Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Grapevine leafroll-associated virus 3 (GLRaV-3) is the type member of the genus Ampelovirus in the family Closteroviridae and is considered to be the main contributing agent of grapevine leafroll disease (GLD) worldwide. A metagenomic sequencing study of a grapevine leafroll-diseased vineyard led to the discovery of a new variant of GLRaV-3 in South Africa. This new variant was most related to a New Zealand isolate, NZ-1. In this study, we sequenced two isolates, GH11 and GH30, of the new variant group of GLRaV-3. These isolates have less than 70% nucleotide (nt) identity to other known GLRaV-3 variants, indicating that they should be considered variants of a different strain of GLRaV-3. We propose that the GLRaV-3-like virus identified in this study be grouped together with NZ-1 and some Napa Valley isolates as Group VI of GLRaV-3. This study also provided further evidence that next-generation sequencing is an invaluable approach to identify novel viruses and variants, in that the draft sequence generated with bioinformatic tools in this study was 98% identical to the GH11 sequence generated using Sanger sequencing. The study further confirmed that the industry standard ELISA is still an effective GLRaV-3 diagnostic method and that it is able to detect all known variant groups of GLRaV-3. However, this assay is not able to differentiate between GLRaV-3 variant groups. In the current study therefore, a real-time RT-PCR was designed that is able to detect GLRaV-3 variant groups I, II, III and VI, using a single primer pair targeting the Hsp70h gene of GLRaV-3. If high-resolution melting (HRM) curve analysis is added to the real-time RT-PCR, it is possible to differentiate between variant groups based on three melting point intervals. The RT-PCR HRM assay provides a more sensitive and rapid tool to detect and differentiate between different GLRaV-3 variant groups. Finally, a multiplex RT-PCR was designed to differentiate between the variant groups present in South Africa. This multiplex RT-PCR offers a validation method for the RT-PCR HRM and provides an end-point PCR alternative for variant identification. In order to investigate the spread and impact of different GLRaV-3 variants in vineyards, sensitive diagnostic techniques are a necessity. The abovementioned tools will contribute to the understanding of the pathogenesis of GLD and aid epidemiological studies to investigate how these different GLRaV-3 variant groups are spreading, the association of specific GLRaV-3 variants to disease symptoms and the mealybug vector transmission efficiency for each GLRaV-3 variant. / AFRIKAANSE OPSOMMING: Grapevine leafroll-associated virus 3 (GLRaV-3) is ’n lid van die genus Ampelovirus in die familie Closteroviridae en word beskou as die hoof bydraende faktor van wingerd-rolbladsiekte wêreldwyd. ’n Metagenomiese studie het bewys dat daar ’n nuwe variant van GLRaV-3 bestaan wat nog nie voorheen in Suid Afrika opgespoor kon word met die huidige opsporingsmetodes nie. Hierdie nuwe variant was naaste verwant aan ’n Nieu-Seelandse isolaat, NZ-1. In hierdie studie is die genoomvolgorde van twee isolate, GH11 en GH30, van hierdie nuwe GLRaV-3 variant groep bepaal. Hierdie twee isolate was minder as 70% identies aan ander GLRaV-3 variante, wat daarop dui dat hulle as variante van ’n nuwe virus-ras beskou behoort te word. Ons beveel aan dat hierdie GLRaV-3-verwante virus geklassifiseer word saam met die NZ-1 isolaat en ander isolate uit Kalifornië, as groep VI van GLRaV-3. Hierdie studie het ook verdere bewyse verskaf dat volgende-generasie volgordebepalingstegnologie ’n waardevolle benadering is om nuwe virusse en variante te identifiseer, deurdat die huidige studie gewys het dat die voorlopige volgorde, wat gegenereer is deur bioinformatika-instrumente, 98% identies was aan die GH11 volgorde wat met Sanger volgordebepaling verkry was. Hierdie studie het ook gevind dat die industrie-standaard ELISA, nog steeds ’n effektiewe GLRaV-3 diagnostiese metode is en wel infeksies, veroorsaak deur al die variant-groepe, sal kan identifiseer. Die ELISA toets is egter nie in staat om te onderskei tussen GLRaV-3 variant-groepe nie. In hierdie studie is ’n variant-identifiseerbare in-tyd tru-transkripsie polimerase ketting reaksie (PKR) ontwerp wat GLRaV-3 variant-groepe I, II, III en VI kan identifiseer deur middel van ’n enkele inleier-stel wat die GLRaV-3 Hsp70h-geen teiken. As hoë-resolusie smeltingskurwe-analise bygevoeg word by die in-tyd tru-transkripsie PKR, is dit moontlik om te onderskei tussen variant-groepe op grond van drie smeltingspunt intervalle. Die tru-transkripsie hoë-resolusie smeltingskurwe-toets verskaf meer sensitiewe en geoutomatiseerde metodes om GLRaV-3 variant-groepe te identifiseer en te onderskei. ’n Veelvuldige tru-transkripsie PKR is ook ontwerp om tussen variante wat tans in Suid-Afrika aangetref word, te onderskei en te dien as ’n valideringsmetode vir die in-tyd tru-transkripsie hoë-resolusie smeltingskurwe-toets. Sensitiewe en akkurate toetse, soos bogenoemde, is noodsaaklik vir die bestudering van die verspreiding en impak van die verskillende GLRaV-3 variante in wingerd. Hierdie metodes kan gebruik word om kennis ten opsigte van rolblad patogenese te verbreed en om by te dra tot epidemiologiese studies wat ondersoek hoe hierdie variant-groepe versprei, of daar ’n assosiasie bestaan tussen ’n spesifieke variant en siekte-simptome en of daar ’n verskil is in die witluisvektor oordragseffektiwitiet vir elke GLRaV-3 variant.

Thesis (MSc)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Metagenomic studies that make use of next-generation sequencing (NGS) generate large amounts of sequence data, representing the genomes of multiple organisms of which no prior knowledge is necessarily available. In this study, a metagenomic NGS approach was used to detect multiple novel mycoviral sequences in grapevine phloem tissue. Individual sequencing libraries of doublestranded RNA (dsRNA) from two grapevine leafroll diseased (GLD) and three shiraz diseased (SD) vines were sequenced using an Illumina HiScanSQ instrument. Over 3.2 million reads were generated from each of the samples and these reads were trimmed and filtered for quality before being de novo assembled into longer contigs. The assembled contigs were subjected to BLAST (Basic Local Alignment Search Tool) analyses against the NCBI (National Centre for Biotechnology Information) database and classified according to database sequences with which they had the highest identity. Twenty-six putative mycovirus species were identified, belonging to the families Chrysoviridae, Endornaviridae, Narnaviridae, Partitiviridae and Totiviridae. Two of the identified mycoviruses, namely grapevine-associated chrysovirus (GaCV) and grapevine-associated mycovirus 1 (GaMV-1) have previously been identified in grapevine while the rest appeared to be novel mycoviruses not present in the NCBI database. Primers were designed from the de novo assembled mycoviral sequences and used to screen the grapevine dsRNA used for sequencing as well as endophytic fungi isolated from the five sample vines. Only two mycoviruses, related to sclerotinia sclerotiorum partitivirus S and chalara elegans endornavirus 1 (CeEV-1), could be detected in grapevine dsRNA and in fungus isolates. In order to validate the presence of mycoviruses in grapevine phloem tissue, two additional sequencing runs, using an Illumina HiScanSQ and an Applied Biosystems (ABI) SOLiD 5500xl instrument respectively, were performed. These runs generated more and higher quality sequence data than the first sequencing run. Twenty-two of the putative mycoviral sequences initially detected were detected in the subsequent sequence datasets, as well as an additional 29 species not identified in the first HiScanSQ sequence datasets. The samples harboured diverse mycovirus populations, with as many as 19 putative species identified in a single vine. This indicates that the complete virome of diseased grapevines will include a high number of mycoviruses. Additionally, the complete genome of a novel endornavirus, for which we propose the name grapevine endophyte endornavirus (GEEV), was assembled from one of the second HiScanSQ sequence datasets. This is the first complete genome of a mycovirus detected in grapevine. Grapevine endophyte endornavirus has the highest sequence similarity to CeEV-1 and is the same virus that was previously detected in fungus isolates using the mycovirus primers. The virus was detected in two fungus isolates, namely Stemphylium sp. and Aureobasidium pullulans, which is of interest since mycoviruses are not known to be naturally associated with two distinctly different fungus genera. Mycoviral sequence data generated in this study can be used to further investigate the diversity and the effect of mycoviruses in grapevine. / AFRIKAANSE OPSOMMING: Metagenomiese studies, wat gebruik maak van volgende-generasie volgordebepalingstegnologie, het die vermoë om die genetiese samestelling van veelvoudige onbekende organismes te bepaal deurdat dit groot hoeveelhede data genereer. Die bogenoemde tegniek was in hierdie studie aangewend om aantal nuwe mikovirusse in die floëem weefsel van wingerd te identifiseer. Dubbelstring-RNS was gesuiwer vanuit twee druiwestokke met rolbladsiekte en drie met shirazsiekte en Illumina HiScanSQ instrument is gebruik om meer as 3.2 miljoen volgorde fragmente te genereer van elk van die monsters. Lae-kwaliteit volgordes was verwyder en die oorblywende kort