 |
| publishDateRange | 2020 |
| publishDateSort | 2020 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/109399 The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) Smith, Katrin Miller, Michele Goosen, Wynand Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Mycobacterium bovis Tuberculosis Syncerus caffer Thesis (MSc)--Stellenbosch University, 2021. ENGLISH ABSTRACT: Mycobacterium bovis infection and the resulting bovine tuberculosis (bTB) disease affects a broad range of mammals including domestic animals, wildlife and humans. The presence of global wildlife maintenance hosts impedes efforts to control M. bovis transmission and bTB. African buffaloes (Syncerus caffer) are reservoir hosts of M. bovis in South Africa and thus pose a threat to multiple other wildlife species, adjacent livestock and their respective communities. Therefore, the efficient and early detection of M. bovis-infected buffaloes can help to minimize the spread of M. bovis and bTB disease. Considering that current standard bTB screening tools for buffaloes remain suboptimal, and in-field application presents several limitations, the validation of more optimal and approved diagnostic tools for M. bovis and bTB testing is required. This project investigated three such approaches to enhancing M. bovis detection in buffaloes. Firstly, the tuberculin skin test (TST), the primary ante mortem diagnostic tool for M. bovis detection in South Africa, was investigated for species-specific use. Buffalo specific cut-off values were calculated for the first time and TST test performance parameters evaluated, using a well-defined negative cohort and a gold standard positive cohort. The results present evidence for current South African guidelines for TST interpretation in buffaloes, in addition to providing alternative recommendations as required for different bTB testing scenarios. Moreover, the high specificity (Sp) yet suboptimal sensitivity (Se) of the TST was demonstrated, reiterating the necessity for additional tests to increase detection of infected animals. The most widely used ancillary test to the TST is the interferon-gamma (IFN-γ) release assay (IGRA). The second section of this project validated the new commercial Mabtech ELISAPRO bovine IFN-γ enzyme-linked immunosorbent assay (ELISA) that demonstrated promising test performance, with buffalo-specific cut-off values, in an IGRA using the QuantiFERON® TB-Gold Plus (QFT-Plus) stimulation platform and cohorts as described above. Furthermore, the QFT-Plus/Cattletype® IGRA, previously validated for buffalo use, was compared with the Mabtech IGRA after calculating species specific cut-off values to replace the manufacturer’s cattle threshold value. This served to significantly enhance Se. Finally, as a novel approach to in vitro measurement of cell-mediated immune responses to M. bovis, a MILLIPLEX® bovine cytokine/chemokine multiplex assay was investigated for use in buffaloes. The results demonstrated the detection of all fifteen target biomarkers with significant differences observed between mitogen-stimulated and/or antigen-stimulated, and unstimulated plasma samples in M. bovis-infected buffaloes. Overall, the results from this project reveal the importance of species-specific validation of diagnostic tests intended for field use. For wildlife species this is particularly challenging, and relevant positive and negative reference samples are not always readily available. Furthermore, the optimisation of current (and candidate) tests, as demonstrated in this project for the TST and IGRAs, should be performed in line with official validation standards (when possible). This will better enable the recognition and in-field application of enhanced and promising diagnostic tools for improved detection of M. bovis infection in African buffaloes. AFRIKAANS OPSOMMING: Mycobacterium bovis infeksie is verantwoordelik vir die siekte bekend as bees tuberkulose (bTB) in verskeie soogdiere soos plaasdiere- en wilde diere, asook mense. Die teenwoordigheid van ‘n instandhoudingsgasheer soos die Kaapse buffel (Syncerus caffer) in ‘n wildsisteem, ondermyn alle pogings om sulke infeksies in verskeie omgewings te beheer. Om verdere verspreiding van M. bovis