Thesis (MscAgric (Viticulture and Oenology))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: Little scientific information regarding the effect of different geological parent materials on grapevine performance is currently available in South Africa. This aspect is of special significance for the Helderberg area, where parent material may change from granite to shale over a short distance. This results in shale- as well as granite-derived soils often occurring within the same vineyard. The objectives of this study were to (i) quantify the nutritional status and other soil properties of different parent materials (shale and granite) and overlying soils (ii) investigate the impact of geological differences in the soil on the vine nutritional status and certain vine parameters. This study was done over two seasons (2006/2007 and 2007/2008). Two Sauvignon blanc and two Cabernet Sauvignon vineyard blocks were selected at two different localities for each cultivar in the Helderberg area, South Africa. Shale- and granitederived soils were identified within each block. Kaolinite was the dominant mineral, whereas quartz and feldspar were sub-dominant. Traces of mica were also present in some shale- and granite-derived soils. Granite- contained significantly higher contents of coarse sand than shale-derived soils, whilst the opposite occurred in terms of fine sand. These differences affected the water holding capacity, in general making it higher in shale- than granite-derived soils. Shale-derived soils had higher concentrations of total K but granite-derived soils had a higher ability to release K as they contained higher concentrations of soluble K. The Q/I parameters, potential buffering capacity of K (PBCK) and equilibrium activity ratio of K (ARK) showed no consistent responses to geological differences. Potassium concentrations were higher in the leaf blades (obtained before harvest in 2007) from Sauvignon blanc grapevines on granite- than on shale-derived soils. Potassium concentrations in the Cabernet Sauvignon juice (obtained in 2007) tended higher in juice from granite- than from shale-derived soils. In 2008, K concentrations tended higher in juice from shale- than from granite-derived soils for both cultivars. The pH of the Sauvignon blanc juice (obtained in 2008) tended higher in juice from shale-than from granite-derived soils, thus corresponding with the K concentrations in the juice in this season. Nitrogen concentrations were higher in Cabernet Sauvignon juice (obtained in 2007) and Sauvignon blanc juice (obtained in 2008) from shalethan from granite-derived soils. In terms of vine water status, vines on granite-derived soils appeared more stressed than those on shale-derived soils in both seasons at one of the vineyards. In these Sauvignon blanc and Cabernet Sauvignon vineyards, the K nutritional status was not affected by geology in a consistent manner but there were some noticeable tendencies for a specific cultivar during certain seasons. On account of vines being planted on shale- and granite-derived soils within the same block, soil preparation was done similarly for both soils, and they were exposed to similar irrigation schedules, canopy management strategies and climatic conditions. Therefore, there is a high probability that all these practices may have negated the effect of geology on the K status of soils and especially on juice K concentration at the time of harvest. It was clear that seasonal differences and fertilisation affected the nutritional status of the vines more than geology. / AFRIKAANSE OPSOMMING: In Suid-Afrika is daar tans min wetenskaplike inligting oor die effek van verskillende geologiese moedermateriale op die prestasie van wingerd beskikbaar. Hierdie aspek is veral van belang in die Helderberg-area, waar moedermateriaal oor ‘n baie kort afstand van graniet na skalie kan wissel. Dit lei daartoe dat skalie-, sowel as granietgronde, dikwels binne dieselfde wingerd voorkom. Die doelwitte van die studie was om: (i) die voedingstatus en ander grondkundige eienskappe van die verskillende moedermateriale (skalie en graniet) en oorliggende gronde te kwantifiseer (ii) die impak van geologiese verskille in die grond op wingerd se voedingstatus en sekere wingerdkundige parameters, te ondersoek. Hierdie studie is oor twee seisoene (2006/2007 en 2007/2008) gedoen. Twee Sauvignon blanc en twee Cabernet Sauvignon wingerdblokke is geselekteer by twee verskillende lokaliteite vir elke kultivar in die Helderbergarea, Suid-Afrika. Beide skalie- en granietgrond is binne elke blok geïdentifiseer. Kaoliniet was die dominante mineraal, met kwarts en veldspaat sub-dominant, terwyl spore van mika ook in beide skalie- en granietgronde gevind is. Granietgronde het betekenisvol hoër hoeveelhede growwe sand bevat, terwyl skaliegronde meer fyn sand bevat het. Hierdie verskille het ‘n effek op waterhouvermoë gehad en daartoe gelei dat waterinhoude oor die algemeen hoër was vir skaliegronde. Skaliegronde het groter hoeveelhede totale K bevat, maar granietgronde se vermoë om K vry te stel was hoër, omdat hulle ‘n hoër konsentrasie oplosbare K bevat het. Die Q/I parameters, potensiële buffervermoë vir K (PBCK) en ewewig aktiwiteitsverhouding vir K (ARK), is nie op ‘n konsekwente wyse deur geologiese verskille beïnvloed nie. Vir die Sauvignon blanc wingerde was kalium konsentrasies in blaarskywe (gemonster voor oes in 2007) hoër vir