volgorde fragmente was saamgestel om langer konstrukte te vorm wat met behulp van BLAST soektogte teen die NCBI databasis geïdentifiseer kon word. Ses-en-twintig mikovirus spesies, wat aan die families Chrysoviridae, Endornaviridae, Narnaviridae, Partitiviridae en Totiviridae behoort, was geïdentifiseer. Twee van die geïdentifiseerde mikovirusse, naamlik grapevine-associated chrysovirus (GaCV) en grapevine-associated mycovirus 1 (GaMV-1), was voorheen al in wingerd gekry terwyl die res nuwe mikovirusse is wat tans nie in die NCBI databasis voorkom nie. Inleiers was ontwerp vanaf die saamgestelde mikovirus basisvolgordes en gebruik om wingerd dubbelstring-RNS sowel as swamme wat vanuit die wingerd geïsoleer is te toets vir die teenwoordigheid van hierdie mikovirusse. Slegs twee mikovirusse, wat onderskeidelik verwant is aan sclerotinia sclerotiorum partitivirus S en chalara elegans endornavirus 1 (CeEV-1), kon deur middel van die inleiers in wingerd en swam isolate geïdentifiseer word. Twee addisionele volgordebepalingsreaksies, wat gebruik gemaak het van die Illumina HiScanSQ en ABI SOLiD 5500xl volgordebepalingsplatforms, was gebruik om die teenwoordigheid van mikovirusse in wingerd te bevestig. Groter hoeveelheid volgorde fragmente was geprodusser wat ook van hoër gehalte was as dié van die eerste volgordebepalingsreaksie. Twee-en-twintig mikovirus spesies kon weer geïdentifiseer word, sowel as 29 spesies wat nie in die eerste HiScanSQ basisvolgorde datastelle gevind was nie. Die wingerdstokke wat in hierdie studie ondersoek was, het hoë diversiteit van mikovirusse bevat aangesien daar tot 19 mikovirus spesies in enkele wingerdstok geïdentifiseer was. Dit is aanduiding dat volledige virus profiele van siek wingerdstokke aantal mikovirusse sal insluit. Die vollengte genoomvolgorde van voorheen onbekende endornavirus was saamgestel vanuit een van die tweede HiScanSQ volgorde datastelle. Dit is die eerste mikovirus wat in wingerd gevind word waarvan die volledige genoomvolgorde bepaal is en ons stel die naam grapevine endophyte endornavirus (GEEV) voor vir hierdie virus. Grapevine endophyte endornavirus is die naaste verwant aan CeEV-1 en is dieselfde virus wat voorheen in wingerd dubbelstring-RNS en swam isolate gevind was deur middel van die mikovirus inleiers. Swam isolate waarin GEEV gevind is, was geïdentifiseer as Stemphylium sp. en Aureobasidium pullulans. Dit is van belang dat GEEV in twee swam isolate gevind is wat aan verskillende genusse behoort aangesien hierdie verskynsel nog nie voorheen in die natuur gevind is nie. Mikovirus nukleiensuurvolgordes wat in hierdie studie bepaal was kan gebruik word in toekomstige studies om die verskeidenheid en impak van mikovirusse in wingerd verder te ondersoek. / National Research Foundation (NRF) / Stellenbosch University

Thesis (MSc)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: South Africa is ranked eighth in the world as far as international wine production is concerned and in terms of area under bearing vines South Africa is ranked 12th. In 2011 the wine industry contributed R4 204.4 million to the South African economy in state revenue from wine products. The importance of viticulture to the economy of South Africa forces the industry to limit the effect of all disease causing pathogens in order to keep their competitive edge. Aster yellows (AY) phytoplasma 16SrI-B subgroup was reported for the first time in grapevine (Vitis vinifera L. (Vitaceae)) in South Africa in 2006. Worldwide phytoplasma diseases of grapevine cause serious damage ranging from lower yields to the death of vines. The lack of knowledge about the epidemiology of AY disease makes it difficult to determine the impact of the disease on the South African wine industry. The aim of this study was to conduct surveys in disease-affected vineyards in the Vredendal region to determine the incidence and spatial distribution of the disease in a variety of cultivars. The field surveys based on visual symptoms of AY disease were confirmed by polymerase chain reaction (PCR). A survey was also conducted in and around AY-infected vineyards in search of possible alternative host plants of the phytoplasma. Spatial distribution of AY-affected vines were analysed using the PATCHY spatial analysis package. A rapid decline of AY-affected Chardonnay eventually leading to the death of vines was observed, confirming the sensitivity of Chardonnay towards grapevine yellows infections. Symptomless AY infections occurred and AY could not be detected in all symptomatic vines, which indicate uneven distribution of AY in individual vines. Molecular analyses using PCR-RFLP showed that all vines sampled in the Vredendal vicinity contained AY phytoplasma only. No phytoplasmas were present in any weeds or other possible host plants tested. Although the mean yearly disease incidences of Chardonnay (29.95%) and Chenin blanc (16.64%) were higher than Pinotage (5.80%) over the four-year survey period, there was no statistically significant difference between the disease incidences of these three cultivars. The mean yearly disease incidence showed a trend over time and the disease incidence of the first year was significantly lower than that of the other years. Chardonnay showed a cumulative disease incidence of 37.77% at the end of the 4-year study which means that Chardonnay vineyards can be 100% AY infected in ten years’ time. Spatial distribution patterns of AY-infected vines were mostly non-random with clustering of disease affected vines along and across vine rows. With the exception of one vineyard, aggregation of AY-affected vines mostly occurred on the edge of vineyards adjacent to infected vineyards. This epidemiological study gives an indication of the sensitivity of the different cultivars towards AY, the tempo of spreading and the future impact of the disease on the South African wine industry. It also contributes valuable information towards the development of a management strategy for grapevine yellows disease in South African vineyards. / AFRIKAANSE OPSOMMING: Suid- Afrika is op agtste op die wêreld ranglys wat internasionale produksie van wyn aan betref, en in terme van oppervlakte onder wingerd, is Suid-Afrika 12de. In 2011 het die wynbedryf R4 204.4 miljoen tot die Suid-Afrikaanse ekonomie bygedra in staats inkomste uit wyn produkte. Die belangrikheid van wingerd tot die ekonomie van Suid-Afrika dwing die bedryf om die effek van alle siekteveroorsakende patogene te beperk, om sodoende hul kompeterende voordeel te behou. Aster vergeling (AY) fitoplasma 16SrI-B subgroep is vir die eerste keer in 2006 in wingerd (Vitis vinifera L. (Vitaceae)) in Suid-Afrika waargeneem. Fitoplasma siektes van wingerd veroorsaak wêreldwyd ernstige skade wat wissel van laer opbrengste tot die afsterf van wingerdstokke. Die gebrek aan kennis oor die epidemiologie van astervergeling siekte maak dit moeilik om die impak van die siekte op die Suid-Afrikaanse wynbedryf te bepaal. Die doel van hierdie studie was om ‘n opname te maak in siekte geaffekteerde wingerde in die Vredendal omgewing om sodoende siekte voorkoms en verspreidingspatrone van die siekte in 'n verskeidenheid van kultivars te bepaal. Die veld opnames, gebaseer op visuele simptome van aster vergeling siekte, was bevestig deur polimerase kettingreaksie (PKR). ‘n Opname is ook in en om aster vergeling geaffekteerde wingerde uitgevoer, op soek na moontlike alternatiewe gasheer plante van die fitoplasma. Verspreidingspatrone van astervergeling geaffekteerde wingerde is ontleed met behulp van die PATCHY ruimtelike analise pakket. 'n Vinnige agteruitgang van AY geaffekteerde Chardonnay, wat uiteindelik gelei het tot die afsterf van wingerde, is waargeneem, wat die sensitiwiteit van Chardonnay teenoor wingerdvergeling infeksie bevestig. Simptoomlose astervergeling fitoplasma infeksies kom voor en astervergeling fitoplasma kon nie opgespoor word in alle simptomatiese wingerdstokke nie, wat op oneweredige verspreiding van AY fitoplasma in individuele wingerdstokke dui. Molekulêre ontledings met behulp van PKR-RFLP het getoon dat alle wingerdstokke, wat in die Vredendal omgewing getoets is, slegs astervergeling fitoplasma bevat. Geen fitoplasmas was teenwoordig in enige onkruide of ander moontlike gasheer plante. Hoewel die gemiddelde jaarlikse siekte voorkoms van Chardonnay (29,95%) en Chenin Blanc (16,64%) oor die vier-jaar opname periode hoër was as dié van Pinotage (5,80%), was daar geen statisties beduidende verskil tussen die siekte voorkoms van hierdie drie kultivars nie. Die gemiddelde jaarlikse siekte voorkoms het 'n tendens oor tyd getoon, en die siekte voorkoms van die eerste jaar was betekenisvol laer as dié van die ander jare. Chardonnay het ‘n kumulatiewe siekte voorkoms van 37.77% aan die einde van die 4-jaar studie getoon, wat beteken dat Chardonnay wingerde binne 10 jaar 100% besmet kan wees met AY. Verspreidingspatrone van AY geaffekteerde wingerdstokke was meestal nie-ewekansig met bondeling van geaffekteerde wingerdstokke in en oor wingerd rye. Bondeling van AY geaffekteerde wingerdstokke het, met die uitsondering van een wingerd, meestal op die kant van wingerde aanliggend aan besmette wingerde, voorgekom. Die epidemiologiese studie gee 'n aanduiding van die sensitiwiteit van die verskillende kultivars ten opsigte van AY, die tempo van die verspreiding en die toekomstige impak van die siekte op die Suid-Afrikaanse wynbedryf. Dit dra ook waardevolle inligting by tot die ontwikkeling van 'n strategie vir die bestuur van wingerdvergeling siekte in Suid-Afrikaanse wingerde.

