An investigation into the wheat (Triticum aestivum L.) host response to Russian wheat aphid (Diuraphis noxia Kurd.) feeding

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dc.contributor.advisorBotha-Oberholster, Anna-Mariaen_ZA
dc.contributor.advisorVan der Vyver, Christellen_ZA
dc.contributor.authorFisher-Smith, Nadiaen_ZA
dc.contributor.otherStellenbosch University. Faculty of Agrisciences. Dept. of Genetics.en_ZA
dc.date.accessioned2023-09-22T11:15:54Z
dc.date.accessioned2024-01-08T13:09:33Z
dc.date.available2023-09-22T11:15:54Z
dc.date.available2024-01-08T13:09:33Z
dc.date.issued2023-12
dc.descriptionThesis (PhDAgric)--Stellenbosch University, 2023. en_ZA
dc.description.abstractENGLISH ABSTRACT: The Russian wheat aphid (RWA), Diuraphis noxia (Kurdjomov), is a major pest of wheat, causing damage and high yield losses worldwide. One of the undesirable effects of aphid feeding is leaf rolling, which serves as a shelter for the aphids protecting them from their natural predators and from insecticide spraying. Leaf rolling reduces the plant’s ability to photosynthesise and grow effectively, in addition, leaf rolling can also increase the aphid fitness, as it provides an ideal environment for growth. Therefore, identifying natural sources of resistance and introducing them into susceptible cultivars seems to be the most efficient strategy against RWA feeding. The use of genetic resistance is an efficient and environmentally safe method for controlling the RWA. This study aimed to ascertain if enhanced tolerance to biotic stress can be achieved by modifying plants either through genetic manipulation or chemical mutagenesis. Various studies have been done at a transcriptome level, allowing the identification of genes likely involved in RWA resistance. Utilising proteomics data in this study, allowed for the identification of differentially expressed peptides between resistant and susceptible wheat lines. Among the identified proteins were: glutathione-S-transferase (GST) and peroxidase. Literature suggests that GST forms part of the detoxification system in plants against biotic stress. This protein was uniquely expressed in the W1320-W1278 line that contains the Dn5 resistance gene. Peroxidase is associated with the oxidative burst, usually in response to stress, was identified in this study in the Gamtoos-S (Dn0) susceptible cultivar. Utilising genetic manipulation, a partial gene fragment of glutathione-S-transferase F6 (GSTF6b) was isolated from wheat and sequenced to confirm its identity. The gene fragment was cloned into a plant expression vector in the antisense orientation and bombarded into four- to six-day-old wheat immature embryos. Resulting in a putative transgenic plant, namely Gamtoos-S (Dn0)-pUBI-510:GSTF6b. Quantitative reverse- transcriptase-linked polymerase chain reaction (RT-qPCRs) were conducted to quantify the expression of the GSTF6b gene with/without RWA infestation. A reduction of nearly 50% was observed in GSTF6b expression in the respective transgenic plants when compared with the control. The T₁ was successfully hardened off, and allowed to seed and a T₂ generation was generated, which was functionally analysed through phenotypic screening, aphid fecundity, enzymatic responses and measuring oxidative burst. A decrease GST transcript level was observed post-infestation in the transgenic plants suggesting that plant susceptibility can probably be linked to a decrease in GST transcript promoting aphid growth and increasing the rate of reproduction. The last part of the study involved chemical mutagenesis, whereby drought-tolerant mutagenic M6 lines were screened for aphid resistance. A phenotyping assessment was performed on available mutant lines infested with South African (SA) biotype 1. A total of 33 mutant lines selected for drought tolerance, consisting of 21 ethyl methanesulfonate (EMS) and 12 Sodium azide (NaN₃) mutants, showed variation in aphid tolerance. Furthermore, drought-tolerant mutants were found to be more susceptible to aphid infestation, excluding the M12 (RYNOB8.012) line, shown to be intermediate to aphid feeding. The anti-oxidative enzyme GSTF6b expression was found to be significantly up-regulated in the mutagenic lines before infestation, therefore, contributing to the notion that GSTF6b is present at the basal level. A positive correlation was observed between GSTF6b gene expression and the intrinsic rate of increase (rm) in 25 mutagenic lines.en_ZA