graniet- as vir skaliegronde. Kalium konsentrasies in die sap vanaf Cabernet Sauvignon (gemonster in 2007) het hoër geneig vir granietgronde. In 2008 het die kalium konsentrasies, vir beide kultivars, hoër geneig in sap vanaf skaliegronde. Gedurende dié seisoen het die pH van sap ook hoër geneig vir Sauvignon blanc vanaf skaliegronde, wat dus ooreenstem met die K inhoud van die sap. Stikstof konsentrasies was hoër in sap vanaf skaliegronde vir Cabernet Sauvignon (2007) en vir Sauvignon blanc (2008). In terme van die wingerde se waterstatus, het stokke op die granietgrond, by een van die lokaliteite, geneig om gedurende beide seisoene onder groter stremming te wees op graniet as op skaliegrond. In hierdie Sauvignon blanc en Cabernet Sauvignon wingerde, is K voedingstatus nie op ‘n konsekwente wyse deur geologie geaffekteer nie, maar gedurende sommige seisoene was daar wel duidelike tendense vir ‘n spesifieke kultivar. Omdat die stokke binne dieselfde blok op skalie- en graniet gronde geplant is, was grondvoorbereiding eenders vir die twee grondtipes terwyl besproeiingskedule, lowerbestuur en klimaatstoestande ook identies was. Daar is dus ‘n hoë waarskynlikheid dat al hierdie faktore daartoe kon bygedra het dat die effek van geologie op die K status van van gronde versluier is, veral die effek op die K inhoud van sap teen oestyd. Dit was duidelik dat seisoenale klimaatsverskile en bemestingspraktyke ’n groter effek as geologie op die voedingstatus van die wingerd gehad het.

Thesis (MScAgric (Viticulture and Oenology))--University of Stellenbosch, 2007. / Biogenic amines are nitrogenous compounds of low molecular weight found in most fermented foods, including wine. These biologically produced amines are essential at low concentrations for normal metabolic and physiological functions in animals, plants and micro-organisms. However, biogenic amines can have adverse effects at high concentrations and pose a health risk for sensitive individuals. Symptoms include nausea, hot flushes, headaches, red rashes, respiratory distress and fluctuations in blood pressure. A number of countries have implemented upper limits for histamine in food and wine. This development has already started to threaten commercial export transactions and may become more serious in the near future, especially in the competitive wine industry of today. The most important biogenic amines in wine include histamine, tyramine, putrescine, cadaverine and phenylethylamine which are produced from the amino acids histidine, tyrosine, ornithine, lysine and phenylalanine respectively. Biogenic amines are mainly produced in wine by microbial decarboxylation of the corresponding precursor amino acid. It may be produced by yeast during alcoholic fermentation, by lactic acid bacteria during malolactic fermentation, or potentially by spoilage microbes such as acetic acid bacteria and Brettanomyces. However, lactic acid bacteria are widely accepted as the main causative agents. Inoculation with commercial malolactic fermentation starter cultures that do not possess the relevant decarboxylase genes may inhibit the growth and activity of decarboxylase positive indigenous bacteria and as such control the production of biogenic amines in wine. In this study it was shown that co-inoculation of malolactic starter cultures together with alcoholic fermentation could reduce the incidence of biogenic amines in wine compared to conventional inoculation protocols; presumably because undesirable activities were restrained at an earlier stage during co-inoculation. It was also indicated in this work that in some cases the effect of co-inoculation on biogenic amine reduction may only be visible after a period of ageing. The frequency of biogenic amine occurrence in wines aged for a short period was generally higher in the presence of fermentation lees than in its absence. This work also included a preliminary investigation into the contribution of commercial wine yeast starter cultures to biogenic amine production. Diamines and polyamines (putrescine, spermidine and cadaverine) were produced to variable extents by all yeasts with very little or no production of physiologically important biogenic amines (histamine, tyramine and phenylethylamine). Another objective of this study was to evaluate the influence of common winemaking practices on biogenic amine production under winemaking conditions. We have shown that biogenic amine production by lactic acid bacteria could be influenced, amongst others, by the presence of precursor amino acids in the grape must or wine, the time of contact between juice or wine and grape skins, the time of contact between wine and yeast lees, the presence of microbial nutrients, wine pH, sulphite and ethanol levels, the phenolic composition of the wine and the number of decarboxylase positive lactic acid bacteria present in the wine. Lately, the wine industry is under increasing pressure to increase measures to ensure food safety and security and to eliminate any compound, present even in trace amounts that could reduce the wholesomeness of the wine. The need arises for a rapid and inexpensive method for quality control. In this study we investigated the potential to use Fourier transform infrared spectroscopy to rapidly screen for the presence of elevated levels of biogenic amines. This presents a novel method for the detection and quantification of total biogenic amines in wines.