Thesis (MSc)--Stellenbosch University, 2015. / ENGLISH ABSTRACT: Grapevine leafroll disease (GLD), caused by the members of the family Closteroviridae, is one of the most economic important viral diseases affecting grapevine. Grapevine leafroll associated virus 3 (GLRaV-3), of the genus Ampelovirus, is the most widespread member of the leafroll associated virus family. To prevent the spread of GLD, management strategies such as rogueing and insect vector control are required to limit crop losses. Alternative control strategies based on genetic modification of the grapevine genome, such as pathogen-derived resistance (PDR), is proven to be effective in conferring resistance to several viruses. Therefore, the focus of this study was to evaluate pathogen-derived resistance strategies for GLRaV-3 using the following two approaches; 1) evaluation of transgenic plants expressing a dysfunctional GLRaV-3 heat shock protein 70 homolog (HSP70h) in order to confer resistance against GLRaV-3, and 2) the construction of artificial microRNAs (amiRNAs) to use as a tool for silencing specific sequences of GLRaV-3 in the grapevine host and the development of an amiRNA-mediated silencing validation system. In the first part of this study, six transgenic plant lines (plant lines #1, #3, #9, #14, #15 and #17) as well as a non-modified plant line, were inoculated with GLRaV-3 by grafting buds of each onto GLRaV-3 infected plant material. After approximately five months, GLRaV-3 virus titres of all grafted plants were quantified relative to two reference genes using RT-qPCR. Results were evaluated by comparing the relative virus titre of each transgenic plant line to that of the non-modified control plant line. Results showed that resistance levels of plant line #3 was significantly enhanced (>99%) and remarkably, plant line #14, showed to be more susceptible to the virus. The second part of the study was the construction and validation of amiRNAs targeting GLRaV-3 sequences. Two 21 nt regions of GLRaV-3 were successfully incorporated into miRNA backbone vvi167b of grapevine. Moreover, target constructs were developed by incorporating corresponding GLRaV-3 target sequences into the 3’ UTR of a green fluorescence protein (GFP) gene. Subsequently, the target constructs were co-infiltrated with the constructed amiRNA in Nicotiana benthamiana and GFP expression levels were quantified to determine the silencing efficiency of the amiRNAs. Results showed that the amiRNAs were successful in silencing the GFP target construct, however, they were not specific in silencing exclusively their corresponding target. These amiRNA constructs are ideal for further viral studies to determine the efficiency of silencing GLRaV-3 in GLD infected grapevines. / AFRIKAANSE OPSOMMING: Wingerd rolblaar siekte (GLD), wat veroorsaak word deur die lede van die familie Closteroviridae, is een van die ekonomies mees belangrike virus siektes van wingerd. Grapevine leafroll-associated virus 3 (GLRaV-3), van die genus Ampelovirus, is die mees wydverspreide lid van die rolblaar geassosieerde virus familie. Om die verspreiding van GLD te voorkom, is bestuur strategieë, soos die verwydering van geïnfekteerde plante en insekvektor beheer, ’n vereiste om oes verliese te beperk. Alternatiewe beheer strategieë gebaseer op genetiese modifikasie van die wingerdgenoom, soos patogeen-afgeleide weerstand (PDR), is bewys om effektief te wees in die verlening van weerstand teen verskeie virusse. Daarom was die fokus van hierdie studie om patogeen-afgeleide weerstand strategieë vir GLRaV-3 te evalueer met behulp van die volgende twee benaderings; 1) die evaluering van transgeniese plante wat 'n disfunksionele GLRaV-3 hitte-skok proteïen 70 homoloog (HSP70h) uitdruk, ten einde weerstand te verleen teen GLRaV-3, en 2) die konstruksie van kunsmatige mikroRNAs (amiRNAs) om te gebruik as 'n instrument vir die ondrukking van spesifieke genoomvolgordes van GLRaV-3 in die wingerd gasheer en die ontwikkeling van ’n stelsel om amiRNA-bemiddelde onderdrukking te bevestig. In die eerste deel van hierdie studie, is ses transgeniese plant lyne (plant lyne # 1, # 3, # 9, # 14, # 15 en # 17) sowel as 'n nie-gemodifiseerde gesonde plant lyn, geïnokuleer met GLRaV- 3 deur enting van ogies van elk op GLRaV-3 besmette plantmateriaal. Na ongeveer vyf maande, is GLRaV-3 virus konsentrasies van alle ingeënte plante gekwantifiseer relatief tot twee verwysing gene deur gebruik te maak van tru-transkripsie kwantitatiewe PCR (RTqPCR). Resultate is geëvalueer deur die relatiewe virus konsentrasie van elke transgeniese plant lyn te vergelyk met dié van die nie-gemodifiseerde kontrole lyn. Resultate het getoon dat weerstand vlakke van plant lyn # 3 beduidend verbeter is (> 99%) en merkwaardig is plant lyn # 14 bewys om meer vatbaar vir die virus te wees. Die tweede deel van die studie was die konstruksie en bevestiging van kunsmatige mikroRNAs (amiRNAs) wat GLRaV-3 genoomvolgordes teiken. Twee 21 nt streke van GLRaV-3 is suksesvol geïnkorporeer in die ruggraat van wingerd mikroRNA vvi167b. Verder is teiken konstrukte ontwikkel deur die inkorporering van ooreenstemmende GLRaV-3 teiken genoomvolgordes in die 3'UTR (3’ ongetransleerde area) van 'n groen fluoressensie proteïen (GFP) geen. Daarna is die teiken konstrukte gesamentlik geïnfiltreer met die gekonstrueerde amiRNA in Nicotiana benthamiana en GFP uitdrukkingsvlakke is gekwantifiseer deur die onderdrukkingsdoeltreffendheid van die amiRNAs te bepaal. Resultate het getoon dat die amiRNAs suksesvol was in die onderdrukking van die GFP teiken konstruk, maar hulle was egter nie-spesifiek in die eksklusiewe onderdrukking van die ooreenstemmende teiken. Hierdie amiRNA konstrukte is ideaal vir verdere virus studies om die doeltreffendheid van GLRaV-3 onderdrukking in GLD besmette wingerdstokke te bepaal.

Thesis (MScAgric) -- University of Stellenbosch, 2002. / ENGLISH ABSTRACT: The table grape industry is a major contributor to the South African economy, directly through foreign earnings from this predominantly export-based industry, as well as indirectly through the employment of thousands of people. It is a growing industry and consists of several production areas. The fastest growing table grape production area in South Africa is the Lower Orange River area, which produces some of the earliest grapes in the Southern Hemisphere. The biggest river in South Africa irrigates this area and it has an extreme climate characteristic of semi-desert areas. This area is considered to be optimal for the production of high quality, early, white seedless grapes. Previously, this area was predominantly planted to Sultanina vines for the purpose of raisin production. When seedless table grapes became a consumer preference, the producers very successfully converted their production practices to yield export quality seedless grapes from the established Sultanina vineyards. Extensive new plantings as well as re-plantings occurred in this area, also including newer cultivars from local and overseas breeding programmes. Being a viticultural and economical hot-spot, the Lower Orange River area is attracting much attention as a table grape production area and it also formed the backdrop to this study. The cultivar profile is changing in the area and it is projected that Sultana-, Regal-, Prime Seedless and Sugraone will be the four major early, white seedless cultivars in 2005. Based on this knowledge and prompted by a lack of information regarding production costs and general profitability of the new cultivars, this study was initiated in the form of a comparative analysis between the four mentioned cultivars spanning the early, middle and late regions of the Lower Orange River area. The approach used extracted information regarding cultural input costs (specifically labour as man-hours and the consequent costs) per manipulation performed in the vineyards. This approach is different from the more general method of obtaining input costs for a specific area based on combined mean values, often not distinguishing between cultivars. The specific aims of the study included a comparative analysis of input costs for production cultural practices per main manipulation action, as well as a comparative analysis taking into account productivity, value and extraordinary costs related to each of the four cultivars. To this end, 22 experimental plots were identified for use in the study. Collaboration of the production managers of each of the experimental plots were procured and information regarding production costs per manipulation and productivity of each cultivar and experimental plot were extracted from their own record keeping systems or from documents provided to the production managers. The value (price achieved) of the various cultivars for the 2001/2002 table grape season were put into perspective by using data from a survey which included information regarding payments for the various cultivars during the season in the Lower Orange River area. Information regarding fruit and vine royalties was obtained from the various plant breeders' rights holders of the various cultivars, where applicable. Primary descriptions of each experimental plot concerning general cultivation practices and information regarding the specific season were used to qualify results obtained from the various blocks. Several complicating factors impacted on the study and specifically the subsequent analyses of the results. Some of the factors were already identified as complicating factors in the planning stage of the study and were mostly linked to the recent introduction of two of the cultivars to the Orange River area. From the data gathered and the analyses performed it became clear that it would be difficult to discern significant differences (where significant is defined as PS0.10), but clear trends were observed and indications obtained. Based on the input cost analyses of this study it is proposed that mature Prime Seedless will have the highest labour input and cultural production cost of the four cultivars, followed by Sugraone. The labour input and the cost for the production cultural practices studied for young Prime Seedless vines were very high in comparison to the mature Sultana Seedless and Sugraone vines, especially for the canopy management and bunch manipulation actions. Prime Seedless was especially prone to the set of small and uneven berries, which lead to very high labour input requirements and subsequent cost for bunch manipulations. Sugraone is also known for the set of small and uneven berries in the Lower Orange River area, especially in difficult climatic seasons, also requiring high labour input for bunch manipulations. The initial indication is that mature Sultana- and Regal Seedless will require similar labour inputs for cultural production practices. The fact that Regal Seedless does not require expensive gibberellic acid (GA) applications, or girdling for thinning and berry enlargement purposes, is a tremendous advantage from a production cost point of view. Accordingly, initial indications are that Regal Seedless will have the lowest cultural production cost of the four cultivars. Regal Seedless was prone to the set of uneven berries during the year of study and accordingly it is suspected that this factor will ultimately