dc.description.abstractAFRIKAANSE OPSOMMING: Die Russiese koring luis, Diuraphis noxia (Kurdjomov), is 'n groot plaag van koring, dit veroorsaak geweldige skade wêreldwyd met hoë verliese in opbrengs. Een van die newe effekte van plantluis voeding is blaarrol, wat dien as skuiling teen roofdiere en insekdoderbespuiting. Blaarrol verminder die plant se vermoë om te fotosinteer en om effektief te groei, asook dra dit by tot ’n toename in die plantluis se fiksheid, aangesien dit 'n ideale omgewing vir groei bied. Daarom is die identifisering van natuurlike weerstandsbronne en vrystelling van weerstandbiedende die mees effektiewe strategie om die plante tee luisskade te beskerm. Die gebruik van genetiese weerstand is 'n doeltreffende en omgewings-veilige metode om die koringluis te beheer. Die doel van die studie was om te toets of verhoogde weerstand teen biotiese stres bereik kan word deur plante te modifiseer, hetsy deur genetiese manipulasie of chemiese mutagenese. ‘n Verskeidenheid van studies was gedoen op transkripsievlak, wat die identifikasie van gene moontlik gemaak het, wat waarskynlik betrokke is by koringluis-weerstand. Die gebruik van proteomika-data in hierdie studie het dit moontlik gemaak om differensiëel uitgedrukte peptiede tussen weerstand biedende en vatbare koring lyne te identifiseer. Onder die geïdentifiseerde proteïene was: glutathioon-S-transferase (GST) en peroksidase. Literatuur dui daarop dat GST deel vorm van die ontgiftingssisteem in plante teen biotiese stres. Hierdie proteïen is uniek uitgedruk in die W1320-W1278-lyn wat die Dn5-weerstandsgeen bevat. Peroksidase word geassosieer met die oksidatiewe uitbarsting, gewoonlik in reaksie tot stres. In hierdie studie was peroksidase in die vatbare kultivar Gamtoos-S (Dn0) geïdentifiseer. Deur gebruik te maak van genetiese manipulasie, is ’n gedeelte van die geenfragment van glutathioon-S-transferase F6 (GSTF6b) uit koring geïsoleer en die identiteit daarvan is bevestig. Die geenfragment is in n plant- uitdrukkingsvektor in die teenrigting-oriëntasie gekloneer en gebombardeer in vier- tot ses dae oue koring-onvolwasse embrio's. Die transgeniese plant, Gamtoos-S (Dn0)-pUBI- 510:GSTF6b was geproduseer deur die proses. Kwantitatiewe omgekeerde transkriptase- gekoppelde polimerase ketting reaksie (RT-qPCR's) is uitgevoer om die uitdrukking van die GSTF6b geen met/sonder koringluis blootstelling te kwantifiseer. 'n Vermindering van byna 50% is waargeneem in GSTF6b-uitdrukking in die onderskeie transgeniese plante in vergelyking met hulle kontroles. Die T₁ generasie is suksesvol gegenereer, en toegelaat om te groei en 'n T₂-generasie te produseer, wat funksioneel ontleed is deur fenotipiese sifting, plantluis vrugbaarheid bepalings, ensiematiese reaksies en die meting van oksidatiewe uitbarsting. 'n Afname in GST-transkripsievlak was waargeneem na-infestasie in die transgeniese plante wat daarop dui dat plant vatbaarheid waarskynlik gekoppel kan word aan 'n afname in GST-transkripsie vlak. Laasgenomede bevorder plantluis groei en verhoog die voortplantingstempo. Die laaste deel van die studie het chemiese mutagenese behels, waardeur droogte-weerstand mutageniese M6-lyne vir plantluis-weerstand getoets was. ’n Fenotiperings bepaling is uitgevoer op beskikbare mutante lyne wat aan Suid-Afrikaanse (SA) biotipe 1 blootgestel was. 'n Totaal van 33 mutante lyne wat vir droogte toleransie geselekteer was, bestaande uit 21 etielmetaansulfonaat (EMS) en 12 Natriumasied (NaN₃) mutante, het ‘n variasie in plantluis-weerstand getoon. Verder is gevind dat droogte-weerstand mutante lyne meer vatbaar was vir plantluis blootstelling, uitgesluit die M12 (RYNOB8.012)-lyn, wat getoon is as intermediêr teen plantluis-voeding. Daar is gevind dat die anti-oksidatiewe ensiem GSTF6b-uitdrukking beduidend opgereguleer word in die mutageniese lyne voor blootstelling, wat dus bydra tot die idee dat GSTF6b teenwoordig is op die basale vlak. 'n Positiewe korrelasie is waargeneem tussen GSTF6b-geenuitdrukking en die intrinsieke tempo van toename (rm) in 25 mutageniese lyne.af_ZA
dc.description.versionDoctoralen_ZA
dc.format.extent141 pages : illustrations (some color)en_ZA
dc.identifier.urihttps://scholar.sun.ac.za/handle/10019.1/128844
dc.language.isoen_ZAen_ZA
dc.language.isoen_ZAen_ZA
dc.publisherStellenbosch : Stellenbosch Universityen_ZA
dc.rights.holderStellenbosch Universityen_ZA
dc.subject.lcshWheat (Triticum aestivum L.) -- Breedingen_ZA
dc.subject.lcshWheat -- Disease and pest resistance -- Genetic aspects -- Economic aspectsen_ZA
dc.subject.lcshRussian wheat aphid -- Host plantsen_ZA
dc.subject.lcshWheat -- Lossesen_ZA
dc.subject.lcshWheat -- Effect of stress on -- Genetic aspectsen_ZA
dc.subject.lcshWheat -- Environmental aspectsen_ZA
dc.subject.lcshChemical mutagenesisen_ZA
dc.subject.lcshWheat -- Growth -- Effect of chemicals onen_ZA
dc.subject.nameUCTDen_ZA
dc.titleAn investigation into the wheat (Triticum aestivum L.) host response to Russian wheat aphid (Diuraphis noxia Kurd.) feedingen_ZA
dc.typeThesisen_ZA
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