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Characterisation of citrus tristeza virus-induced stem pitting in citrus
Thesis (PhDAgric)--Stellenbosch University, 2023.
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| Format: | Thesis |
| Language: | en_ZA en_ZA |
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Stellenbosch : Stellenbosch University
2023
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| _version_ | 1867613949748314112 |
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| access_status_str | Open Access |
| author | Aldrich, Dirk Jacobus |
| author2 | Maree, Hans Jacob |
| author_browse | Aldrich, Dirk Jacobus Maree, Hans Jacob |
| author_facet | Maree, Hans Jacob Aldrich, Dirk Jacobus |
| author_sort | Aldrich, Dirk Jacobus |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (PhDAgric)--Stellenbosch University, 2023. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/128835 |
| institution | Stellenbosch University (South Africa) |
| language | en_ZA en_ZA |
| last_indexed | 2026-06-10T12:44:16.501Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2023 |
| publishDateRange | 2023 |
| publishDateSort | 2023 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/128835 Characterisation of citrus tristeza virus-induced stem pitting in citrus Aldrich, Dirk Jacobus Maree, Hans Jacob Bester, Rachelle Burger, Johan Theodorus Stellenbosch University. Faculty of Agrisciences. Dept. of Genetics. Citrus tristeza disease Citrus tristeza virus -- Genetic engineering Reverse genetics Stem pitting -- Pathogenesis Stems (Botany) -- Diseases and pests Plant biotechnology Citrus -- Diseases and pests -- Economic aspects UCTD Thesis (PhDAgric)--Stellenbosch University, 2023. ENGLISH ABSTRACT: Citrus tristeza virus (CTV) is the most important viral pathogen of citrus and causes several disease syndromes in different citrus hosts. CTV-induced stem pitting leads to substantial economic losses in sensitive citrus varieties, including grapefruit. The exact mechanisms of stem pitting development in CTV-infected citrus remain unclear. This study aimed to utilise CTV infectious clone mutants in a reverse genetics approach to study stem pitting induction. A panel of recombinant CTV clones was generated focussing on the open reading frames (ORFs) implicated in stem pitting induction and pathogenicity, namely p33, p18, p13 and p23. ORF replacements from severe- and mild-pitting South African CTV isolates were introduced into the mild-pitting infectious clone (CTV-fl6 - genotype T36) to determine if severe stem pitting could be induced. Stem pitting assessments were complemented with the determination of virus concentration ratios in ‘Mexican’ lime and ‘Duncan’ grapefruit to relate stem pitting outcomes to virus concentration. The various infectious clone and wild type CTV infections yielded a broad range of stem pitting outcomes and specific ORF replacements associated with enhanced stem pitting were identified. Plant responses to different stem pitting pressures were further assessed by untargeted metabolite profiling and the quantitation of the stress-responsive phytohormones, abscisic acid, jasmonic acid and salicylic acid. In both citrus hosts, the metabolite profiling yielded fourteen statistically significant compounds that differed between stem pitting groups. These compounds were mainly phenolic acids and phenolic glycosides and are known to function as antioxidant and stress signalling molecules. Significant differences in phytohormone content were also found between test groups, particularly in plants that were severely stunted. Stem pits were also characterised at the molecular anatomical level using a combination of known and novel techniques to better understand the nature of the xylem and phloem tissues impacted by severe pitting. Established methods such as biological staining and fluorescence microscopy were used to visualise CTV-induced stem pitting and virus localisation. The utility of two novel technologies that have not previously been used to study CTV-induced stem pitting, namely high-resolution CT scanning and serial block face scanning electron microscopy was also evaluated. This study contributed to our understanding of CTV-induced stem pitting in citrus. Importantly, the use of CTV infectious clones for South African CTV research was established, which can allow for targeted follow-up experiments to further characterise the local citrus-CTV pathosystem. This can also provide novel opportunities for potentially employing CTV infectious clones as a crop protection tool against other important citrus diseases on the African continent. AFRIKAANSE OPSOMMING: Sitrus tristeza-virus (CTV) is die belangrikste virale patogeen van sitrus en veroorsaak verskeie siektesindrome in verskillende sitrusgashere. CTV-geïnduseerde stampitting lei tot aansienlike ekonomiese verliese in sensitiewe sitrusvariëteite, insluitend pomelo. Die presiese meganismes van stampit-ontwikkeling in CTV-geïnfekteerde sitrus bly onduidelik. Hierdie studie het ten doel gehad om CTV-aansteeklike kloonmutante te gebruik in 'n omgekeerde genetika-benadering om stampit-induksie te bestudeer. 