determine the labour requirements and cultural production input cost, especially in difficult climatic seasons. The labour input and ultimately the cultural production cost for Sultana Seedless will be determined by the correct timing and concentration of the GA applications for thinning and berry sizing. Sultana Seedless and Sugraone produced high yields during the 2001/2002 table grape season in the Lower Orange River area. Yield information from the various experimental plots confirmed that there is little to choose between the two cultivars in terms of yield when cultivation conditions and practices are optimal. Large variation was observed in the yield results from the Regal- and Prime Seedless experimental plots. This is largely due to the recent introduction of the cultivars to the area and the consequent scarcity of blocks of these cultivars that are in full production. It was impossible to identify clear trends in terms of the future productivity of mature Regal- and Prime Seedless, but some indications of labour inputs could be extracted and qualified. Early maturing Prime Seedless and Sugraone performed very well in terms of price, especially in the harvest period prior to week 50. This advantage of high prices early in the season is, however, not always applicable to early cultivars in the later maturing regions of the Lower Orange River area. Later during the season, after week 50, when the supply of table grapes to the overseas markets has increased sharply, Sultana Seedless is usually the best performer in terms of price of the four cultivars. The ultimate price obtained by a cultivar is to a large extent determined by supply and demand, quality and acceptance of the specific cultivar. This study and its outcomes have a strong regional (Lower Orange River) and local (South Africa) impact and the specific results will undoubtedly be valuable to the producers, exporters and other role-players with vested interest in the cultivars studied or in table grape production per se. The methodology adopted in this study, however, is of broader interest and dearly shows the advantage of having detailed and qualified information regarding cultivation practices and bringing it in relation to the labour and consequent costs required per action. This should lead to more business intelligence and realistic planning on the producer side when decisions regarding the choice of a cultivar for a specific production area with a particular marketing scope have to be made. This study has also paved the way for similar studies, specifically with regard to the detailed description of the methodology that was established. Knowledge of the problems experienced in this study provides a useful reference for the planning and execution of similar studies. / AFRIKAANSE OPSOMMING: Die tafeldruifindustrie dra grootskaals by tot die Suid- Afrikaanse ekonomie: regstreeks deur middel van buitelandse valuta vanaf hierdie hoofsaaklik uitvoer-gebaseerde industrie, asook indirek deur werkverskaffing aan duisende mense. Dit is 'n vinnig groeiende industrie en bestaan uit verskeie produksie-areas waarvan die Benede-Oranjerivierarea, waar van die vroegste druiwe in die suidelike halfrond geproduseer word, tans die meeste groei toon. Die grootste rivier in Suid-Afrika vloei deur hierdie gebied wat deur uiterste klimaatstoestande, soortgelyk aan die van semi-woestyngebiede, gekenmerk word. Hierdie gebied is baie gunstig vir die produksie van hoë-gehalte, vroeë, wit pitlose druiwe. In die verlede is hoofsaaklik Sultanina vir die produksie van rosyne in hierdie gebied verbou. Namate pitlose tafeldruiwe voorkeur begin geniet het onder verbruikers wêreldwyd, het produseerders in die area hul verbouingspraktyke suksesvol aangepas vir die produksie van uitvoergehalte tafeldruiwe vanaf die grootskaalse, reeds gevestigde Sultanina-wingerde. Uitgebreide aanplantings en heraanplantings, wat nuwe cultivars van plaaslike en oorsese teelprogramme ingesluit het, is in hierdie gebied gedoen. Die vinnige groei in tafeldruifaanplantings en -uitvoere, asook die ekonomiese impak van die industrie in die Benede-Oranjeriviergebied, het die afgelope aantal jaar sterk op die voorgrond getree en het gevolglik gedien as agtergrond vir hierdie studie. Die cultivarprofiel in dié area is besig om te verander en volgens vooruitskattings gaan Sultana, Regal, Prime Seedless en Sugraone die vier prominente vroeë, wit, pitlose tafeldruifcultivars in 2005 wees. Gebaseer op hierdie feit en na aanleiding van 'n behoefte aan meer inligting met betrekking tot produksiekostes en algemene winsgewendheid van die nuwe cultivars, is 'n vergelykende studie aangaande die vier genoemde cultivars in die Benede-Oranjeriviergebied geloods. Die benadering wat gedurende die studie gevolg is, het inligting aangaande produksie-insetkoste (spesifiek arbeid in man-ure en gevolglike koste) per manipulasie onttrek. Hierdie benadering verskil van die meer algemene metodiek om insetkoste-inligting van 'n spesifieke area van gekombineerde gemiddelde waardes te verkry. Met so 'n benadering word gewoonlik geen onderskeid tussen cultivars getref nie. Die spesifieke doelwitte van hierdie studie het 'n vergelykende analise aangaande die insetkoste van die produksiepraktyke per hoofmanipulasie/aksie ingesluit, asook 'n analise waar produktiwiteit, waarde en buitengewone koste van die vier cultivars in ag geneem is. In totaal is 22 eksperimentele persele gebruik in die studie. Samewerking van die produksiebestuurders van die onderskeie esperimentele persele is verkry ten opsigte van die verskaffing van inligting oor produksiekoste per manipulasie, en die produktiwiteit per cultivar en eksperimentele perseel. Die produksiebestuurders het die nodige dokumente ontvang om die inligting te onttrek, of kon die inligting verskaf soos dit in hul rekordhoudingsisteem voorgekom het. Die waarde (prys behaal) van die onderskeie cultivars vir die 2001/2002-seisoen is in perspektief gestel deur gebruik te maak van 'n opname wat in die Benede Oranjeriviergebied plaasgevind het. Hierdie opname het inligting oor die uitbetalings van die onderskeie cultivars in die area vir die 2001/2002- seisoen ingesluit. Inligting rakende die stok- en vrugproduksie-tantieme is vanaf die onderskeie plantttelersregtehouers van die cultivars verkry. Primêre beskywings van die algemene verbouingspraktyke van elke eksperimentele blok en inligting oor die spesifieke seisoen is gebruik om die data wat vanaf die esperimentele persele verkry is, in perskektief te stel. Verskeie kompliserende faktore het die studie en die ontleding van data beïnvloed. Verskeie van hierdie faktore is reeds geidentifiseer met die beplanning van die studie en was meestal gekoppel aan die onlangse bekendstelling van Regal en Prime Seedless aan die Benede-Oranjeriviergebied. Na aanleiding van die data wat ingesamel en ontleed is, was dit duidelik dat dit moeilik sou wees om betekenisvolle verskille (waar "betekenisvol" as PS0.10 gedifinieer is) tussen die cultivars uit te lig, maar dat dit egter wel moontlik sou wees om aanvanklike indikasies en tendense te kry. Gebaseer op die insetkoste-ontleding van die studie blyk dit dat volwasse Prime Seedless die hoogste arbeidsinsetle en produksiekoste van die vier cultivars gaan hê, gevolg deur Sugraone. Die arbeidsinsetle en koste van die produksie-aksies wat van jong Prime Seedless bestudeer is, was baie hoog in vergelyking met volwasse Sultana Seedless- en Sugraone-stokke, veral ten opsigte van lowerbestuur en trosmanipulasies. Prime Seedless was veral geneig tot die set van klein, oneweredige korrels, wat tot baie hoë arbeidsinsetle en gevolglik koste vir trosmanipulasies gelei het. Sugraone is ook daarvoor bekend dat dit geneig is tot die set van klein, oneweredige korrels in die Benede-Oranjeriviergebied (veral in moeilike klimaatseisoene), wat gevolglik tot hoë arbeidsinstle vir trosmanipulasie lei. Die aanvanklike aanduiding is dat volwasse Sultana en Regal Seedless min of meer die dieselfde arbeidsinsetle vir verbouing sal vereis. Die feit dat Regal Seedless nie duur gibberelliensuur (GS)-behandelings vir blomtrosuitdunning of korrelvergroting benodig nie, is 'n enorme voordeel in terme van produksiekoste. Gevolglik is die aanvanklike aanduiding dat Regal Seedless die laagste produksieskoste van die vier cultivars sal hê. In die studiejaar was Regal Seedless egter geneig tot die set van onweredige korrels en gevolglik word verwag dat hierdie faktor uiteindelik die arbeidsinsetle en produsiekoste van die cultivar sal bepaal, veral in moeilike klimaatseisoene. Die arbiedsinsetle en produksiekoste van Sultana Seedless sal bepaal word deur die korrekte tydsberekening en konsentrasie van die GS-behandelings vir uitdunning en korrelvergroting. Sultana Seedless en Sugraone het gedurende die 2001/2002-seisoen hoë opbrengste in die Benede-Oranjeriviergebied geproduseer. Oesdata inligting van die onderskeie esperimentele persele het bevestig dat daar min te kies is tussen die twee cultivars in terme van produktiwiteit wanneer verbouingstoestande en -praktyke optimaal is. Groot variasie is egter waargeneem in die opbrengsresultate van die Regal en Prime Seedless. Dit is hoofsaaklik as gevolg van die onlangse bekendstelling van die twee cultivars in die area en dus ook die beperkte aantal blokke van die cultivars wat reeds in vol produksie was. Dit was dus onmoontlik om duidelike tendense in terme van die toekomstige produksie van volwasse Regal en Prime Seedless te identifiseer. Indikasies van arbeidsinsette en produksiekoste kon egter wel verkry word. Vroeg rypwordende Prime Seedless en Sugraone vaar baie goed in terme van die prys wat dit behaal, veral in die oesperiode voor week 50. Hierdie voordeel van hoë pryse behaal vroeg in die seisoen is egter nie altyd van toepassing op vroeë cultivars in die later rypwordende areas van die Benede-Oranjeriviergebied nie. Later in die seisoen (na week 50), wanneer die aanbod van tafeldruiwe op oorsese markte skerp toegeneem het, is Sultana Seedless gewoonlik die beste presteerder in terme van prys van die vier cultivars. Die uiteindelike prys wat deur cultivars behaal word, word tot 'n groot mate bepaal deur vraag en aanbod, kwaliteit en aanvaarding van die cultivar deur die verbruiker. Die studie en die uitkomste daarvan het 'n sterk streeks (Benede-Oranjerivier) en plaaslike (Suid-Afrika) impak, en die spesifieke resultate salongetwyfeld van waarde wees vir produseerders, uitvoerders en ander rolspelers met bestaande belange in die cultivars of vir tafeldruifproduksie as sulks. Die metodiek wat in hierdie studie gebruik is, is egter van breêr belang en wys duidelik die voordele daarvan om gedetailleerde en gekwalifiseerde inligting aangaande produksiepraktyke te hê, wat dit ook in verband bring met arbeid en gevolglike koste per aksie. Dit behoort te lei tot meer besigheidsintelligensie en realistiese beplanning deur die produseerder met betrekking tot cultivarkeuse vir 'n spesifieke produksiearea met 'n spesifieke bemarkings geleentheid. Hierdie studie het ook die weg gebaan vir soortgelyke studies, spesifiek ten opsigte van die gedetailleerde beskrywing van die metodiek wat gevestig is. Kennis van die probleme wat in hierdie studie ondervind is, kan dien as nuttige verwysing vir die beplanning en uitvoer van soortgelyke studies.