onder soogdiere te voorkom is dit dus belangrik om M. bovis infeksie in ‘n dier so vroeg as moontlik te identifiseer deur gebruik te maak van die mees optimale toetse. Daar word egter steeds geglo dat die diagnostiese toetse wat gebruik word gedurende bTB-beheerprogramme, soos die enkele intradermale vergelykende tuberkulien-toets (“single intradermal comparative tuberculin test”; SICTT) en kommersieel beskikbare interferon-gamma (IFN-γ)-vrystellingstoetse (“IFN-γ release assays”; IGRAs), suboptimaal is vir die diagnose van bTB in beesverwante hoefdiere. Die doelwit van hierdie studie was om die beskikbare toetse, soos vroeër beskryf, vir die eerste keer te optimiseer vir gebruik in buffels. Buffel spesifieke afsnypunte vir die veltoets (SICTT) was bepaal deur gebruik te maak van historiese negatiewe buffel populasies asook wel bekende positiewe populasies. Resultate verkry gedurende hierdie studie ondersteun huidige Suid-Afrikaanse veltoets riglyne vir die interpretasie van veltoets reaksies in buffels en voorsien ook addisionele gebruike en riglyne vir ander tipe buffel populasies. Die volgende komponent van hierdie studie was om ‘n nuwe IFN-γ-vrystellingstoets, die ‘Mabtech ELISAPRO bovine’ IFN-γ ensiem-gekoppelde immunosorbente toets (ELISA) te evalueer in bogenoemde negatiewe en postiewe buffel populasies en die diagnostiese potensiaal van hierdie toets te vergelyk met die meer bekende ‘Cattletype’ ELISA. Vir hierdie toetse is die plasma van die diere versamel nadat bloed in die QuantiFERON® TB-Gold Plus (QFT-Plus) sisteem gestimuleer was. Na buffel-spesifieke afsnypunte bepaal is vir albei ELISAs, was dit duidelik dat albei toetse se diagnostiese sensitiwiteit eenders was. Laastens, om verdere moontlike immunologiese kandidaat merkers van M. bovis infeksie te identifiseer vir moontlike diagnostiese gebruik in buffels, is die ‘MILLIPLEX® bovine cytokine/chemokine multiplex’ toets vir die eerste keer in buffels getoets. Vyftien moontlike merkers met diagnostiese potensiaal was geïdentifiseer, omdat hulle suksesvol tussen mitogeen gestimuleerde plasma, M. bovis antigeen-gestimuleerde plasma en ongestimuleerde plasma van M. bovis-geïnfekteerde buffels kon onderskei. Ten slotte beklemtoon, hierdie studie die belangrikheid van spesie-spesifieke optimisering en validasie van alle diagnostiese toetse wat in die veld op diere gebruik mag word. In wilde diere is hierdie tipe optimiserings en validasies aansienlik moeiliker in vergelyking met ander diere soos beeste. Verder is dit baie moeilik om bevestigde M. bovis negatiewe buffels te bekom. As gevolg van bogenoemde redes is hierdie studie se bevindinge waardevol vir siekte navorsing in wilde diere. Wanneer toetse gevalideer word soos in hierdie studie, moet die amptelike validasie riglyne van die land gevolg word (waar moontlik) om verbeterde diagnostiese toetse vir M. bovis infeksies in die Kaapse buffels te ontwikkel. Sodoende, sal toetse vinniger deur die staat aanvaar word en die gebruik daarvan in die veld grootliks ondersteun word. Masters 2020-09-07T06:27:54Z 2021-02-01T07:53:46Z 2020-09-07T06:27:54Z 2021-02-01T07:53:46Z 2020-12 Thesis http://hdl.handle.net/10019.1/109399 en_ZA Stellenbosch University application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Mycobacterium bovis Tuberculosis Syncerus caffer Smith, Katrin The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) |
| title | The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) |
| title_full | The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) |
| title_fullStr | The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) |
| title_full_unstemmed | The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) |
| title_short | The evaluation and validation of cell-mediated immunological responses for the improved detection of Mycobacterium bovis infection in African buffaloes (Syncerus caffer) |
| title_sort | evaluation and validation of cell mediated immunological responses for the improved detection of mycobacterium bovis infection in african buffaloes syncerus caffer |
| topic | Mycobacterium bovis Tuberculosis Syncerus caffer |
| url | http://hdl.handle.net/10019.1/109399 |
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