'n Paneel van rekombinante CTV-klone is gegenereer wat fokus op die oop leesrame (ORF's) wat by stampit-induksie en patogenisiteit geïmpliseer is, naamlik p33, p18, p13 en p23. ORF-vervangings van Suid-Afrikaanse CTV-isolate wat ernstige asook ligte stampitte veroorsaak is in die ligte-pittende-aansteeklike kloon (CTV-fl6 - genotipe T36) ingebring om te bepaal of ernstige stampitting veroorsaak kan word. Stampit-evaluerings is aangevul met die bepaling van viruskonsentrasies in 'Mexikaanse' lemmetjie en 'Duncan' pomelo plante om stampituitkomste met viruskonsentrasie in verband te bring. Die verskillende aansteeklike kloon- en wilde tipe CTV-infeksies het 'n wye reeks stampituitkomste gelewer en spesifieke ORF-vervangings geassosieer met verhoogde stampitting is geïdentifiseer. Plantreaksies op verskillende stampitdrukke is verder geassesseer deur ongeteikende metabolietprofilering en die kwantifisering van die stres-responsiewe fitohormone, absisiensuur, jasmoniese suur en salisielsuur. In beide sitrusgashere het die metaboliet-profilering veertien statisties beduidende metaboliete geïdentifiseer tussen stampitgroepe. Hierdie metaboliete was hoofsaaklik fenoliese sure en fenoliese glikosiede en is bekend daarvoor dat dit funksioneer as antioksidant- en stresseinmolekules. Beduidende verskille in fitohormooninhoud is ook tussen toetsgroepe gevind, veral vir die plante wat belemmerde groei gehad het. Stampitte is ook op die molekulêre anatomiese vlak gekarakteriseer deur 'n kombinasie van bekende en nuwe tegnieke te gebruik om die aard van die xileem- en floëemweefsels wat deur ernstige pitting geaffekteer word, beter te verstaan. Gevestigde metodes soos biologiese kleuring en fluoressensiemikroskopie is gebruik om CTV- geïnduseerde stampitting en viruslokalisering te visualiseer. Die nuttigheid van twee nuwe tegnologieë wat nog nie voorheen gebruik is om CTV-geïnduseerde stampitting te bestudeer nie, naamlik hoë-resolusie CT-skandering en seriële blokgesigskandering-elektronmikroskopie is ook geëvalueer. Hierdie studie het bygedra tot ons begrip van CTV-geïnduseerde stampitting in sitrus. Die gebruik van CTV-aansteeklike klone is ook gevestig vir Suid-Afrikaanse CTV- navorsing, wat gebruik kan word vir geteikende opvolgeksperimente om die plaaslike sitrus- CTV patosisteem verder te karakteriseer. Dit kan ook nuwe geleenthede bied vir die moontlike gebruik van CTV-aansteeklike klone as 'n gewasbeskermingsinstrument teen ander belangrike sitrussiektes op die Afrika-kontinent. Doctoral 2023-10-20T06:42:10Z 2024-01-08T12:49:53Z 2023-10-20T06:42:10Z 2024-01-08T12:49:53Z 2023-12 Thesis https://scholar.sun.ac.za/handle/10019.1/128835 en_ZA en_ZA Stellenbosch University xvii, 187 pages : illustrations (some color) application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Citrus tristeza disease Citrus tristeza virus -- Genetic engineering Reverse genetics Stem pitting -- Pathogenesis Stems (Botany) -- Diseases and pests Plant biotechnology Citrus -- Diseases and pests -- Economic aspects UCTD Aldrich, Dirk Jacobus Characterisation of citrus tristeza virus-induced stem pitting in citrus |
| title | Characterisation of citrus tristeza virus-induced stem pitting in citrus |
| title_full | Characterisation of citrus tristeza virus-induced stem pitting in citrus |
| title_fullStr | Characterisation of citrus tristeza virus-induced stem pitting in citrus |
| title_full_unstemmed | Characterisation of citrus tristeza virus-induced stem pitting in citrus |
| title_short | Characterisation of citrus tristeza virus-induced stem pitting in citrus |
| title_sort | characterisation of citrus tristeza virus induced stem pitting in citrus |
| topic | Citrus tristeza disease Citrus tristeza virus -- Genetic engineering Reverse genetics Stem pitting -- Pathogenesis Stems (Botany) -- Diseases and pests Plant biotechnology Citrus -- Diseases and pests -- Economic aspects UCTD |
| url | https://scholar.sun.ac.za/handle/10019.1/128835 |
| work_keys_str_mv | AT aldrichdirkjacobus characterisationofcitrustristezavirusinducedstempittingincitrus |