Thesis (MScAgric)--University of Stellenbosch, 2002. / ENGLISH ABSTRACT: In an attempt to reduce the pH of juice and wine, different fertiliser applications and canopy management practices were evaluated in a field trial. Fertiliser treatments consisted of no, CaS04, Ca(OH)2, and MgS04 fertilisation. Canopy management was as follows: suckering (leaving only two shoots per bearer), tipping, vertical shoot positioning and removal of lateral shoots and yellow leaves in the bunch zone (Canopy 1); suckering (leaving three shoots per bearer), vertical shoot positioning as well as topping (Canopy 2); vertical shoot positioning and topping (Canopy 3). The field trial was conducted in the Paardeberg region on the farms Meerlus and Kersfontein. The vineyard at Meerlus was Cabernet franc/R99 with a high canopy density and a good root distribution, established on a sandy loam soil of granite origin, with a low subsoil pH and a high K content. The vineyard at Kersfontein was Cabemet Sauvignon/101-14 Mgt with a lower canopy density and a less extensive root distribution, also established on a sandy loam soil of granite origin, but with a low top- and subsoil pH and a higher K content. Fertilisation had no significant influence on the K content of juice and wine. Fertilisation with Ca and Mg reduced the pH of juice significantly in the case of Meerlus. In contrast, Mg fertilisation increased the pH of juice significantly at Kersfontein. Lastly, fertilisation had no significant effect on the pH of the wine. The K content of the juice at Meerlus was significantly reduced by Canopy 3 in comparison with Canopy 1 and 2. However, in contrast with Canopy 1 and 3, Canopy 2 significantly increased the pH of juice at Meerlus. The K content of the juice at Kersfontein was significantly reduced by Canopy 1, compared to Canopy 2 and 3, with no significant effect on the pH of the juice. Canopy management had no significant effect on wine pH. It appears to be possible to reduce the pH of juice in the case of Cabernet franc/99R, situated on granite soils, by means of Ca and Mg fertilisation. Because Ca(OH)2 and MgS04 fertilisation increased the maturity of Cabernet Sauvignon/101-14 Mgt grapes, it was impossible to evaluate the effect of fertilisation on the pH of juice at Kersfontein. The general phenomenon that a high canopy density results in a high pH of juice and wine, was not observed in this field trial. The possible reasons for this were the low canopy density of the control plots, as well as the difference in maturity between canopy management treatments. / AFRIKAANSE OPSOMMING: In 'n poging om die pH van sap en wyn te verlaag, is die effek van verskillende bemesting- en lowerbestuursfaktore in 'n veldproef geêvalueer. Bemesting het bestaan uit geen, CaS04, Ca(OH)2 en MgS04 toediennings. Lowerbestuur was: suier tot twee lote per draer, tip, vertikale lootposisionering, verwydering van sylote en geel blare in trossone (Lower 1); suier tot drie lote per draer, top en vertikale lootposisionering (Lower 2); top en vertikale lootposisionering met geen suier nie (Lower 3). Die veldproef is op twee plase nl. Meerlus en Kersfontein, in die Paardeberg omgewing uitgevoer. Die wingerd by Meerlus was Cabernet franc/R99 met 'n hoë lowerdigtheid en goeie wortelverspreiding, wat op 'n sandleemgrond van graniet oorsprong met In lae ondergrond-pH en hoë K-inhoud gevestig is. Die wingerd by Kersfontein het bestaan uit Cabernet Sauvignon/101-14 Mgt met 'n laer lowerdigtheid en swakker wortelverspreiding, wat op In sandleemgrond van graniet oorsprong met In deurgaans lae grond-pH en baie hoë K-inhoud gevestig is. Bemesting het geen betekenisvolle invloed op die K-inhoud van sap en wyn gehad nie. By Meerlus het Ca- en Mg-bemesting egter die pH van sap betekenisvol verlaag. In teenstelling hiermee het Mg-bemesting die sap-ph by Kersfontein betekenisvol verhoog. Bemesting het verder geen betekenisvolle invloed op die pH van wyn gehad nie. Lower 3 het die K-inhoud van sap by Meerlus betekenisvol verlaag in vergelyking met Lower 1 en Lower 2. By Kersfontein was die K-inhoud van sap by Lower 1 betekenisvollaer as by Lower 2 en Lower 3. Teenoor Lower 1 en Lower 3 het Lower 2 'n betekenisvol hoër sap-pH by Meerlus tot gevolg gehad. Lowerbehandelings het egter geen betekenisvolle invloed op die pH van wyn gehad nie. Die moontlikheid bestaan dus om die sap-pH van Cabernet franc/R99 op granietgrond betekenisvol m.b.v. Ca- en Mg-bemesting te verlaag. Aangesien Ca(OH)2- en MgS04- bemesting die rypheidsgraad van Cabernet Sauvignon/101-14 Mgt se druiwe betekenisvol verhoog het, was dit onmoontlik om die effek van bemesting op sap-pH by Kersfontein te evalueer. Die algemene verskynsel dat 'n hoë lowerdigtheid tot hoë pH's in sap en wyn lei, is nie in die proef ondervind nie. Die lae lowerdigtheid van die kontrole persele en die verskil in rypheidsgraad tussen lowerbehandelings kan moontlik as rede hiervoor aangevoer word.

Thesis (MScAgric)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Plant carotenoid cleavage dioxygenases (CCD) are a family of enzymes that catalyse the oxidative cleavage of carotenoids and/or apocarotenoids. Carotenoids are synthesised in plastids (primarily chloroplasts and chromoplasts), where they are involved in light-harvesting and protecting the photosynthetic apparatus from photo-oxidation. The carotenoid-derived apocarotenoids fulfil a number of roles in plants such as phytohormones, pollinator attractants and flavour and aroma compounds. Due to the floral and fruity characteristics that apocarotenoids contribute to wine, these C13 compounds have received interest in grapevine (Vitis vinifera L.). The CCD gene family in Arabidopsis consists of nine members, all encoding for enzymes that catalyse the cleavage of carotenoids. The enzymes in this family include 9-cis-epoxydioxygenases (NCEDs) and four classes of CCD. NCEDs and CCD7 and CCD8 are involved with plant hormone synthesis, e.g. abscisic acid (ABA) through cleavage by NCED and strigolactone (SL) through the sequential cleavage of carotenoids by CCD7 and CCD8, respectively. SLs are a fairly new class of plant hormone which are involved in several aspects of plant growth and development. The most extensively characterised role of SLs is their involvement in the inhibition of shoot-branching. CCD1 and CCD4 cleave a variety of carotenoids to form pigments and aroma compounds. For example, CCD1 forms β-ionone and β-damascenone, which are important varietal flavours of wine, and CCD4 is involved in synthesis of the pigment and aroma compounds of saffron and annatto. CCD1 enzymes symmetrically cleave the 9,10 (9’,10’) double bonds of multiple carotenoids to produce a C14 dialdehyde and two C13 products. Additional CCD1 cleavage activity at 5,6 (5’,6’) double bonds of lycopene has been reported. Previous studies have shown that CCD1 isolated from V. vinifera (VvCCD1) was able to cleave multiple carotenoid substrates in vitro, namely zeaxanthin, lutein and β-carotene at 9,10 (9’,10’) double bonds and both the 5,6 (5’,6’) and 9,10 (9’,10’) double bonds of lycopene. None of the other VvCCDs, except VvCCD4a have been isolated (but no functionality was illustrated) and characterised yet. CCD4 enzymes also cleave carotenoids at the 9,10 (9’,10’) double bond positions. The presence of plastid-target peptides implies that the CCD4 enzymes have continuous access to carotenoids. Therefore it is suggested that CCD4s are responsible for carotenoid maintenance, where CCD1s contribute towards volatile production. To test this hypothesis VvCCD1, VvCCD4a and VvCCD4b were isolated from V. vinifera (cv Pinotage) cDNA and cloned into a pTWIN1 protein expression vector. Substrate specificity of each VvCCD was tested by co-transforming a carotenoid accumulating E. coli strain with a CCD expression vector. Carotenoids synthesized by the bacteria were identified and quantified by UPLC-analysis, while the concentration of the apocarotenoids, were measured in the headspace of the bacterial cultures using HS-SPME-GC-MS. Several optimisations were done to minimize the natural degradation of the carotenoids; to ensure that the apocarotenoid formation is predominantly due to the enzymatic cleavage by the VvCCDs and not due to oxidation or other non-enzymatic degradation. The HS-SPME-GC-MS analysis indicated that all isoforms cleaved phytoene, lycopene and ε-carotene. Additionally VvCCD1 cleaved a carotenoid involved in photosynthesis, namely β-carotene, while VvCCD4a cleaves neurosporene and VvCCD4b cleaves neurosporene and ζ-carotene, carotenoids not involved in photosynthesis. This study has illustrated that VvCCD1 cleave carotenoids necessary for photosynthesis and VvCCD4s cleave carotenoids which were not present in berry tissue, suggesting their role in carotenoid maintenance. Therefore in planta substrates for CCD1 could possibly be C27 apocarotenoids generated from enzymatic cleavage through CCD4 (role in carotenoid maintenance), CCD7 and/or photo-oxidation, which are then transported from the plastid to the cytosol or possibly C40 carotenoids that are released during senescence or when the plastid membrane is damaged, thus releasing important aroma compounds. Thus the identification of the in vivo substrates has contributed to the understanding the in planta functions of these enzymes / AFRIKAANSE OPSOMMING: Die plant ensiemfamilie van karotenoïedsplitsingdioksigenases (CCDs) kataliseer die oksidatiewe splitsing van karotenoïede en/of apokarotenoïede. Karotenoïede