Characterization of citrus tristeza virus variants and their influence on symptom expression in grapefruit

Cook, Glynnis (2019-04)

Thesis (PhDAgric)--Stellenbosch University, 2019.

Thesis

ENGLISH ABSTRACT: Citrus tristeza virus (CTV), a member of the family Closteroviridae, was responsible for large scale destruction of citrus, especially in the Americas, due to tristeza disease and necessitated a production switch to less sensitive rootstocks. CTV however continues to affect citrus through the stem-pitting disease phenotype which is especially problematic in sweet orange, grapefruit and lime cultivars. In South Africa, the productive lifespan of grapefruit trees was severely affected by stem-pitting, requiring early tree replacement with an associated lag in production. This affect was later mitigated by applying cross-protection, a management strategy using non-stem-pitting sources of CTV, but without prior knowledge of which CTV strains were responsible for stempitting or which strains were present in the cross-protection sources. To understand the disease and unravel mechanisms underlying cross-protection, it is necessary to characterise CTV strains to investigate both virus-host- and strain-interactions. The aim of this study was firstly to identify single-strain isolates belonging to different strains, to characterise them biologically and to determine full-genome sequences. These characterised CTV isolates were further used in a complementation study to investigate possible synergistic interactions affecting stem-pitting. Complete viral genomes of eight single-strain isolates were determined during the study. Two commercial grapefruit cultivars, ‘Star Ruby’ and ‘Marsh’, were used in a glasshouse trial to evaluate the ability of specific strains to induce stem-pitting in single or mixed infections. Evaluation over four years showed that symptom expression of mild strains did not result in altered symptom expression when in combination with each other. Importantly demonstrating that there was no additive effect on stem-pitting expression with multiple isolates. Relative quantitation of the strains in ‘Marsh’ and ‘Star Ruby’ plants indicated that the individual strain concentrations were not significantly altered when in combination with the other strains. A valuable discovery made within this project was the characterisation of two variants of the T68 strain, derived from the same GFMS12 source, but displaying differences in stem-pitting severity in grapefruit. This finding demonstrates the co-existence of severe and mild variants of the same strain in one source and provides an explanation for the presumed strain segregation event observed for the GFMS12 cross-protection source that resulted in the discontinuation of the source for use in cross-protection of grapefruit. The characterisation of these variants will further assist in the identification of the sequence determinants for stem-pitting in grapefruit.

AFRIKAANSE OPSOMMING: Sitrus tristeza virus (CTV), 'n lid van die familie Closteroviridae, was verantwoordelik vir grootskaalse vernietiging van sitrus, weens tristeza siekte en het 'n produksie veradering na minder sensitiewe onderstamme genoodsaak. CTV het egter steeds 'n nadelige invloed op sitrus weens stamgleuf wat veral problematies is in soetlemoen, pomelos en lemmetjie kultivars. In Suid-Afrika is die produktiewe lewensduur van pomelo-bome verlaag weens stamgleuf, wat vroeë boomvervanging met 'n gepaardgaande produksieverlaging vereis het. Hierdie invloed is later verminder deur die toepassing van kruisbeskerming, 'n bestuurstrategie waar ligte bronne van CTV toegepas word, maar sonder voorafgaande kennis van watter CTV-rasse verantwoordelik is vir stamgleuf of watter rasse teenwoordig is in die bronne. Om die siekte te verstaan en meganismes onderliggend aan kruisbeskerming te ontleed, is dit nodig om CTV-rasse te karakteriseer om beide virus-gasheer- en rasinteraksies te ondersoek. Die doel van hierdie studie was om eerstens enkelras isolate wat aan verskillende rasse behoort, te identifiseer, om hulle biologies te karakteriseer en om volledige-genoom-nukleotiedvolgordes te bepaal. Hierdie gekarakteriseerde CTV-isolate is verder gebruik in 'n komplementeringsstudie om moontlike sinergistiese interaksies te ondersoek wat stamgleuf beïnvloed. Vollengte virale genome van agt enkelras-isolate is tydens die studie bepaal. Twee kommersiële pomelo kultivars, 'Star Ruby' en 'Marsh', is in 'n glashuisproef gebruik om die vermoë van spesifieke rasse te evalueer om stamgleuf in enkel- of gemengde infeksies te veroorsaak. Evaluering oor vier jaar het getoon dat simptoom uitdrukking van ligte rasse nie gelei het tot 'n veranderde simptoom uitdrukking wanneer hul in kombinasie met mekaar voorgekom het nie. Dit is belangrik om aan te toon dat daar geen toevoegende effek op stamgleuf uitdrukking met veelvoudige isolate was nie. Relatiewe hoeveelheids bepaling van die rasse in 'Marsh' en 'Star Ruby' plante het aangedui dat die individuele raskonsentrasies nie beduidend verander in kombinasie met die ander rasse nie. 'n Waardevolle ontdekking wat in hierdie projek gemaak is, was die karakterisering van twee variante van die T68-ras, afkomstig van dieselfde GFMS12-bron, maar met verskille in stamgleuf uitdrukking in pomelo's. Hierdie bevinding demonstreer die gelyktydige bestaan van strawwe en ligte variante van dieselfde ras in een bron en verskaf 'n verduideliking vir die vermoedelike ras segregasie gebeurtenis waargeneem in die GFMS12 kruisbeskerming bron, wat gelei het tot die staking van die bron vir gebruik as kruisbeskerming van pomelo's. Die karakterisering van hierdie variante sal verder help met die identifisering van die volgorde-bepaling van stamgleuf in pomelo's.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/105724
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