word in plastiede (primêr chloroplaste en chromoplaste) sintetiseer en is betrokke by lig-absorpsie en die beskerming van die fotosintetiese apparaat teen foto-oksidasie. Die apokarotenïede afkomstig van karotenoïede dien onder meer as planthormone, geur- en aromakomponente en om bestuiwers aan te lok. Aangesien apokarotenoïede bydra tot die vrug- en blomgeure van wyn is die C13-verbindings binne wingerd (Vitis vinifera L.) van belang. Al nege lede van die CCD geenfamilie in Arabidopsis kodeer karotenoïedsplitsingsensieme. Die ensiemfamilie sluit 9-sis-epoksidioksigenases (NCEDs), en vier klasse CCD in. NCEDs en CCD7 en 8 is betrokke by die sintese van planthormone, naamlik absissiensuur (ABA) deur NCED en strigolaktone (SL) deur die opeenvolgende aksie van onderskeidelik CCD7 en CCD8. SLe is redelik onlangs as planthormone indentifiseer en is betrokke by ‘n verskeie aspekte van die groei en ontwikkeling van plante. Die rol van SL in inhibisie van vertakking is die beste gekarakteriseerde van hierdie aspekte. CCD1 en CCD4 splits ‘n verskeidenheid karotenoïede om pigmente en aromakomponente te vorm. CCD1 vorm byvoorbeeld β-jonoon en β-damasenoon, beide belangrike kultivar-spesifieke wyngeure. CCD4 vorm weer die pigment en aromakomponente van saffraan en annatto. Die CCD1 ensieme splits die 9,10 (9’,10’) dubbelbindingsetels van verskeie karotenoïede simmetries en vorm een C14-dialdehied en twee C13-produkte. Daar is voorheen melding gemaak van verdere splitsing deur CCD1 by die 5,6 (5’,6’) dubbelbindingsetels van likopeen. Vroeër is getoon dat die CCD1 isovorm wat uit V. vinifera geïsoleer is, naamlik VvCCD1, in vitro seaxantin, luteïen en β-karoteen by die 9,10 (9’,10’) dubbelbindingsetels kon splits, en likopeen by beide die 9,10 (9’,10’) en 5,6 (5’,6’) dubbelbindingsetels. Geen ander VvCCDs is al isoleer en funksioneel gekarakteriseer. VvCCD4a is isoleer, maar geen funksie is bepaal nie. CCD4 ensieme splits ook die 9,10 (9’,10’) dubbelbindingsetels van karotenoïede. Aangesien CCD4 ensieme ‘n plastied-bestemmingspeptied besit behoort dié ensieme konstant toegang tot karotenoïede te hê, wat dui op hul rol in die handhawing van die karotenoïedbalans, terwyl CCD1-ensieme bydra tot die sintese van vlugtige verbindings. Om hierdie hipotese te toets is VvCCD1, VvCCD4a en VvCCD4b uit V. vinifera (kv Pinotage) kDNS isoleer in binne ‘n pTWIN1 proteïenuitdrukkingsvektor kloneer. Die substraatspesifisiteit van elke VvCCD is getoets deur ‘n karotenoïedakkumulerende E. coil stam te transvormeer met ‘n CCD-uitdrukkingsvektor. UPLC-analise is gebruik om karotenoïede wat deur die bakterium sintetiseer is te kwantifiseer en identifiseer, terwyl die apokarotenoïedinhoud en -konsentrasie van die boruimte van die bakteriële kultuur met HS-SPME-GC-MS bepaal is. Verskeie aspekte van die proses is optimaliseer om natuurlike afbreking van karotenoïede te minimeer. Daardeur is verseker dat die apokarotenoïedvorming primêr vanweë die ensiematiese splitsing deur VvCCDs plaasvind en nie deur oksidasie of ander nie-ensiematiese afbreking. Die HS-SPME-GC-MS metings het aangedui dat al drie isovorme fitoëen, likopeen en ε-karoteen kan splits. VvCCD1 kan daarby β-karoteen splits, terwyl VvCCD4a neurosporeen, en VvCCD4b neurosporeen en ζ-karoteen kan splits, beide karotene wat nie betrokke is by fotosintese nie. Dié studie toon dat VvCCD1 die karotenoïede splits wat benodig word vir fotosintese, terwyl beide VvCCD4 isovorme karotenoïede splits wat nie in druiwekorrels gevind word nie. Dit dui op hulle rol in die handhawing van karotenoïedpoele. Die in planta substrate vir CCD1 mag dus die C27-apokarotenoïede wees wat deur CCD4 (as deel van karotenoïedhandhawing), CCD7 en/of foto-oksidasie gevorm word en na die sitosol vervoer word, of moontlik die C40-karotenoïede wat tydens veroudering óf wanner die plastiedmembraan beskadig is in die sitosol vrygestel word. Die identifisering van die in vivo substrate het dus bygedra to die begrip van die in planta funksies van die ensieme.

CAPES / O objetivo do trabalho foi avaliar a capacidade antioxidante de extratos hidroalcoólicos e microencapsulados, por spray dryer, de coprodutos do vinho e suco da uva das variedades Bordô e Niágara (Vitis labrusca) e aplicar em patê cremoso de carne de frango para avaliar a inibição da oxidação lipídica e a aceitação sensorial do produto. Os coprodutos da uva foram extraídos individualmente com etanol 80% em shaker, a 40 ºC/60 minutos, concentrado em evaporador rotativo e microencapsulado em spray dryer com maltodextrina 10 DE e amido modificado (Capsul®). As amostras pulverizadas em spray dryer sofreram influência do agente encapsulante utilizado, e as amostras microencapsuladas com maltodextrina 10 DE obtiveram maior eficiência. A microencapsulação dos coprodutos originou microesferas lisas, rugosas, homogêneas quanto à forma e estrutura e sem fissuras ou rachaduras. A análise de infravermelho com transformada de Fourrier (IVTF) mostrou que os compostos bioativos permaneceram nos extatos mesmo depois da microencapsulação. Os extratos hidroalcoólicos e microencapsulados apresentaram alta atividade antioxidante in vitro, sendo esta atividade atribuída à presença de compostos fenólicos totais, antocianinas e flavonoides totais. Pela técnica de Cromatografia Líquida de Alta Eficiência (CLAE) foi possível identificar e quantificar seis compostos fenólicos (ácido gálico, cafeico, cumárico, ferrúlico, vanílico e trans-resveratrol). O extrato do coproduto da uva bordô vinho, safra 2014 microencapsulado com maltodextrina (CUBV2M (MD)) e o extrato hidroalcoólico liofilizado coproduto uva bordô vinho, safra 2014 (CUBV2) foram escolhidos para aplicação em patê cremoso de frango por apresentar maior teor de compostos fenólicos com elevada atividade antioxidante. A formulação base do patê cremoso de frango foi elaborada com carne de frango, pele de frango e condimentos e foi dividida em quatro tratamentos: o primeiro foi designado como controle e nenhum ingrediente adicional foi incluído (T1). O segundo lote foi prepardo adicionando eritorbato de sódio (T2). O terceiro lote recebeu o extrato etanólico liofilizado CUBV2 (T3). O quarto lote recebeu o extrato microencapsulado CUBV2M (MD) (T4). A estabilidade oxidativa dos patês foi avaliada pelo teor de substâncias reativas ao ácido tiobarbitúrico (TBARS) e pela coloração no dia do processamento e semanalmente por 41 dias. Pela diferença total de cor (ΔE), em relação ao controle (T1), apenas o T4 apresentou influência visualmente perceptível, durante o período de estocagem. Os patês adicionados de 0,3% de CUBV2 e CUBV2M (MD) demonstraram resultados satisfatórios pela análise de TBARS e estavam microbiologicamente de acordo com à legislação vigente. Os índices de aceitabilidade para a avaliação global foram superiores a 70% para T1, T2 e T3 podendo estes extratos de coprodutos de vinho de uva ser considerados uma alternativa aos antioxidantes sintéticos em patês cremosos. / The objective of this study was to evaluate the antioxidant activity of hydroalcoholic extracts and microencapsulated by spray dryer, of different samples of co-products of wine and juice grape varieties Bordô and Niagara (Vitis labrusca) and apply creamy pate of chicken to verify the inhibition lipid oxidation and the acceptance of this product. The grape co-products were extracted individually with ethanol (80%) in shaker at 40 °C / 60 min, concentrated and microencapsulated in spray dryer with maltodextrin 10 DE and capsul®. The dried samples spray dryer were influenced by the encapsulating agent used, where the microencapsulated samples with maltodextrin obtained better efficiency. Microencapsulation of co-products originated smooth beads and other rough, homogeneous in form and structure, without fissures or cracks. The Infrared Fourier Transform analysis (FTIR) showed that the microencapsulated bioactive compounds in the extracts remained even after subjected to drying by spray drying. The hydroalcoholic and microencapsulated extracts showed high phenolic compounds, anthocyanins and flavonoids with antioxidant activity. For the HPLC technique it was possible to identify and quantify six patterns of phenolic compounds (gallic acid, caffeic, coumaric, ferulic, vanillic and trans-resveratrol). The CBGW2 and CBGW2M (MD) extracts were chosen to be applied in creamy pate due to the best levels of phenolic compounds with antioxidant activity. The basic formulation of creamy chicken pate was made with chicken meat, chicken skin and spices and was divided into four treatments: The first was designated as the control and no additional ingredients were included (T1). The second lot was designated as a positive control and was prepared by adding sodium eritorbate (T2). The third lot received liophilized extract CBGW2 (T3). The fourth lot received the CBGW2M (MD) extracts. The stability of chicken meat pates was performed on the day of processing and weekly at 41 days in 4º C by thiobarbituric acid reactive substances index (TBARS) and color. By (ΔE) in color analyses, compared to the control (T1), only the T4 presented visually perceptible influence during the storage period. The pates added 0.3% CBGW2 and CBGW2M (MD) showed satisfactory results for TBARS analysis and were microbiologically according to law. Acceptability levels for the overall evaluation were greater than 70% for T1, T2 and T3 and these extracts of grape wine co-products can be considered an alternative to synthetic antioxidants in pates creamy.

Made available in DSpace on 2015-04-17T14:49:44Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1284009 bytes, checksum: d97ba3b3d03fb907be34c234b54f032f (MD5) Previous issue date: 2010-09-30 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Phenolic compounds are widely found in the plant kingdom. Among the fruits considered as functional foods, grapes and their derivatives are always in a position of prominence, especially after the association of regular consumption of wine to a healthy life. This work was carried out in order to know the antioxidant properties and resveratrol concentration of grape juices and also of grape pomace samples, a byproduct of winemaking. The skins of wine grapes Vale São Francisco (varieties Syrah and Cabernet Sauvignon), the Syrah grape pomace and commercially available grape juices were studied for their profile of polyphenolic compounds and antioxidant activity. Samples were taken from sun-exposed grape Syrah, coded as sol and grape shade by the foliage canopy, coded as sombra ." The juice samples were composed 11integrals juices,8 reprocessed grape juice and 11 nectars. An extract of grape and pomace was prepared from 0,5mg of sample (dry weight) with 3 ml of a solution of ethanol / water (80/20 v / v). The concentration of the cis and trans-isomers of resveratrol were determined by using a chromatographic method (HPLC-UV/DAD with C8 chromatography column) and spectrophotometric methods were determined phenolic monomeric anthocyanins and DPPH radical scavenging activity (expressed as EC50 μg / mL ). The trans isomer of resveratrol was determined in samples of Syrah sombra , Cabernet Sauvignon, Syrah pomace and on 60% of the juices analyzed. The isomer cis of resveratrol was determined only on the samples Syrah grape sol and this isomer was not found in the nectar juice samples. The total phenolic for grapes ranged between 3,7 and 17,59 ± 0.05 ± 0,61 mg / mL for wine grapes and from 451,7 ± 4,68 and 1935,0 ± 15,06 g / mL for grape juices. Grape pomace was determined in 3268,0 ± 360.7 mg / mL (n = 3). Monomeric anthocyanins for grapes ranged from 1,47 ± 0,08 and 10,0 ± 0,56 mg / mL for wine grapes and between 6,88 ± 0,63 and 292,4 ± 27,7 mg/L the grape juices. In the pomace was determined 0,75 ± 0,07 mg/mL (n = 3). The Syrah grape pomace showed high antioxidant activity (EC50 1,09 μg/mL) compared to ascorbic acid (EC50 2,12 ± 0,0009 μg/mL) The Syrah grape sol and sombra showed significant differences regarding their ability to scavenge the DPPH radical. The results obtained allow us to conclude that the not only the polyphenolic content, but also their profile, influenced the antioxidant activity of the sample. The Syrah grape pomace had a very high content of total phenolics and radical-scavenging activity, and is thus a potential source of these substances that deserve to be used. / Os compostos fenólicos são largamente encontrados no reino vegetal. Dentre as frutas consideradas como alimentos funcionais, as uvas e seus derivados aparecem sempre em posição de destaque. Principalmente, após a associação do consumo regular do vinho a uma vida saudável. Este trabalho foi realizado com o objetivo de conhecer a propriedade antioxidante e concentração de resveratrol de uva, sucos de uva e bagaço, subproduto da vinificação. As cascas de uvas viníferas do Vale de São Francisco (das variedades Syrah e Cabernet sauvignon), o bagaço da uva Syrah e sucos de uva comerciais foram estudados quanto ao seu perfil de compostos polifenólicos e atividade antioxidante. Foram coletadas amostras de uva Syrah expostas ao sol, chamadas de Syrah sol e uvas sombreadas pelas folhagens do dossel vegetativo, chamadas Syrah sombra . As amostras de sucos eram compostas 11 sucos integrais, 8 sucos reprocessados e 11 néctares de uva. Foi preparado extrato das uvas e do bagaço a partir de 0,5mg da amostra (peso seco) com 3 ml de uma solução de Etanol/água (80/20 v/v). A concentração dos isômeros cis e trans do resveratrol foi determinada usando um método cromatográfico (CLAE-UV/DAD, com coluna cromatográfica C8) e por métodos espectrofotométricos foram determinados os fenólicos totais, as antocianinas monoméricas e a capacidade de reduzir 50% do radical DPPH (CE 50 μg/mL). Foi detectado o trans e cis-resveratrol nas uvas Syrah sombra, Cabernet sauvignon, bagaço de uva Syrah e em 60% dos sucos analisados. Foi determinado apenas o cisresveratrol na uva Syrah sol. Os néctares de uva não tiveram teor de cis-resveratrol detectado. Os fenólicos totais para as uvas variaram entre 3,7± 0,05 e 17,59± 0,61 μg/mL para as uvas viníferas e entre 451,7±4,68 e 1935±15,06 mgEAG/L para os sucos de uva. No bagaço foi determinado 3268 ± 360,7mgEAG/g (n=3). As antocianinas monoméricas para as uvas variaram entre 1,47± 0,08 e 10,0± 0,56 mg/mL para as uvas viníferas e entre 6,88±0,63 e 292,4 ± 27,7mg/L para os sucos de uva. No bagaço foi determinado 0,75 ± 0,07mg/mL (n=3). O bagaço de uva Syrah apresentou elevada atividade antioxidante (CE50 1,09μg/mL) comparada ao ácido ascórbico (CE50 2,12±0,0009μg/mL) A uvas Syrah sol e sombra apresentaram diferenças significativa quanto sua capacidade de seqüestrar o radical DPPH, respectivamente. Os resultados obtidos permitem-nos concluir que não apenas o conteúdo de polifenólicos, como também o seu perfil influenciam na atividade antioxidante da amostra. O bagaço de uva Syrah apresentou um elevado teor de fenólicos totais e atividade antiradicalar, sendo assim uma fonte potencial destas substâncias que merece ser melhor reaproveitado.

O Rio Grande do Sul é o maior produtor de uvas do Brasil, e as cultivares de Vitis labrusca representam mais de 90% da produção, dentre as quais se destacam as variedades Isabel e Bordô, que se caracterizam por conter altas concentrações de polifenóis, principalmente antocianinas, que exibem propriedades antioxidantes e que podem ser usados como corantes em alimentos. Entretanto, a utilização destes compostos em alimentos se torna difícil devido à sua alta instabilidade, e uma das formas de proteger estes compostos das condições adversas do meio é a microencapsulação. O bagaço da uva, proveniente da fabricação de sucos e vinhos, contém uma elevada concentração de polifenóis, e é composto em grande parte pela casca da uva. Dessa forma, este trabalho de Tese, teve como primeiro objetivo a extração dos polifenóis da casca das uvas Isabel e Bordô, e seu microencapsulamento utilizando diferentes materiais de parede, o qual foi dividido em três etapas. Na primeira etapa, foi realizada a separação dos compostos fenólicos mediante extração aquosa em meio ácido da casca de uva Isabel. A seguir, o extrato foi submetido à atomização para obtenção das micropartículas, utilizando goma arábica, β-ciclodextrina e hidroxipropil-β-ciclodextrina como agentes encapsulantes, combinadas em concentrações máximas de 5%. Os pós obtidos foram avaliados quanto aos fenóis totais, antocianinas monoméricas totais, flavonoides, flavanols, atividade antioxidante (DPPH, CUPRAC e HRSA), cor, umidade, atividade de água, solubilidade, higroscopicidade, temperatura de transição vítrea e microestrutura. De forma geral, os pós obtidos apresentaram alta higroscopicidade e baixa temperatura de transição vítrea, além de aglomeração das partículas. O tratamento elaborado com 3% de goma arábica e 2% de β-ciclodextrina foi considerado o melhor, com maior retenção de flavonoides (67,2%), flavanols (51,1%), atividade antioxidante por DDPH (55%) e CUPRAC (58,8%), menor higroscopicidade (17,33%) e maior temperatura de transição vítrea (32,85 °C). Na segunda etapa, o extrato fenólico aquoso acidificado da casca de uva Bordô foi atomizado e liofilizado para a obtenção das micropartículas, utilizando goma arábica, goma guar parcialmente hidrolisada e polidextrose, em um total de 10% de material de parede. Os pós obtidos foram avaliados quanto ao conteúdo de fenóis totais, antocianinas monoméricas totais, atividade antioxidante (DPPH, CUPRAC, HRSA), cor, umidade, atividade de água, solubilidade, higroscopicidade, temperatura de transição vítrea, tamanho de partícula e microestrutura. Foram obtidas altas retenções, maiores que 80% para fenóis totais e antocianinas monoméricas totais, e entre 45 e 84% para atividade antioxidante em todos os tratamentos estudados. Os pós atomizados tiveram menor umidade, atividade de água e tamanho de partícula, maior solubilidade e temperatura de transição vítrea, além de melhores características morfológicas do que os pós liofilizados. O pó obtido por atomização com 5% de goma guar parcialmente hidrolisada e 5% de polidextrose foi considerado o melhor tratamento, visto que teve maior retenção de fenóis totais (89,0%), antocianinas monoméricas totais (99,5%) e atividade antioxidante por DPPH (57,3%) e CUPRAC (83,2%). Na terceira etapa, dispersões de extrato de casca de uva Bordô com 5% de goma guar parcialmente hidrolisada e 5% de polidextrose, que foi considerado o melhor tratamento dentre todos os elaborados com uva Isabel e Bordô, foram atomizadas e liofilizadas para obtenção das micropartículas, que foram submetidas a testes acelerados de armazenamento (umidades relativas de 75 e 90% em temperaturas de 35, 45 e 55 °C) e de simulação de digestão gastrointestinal (divididos em duas fases: fase gástrica e fase intestinal). Foram avaliados os conteúdos de fenóis totais, antocianinas monoméricas totais e atividade antioxidante por ABTS. Quanto às provas aceleradas, a temperatura teve um efeito significativo na diminuição no conteúdo de fenóis, com retenções de 82,5% a 93,5%. Na redução do conteúdo de antocianinas monoméricas totais foi significativo o efeito da temperatura, umidade relativa e tempo, com retenções de 3,9 a 42,3%. A redução do teor de antocianinas monoméricas totais exibiu cinética de primeira ordem, e os valores de z e Ea indicaram que o pó liofilizado é mais instável às mudanças de temperatura, quando utilizadas temperaturas mais elevadas. Por outro lado, os valores de D e t1/2 foram muito próximos entre os dois pós, o que indica pouca diferença de estabilidade entre eles nas temperaturas utilizadas neste estudo. Os parâmetros termodinâmicos indicaram que a reação foi endotérmica e não espontânea. A atividade antioxidante teve comportamento similar ao dos fenóis totais, com retenção final de 38,5 a 59,5%. Quanto à simulação da digestão gastrointestinal os dois pós tiveram liberação de fenóis de aproximadamente 80% na fase gástrica, e aumento significativo da liberação na fase intestinal, onde, na última hora de experimento, o pó atomizado teve 90,6% de liberação e o liofilizado 94,9%. Comportamento similar foi observado para a atividade antioxidante, onde o pó atomizado e o pó liofilizado tiveram percentuais próximos a 50% na fase gástrica e aumento significativo na fase intestinal, onde na última hora do experimento o pó atomizado teve 69,4% da atividade antioxidante e o pó liofilizado 67,8%. Entretanto, as antocianinas monoméricas tiveram redução significativa de aproximadamente 50% do seu conteúdo na fase intestinal, onde, na última hora de experimento, o pó atomizado teve 39% de liberação e o liofilizado 39,8%. O método de obtenção das micropartículas não influenciou na estabilidade dos pós, tanto nos testes acelerados de armazenamento quanto na simulação da digestão gastrointestinal. Como segundo objetivo foi avaliado o efeito da irradiação UV-C em uvas Bordô, como tratamento pós-colheita. Foram estudadas duas safras de anos diferentes, sendo que na primeira as uvas foram submetidas a 0, 0,5, 1, 4, 10 e 30 minutos de irradiação (120 W) e armazenadas a 22 °C, enquanto que na segunda safra as uvas foram submetidas à irradiações com UV-C (120 W) por 0, 0,5 e 4 minutos, combinada com ultrassom (40 kHz) por 5 minutos e armazenadas a 22 °C. Na primeira safra, as uvas não apresentaram aumento dos compostos bioativos e da atividade antioxidante. Entretanto, na segunda safra, os resultados indicaram o aumento significativo no conteúdo de fenóis totais, antocianinas monoméricas totais e na atividade antioxidante para as uvas irradiadas por 0,5 minutos com UV-C e para as irradiadas com UV-C por 4 minutos em combinação com ultrassom por 5 minutos, com aumentos de 1,2 a 2,0 vezes em relação ao controle, não havendo mudanças significativas na cor das uvas irradiadas. / The state of Rio Grande do Sul, Brazil, is the greatest producer of grapes in the country. Vitis labrusca cultivars constitute more than 90% of production, underscoring Isabel and Bordô varieties, featuring high concentrations of polyphenols, especially antioxidant anthocyanins used as food coloring. Since the use of the compounds in food is rather difficult due to their unstableness, microencapsulation is one of the methods to protect the compounds from adverse environmental condition. Grape pomace, the residue from the manufacture of juice and wine, has high polyphenol concentration and is mainly formed by grape skin. Current thesis aims at extracting polyphenols from the skin of Isabel and Bordô grape varieties and their micro-encapsulation by different wall materials. Research was divided into three stages. The first stage comprised the separation of phenolic compounds by water extraction in the acid medium of the skin of the Isabel grape variety. The extract was spray-dried for microparticles by means of gum arabic, β-cyclodextrin and hydroxypropyl-β-cyclodextrin as encapsulating agents at maximum 5% concentrations. Powders were assessed for total phenols, total monomer anthocyanins, flavonoids, flavanols, antioxidant activity (DPPH, CUPRAC and HRSA), color, humidity, water activity, solubility, hygroscopicity, glass transition temperature and micro-structure. As a rule, the powders featured high hygroscopicity, low glass transition temperature and particle agglomeration. Treatment with 3% of gum arabic 3% and 2% of β-cyclodextrin was the best, with the highest retention rate of flavonoids (67.2%), flavanols (51.1%), antioxidant activity, and with DDPH (55%) and CUPRAC (58.8%), lowest hygroscopicity (17.33%) and highest glass transition temperature (32.85 °C). In the second stage the acidified water phenolic extract from the Bordô grape skin was spray-dried and freeze-dried for microparticles with gum arabic, partially hydrolyzed guar gum and polydextrose, with a total 10% of wall material. Powders were assessed for total phenols, total monomer anthocyanins, antioxidant activity (DPPH, CUPRAC, HRSA), color, humidity, water activity, solubility, hygroscopicity, glass transition temperature, particle size and micro-structure. High retentions occurred, with more than 80% for total phenols and total monomer anthocyanins; and between 45 and 84% for antioxidant activity in all treatments under analysis. Atomized powders had lower humidity, water activity and particle size, greater solubility, higher glass transition temperature, and better morphological characteristics than the freeze-dried powders. The powder obtain by spray-dried with 5% of partially hydrolyzed guar gum and 5% of polydextrose was the best treatment due to greater retention of total phenols (89.0%), total monomer anthocyanins (99,5%) and antioxidant activity for DPPH (57.3%) and CUPRAC (83.2%). The third stage comprised dispersions of the extract of the Bordô grape skin with partially hydrolyzed guar gum 5% and polydextrose 5%, or rather, the best treatment among those prepared with Isabel and Bordô grapes. Dispersions were spray-dried and freeze-dried to obtain microparticles which underwent fast storage tests (relative humidity rates 75 and 90% at 35, 45 and 55 °C) and gastrointestinal digestion simulations (divided into two phases: gastric and intestinal phases). Total phenols, total monomer anthocyanins and antioxidant activity for ABTS were evaluated. Fast tests revealed that temperature had a significant effect on the decrease in phenol contents, with 82.5 - 93.5% retentions. Temperature, relative humidity and time were significant on the reduction of total monomer anthocyanin contents, with 3.9 – 42.3% retentions. Decrease in total monomer anthocyanin rates revealed a first order kinetics, whilst z and Ea rates indicated highly unstable freeze-dried powder for changes in temperature when higher temperatures are employed. On the other hand, D and t1/2 rates were very close between the two powders, revealing slight stability difference between the two at temperatures employed in current study. Thermodynamic parameters demonstrated an endothermic and non-spontaneous reaction. Antioxidant activity behaved similarly to total phenols with a final retention between 38.5 and 59.5%. In the case of gastrointestinal digestion simulation, the two powders released 80% phenols during the gastric phase and a significant increase in release during the intestinal phase in which spray-dried and freeze-dried powders had 90.6% and 94.9% release during the last hour of the experiment. A similar behavior was detected for antioxidant activity in which spray-dried and freeze-dried powders featured 50% during the gastric phase and a significant increase during the intestinal one, with 69.4% and 67.8% of antioxidant activity respectively by the spray-dried and freeze-dried powders during the last hour of the experiment. However, monomer anthocyanins had a significant 50% reduction of contents in the intestinal phase in which the spray-dried and freeze-dried powders had 39% and 39.8% release respectively during the last hour of the experiment. The methods for obtaining microparticles failed to affect the stability of the powder in fast storage tests and in gastrointestinal simulation tests. Current research also evaluated the effect of UV-C irradiation on Bordô grapes for post-harvest treatment. Two harvests in two different years were analyzed: grapes of the first harvest underwent 0, 0.5, 1, 4, 10 and 30 minutes irradiation (120 W) and stored at 22 °C, whereas grapes of the second harvest underwent UV-C irradiations (120 W) during 0, 0.5 and 4 minutes, coupled to ultrasound (40 kHz) for 5 minutes and stored at 22 °C. The former did not have any increase in bioactive compounds and antioxidant activity. Results of the latter, however, demonstrated a significant increase in total phenols, total monomer anthocyanins and antioxidant activity for grapes irradiated during 0.5 minutes with UV-C and for those irradiated with UV-C for 4 minutes plus ultrasound for 5 minutes. There was 1.2 - 2.0 times increase when compared to control, with no change in color in the irradiated grapes.