Thesis (MScAgric (Viticulture and Oenology. Wine Biotechnology))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT: Necrotrophic fungi infect many economically important crop plants. This results in great losses in the agricultural sector world-wide. Understanding the nature by which plants respond to pathogens is imperative for genetically enhancing disease resistance in plants. Research tools have significantly contributed to our understanding of how the plant responds to pathogen attack, identifying an array of defence mechanisms used by plants upon attack. Many fungal pathogens secrete endopolygalacturonases (endoPGs) when infecting plants. These hydrolytic enzymes are inhibited by polygalacturonase-inhibiting proteins (PGIPs) associated with plant cell walls. PGIPs are well characterised and their current known functions are all linked to endoPG inhibition and the subsequent upregulation of plant defence pathways. Work on grapevine PGIPs have shown that apart from being efficient antifungal proteins, leading to protection of the plant against Botrytis cinerea when overexpressed, PGIPs might also have additional functions linked to cell wall strengthening. This working hypothesis formed the motivation of this study where a cinnamyl alcohol dehydrogenase (CAD) (1.1.1.195) gene was targeted for functional analysis in tobacco (Nicotiana tabacum). Some previous work and genetic resources obtained is relevant to this study, specifically previously characterized transgenic tobacco lines overexpressing the Vitis vinifera pgip1 (Vvpgip1) gene. These lines have confirmed PGIP-specific resistance phenotypes against B. cinerea, as well as increased levels of CAD transcripts in healthy plants. Moreover, preliminary evaluations indicated increased lignin levels as well as differential expression of several other cell wall genes in these overexpressing lines (in the absence of infections). In this study we generated a transgenic tobacco population, overexpressing the native CAD14 gene, via Agrobacterium transformations. The transgene was overexpressed with the Cauliflower Mosaic Virus promoter (CaMV 35Sp). The CAD transgenic population was analyzed for transgene integration and expression and showed active transcription, even from leaves that normally don’t express CAD to high levels. These lines, together with the untransformed control, and a representative transgenic VvPGIP1 tobacco line previously characterized with elevated expression of CAD were used for all further analyses, specifically CAD activity assays of stems and leaves, as well as whole plant infections with B. cinerea. CAD enzyme activity assays were performed on healthy uninfected plant lines, without inducing native CAD expression or resistance phenotypes (i.e. without Botrytis infection). CAD activity was detected in leaves and stems, but a statistically sound separation between the CAD population and the untransformed control was only observed in the stems. The CAD assays also confirmed previous results that indicated that CAD transcription was upregulated in the PGIP line in the absence of infection. Overall, in all plant lines the stems exhibited 10-fold higher levels of CAD activity than the leaves, but the transgenic VvPGIP1 line showed a further 2-3-fold increase in CAD activity in the stems, when compared to the untransformed control and the majority of the CAD overexpressing lines. Disease assessment by whole plant infections with B. cinerea of the CAD transgenic plants revealed reduced disease susceptibility towards this pathogen. A reduction in disease susceptibility of 20 – 40% (based on lesion sizes) was observed for a homologous group of transgenic lines that was statistically clearly separated from the untransformed control plants following infection with Botrytis over an 11-day-period. The VvPGIP1 transgenic line displayed the strongest resistance phenotype, with reduction in susceptibility of 47%. The reduction in plant tissue maceration and lesion expansion was most pronounced in the VvPGIP1 line compared to the CAD transgenic plants, while the CAD transgenic plants showed more reduction than the untransformed control. In combination, the data confirms that CAD upregulation could lead to resistance phenotypes. Relating this data back to the previously observed upregulation of CAD in the VvPGIP1-overexpressing lines, the findings from this study corroborates that increased CAD activity contributes to the observed resistance phenotypes, possibility by strengthening the cell wall. In conclusion, this study yielded a characterized transgenic population overexpressing the CAD14 gene; this overexpression contributed to increased RNA transcription compared to the untransformed control plant, increased CAD activity (most notably in the stems) and a disease resistance phenotype against Botrytis. These findings corroborates the current working hypothesis in our group that PGIPs might have a role in preparing the plant cell for attack by contributing to specific cell wall changes. The exact mechanisms are still currently unknown and under investigation. The transgenic lines generated in this study will be invaluable in the subsequent analyses where these various phenotypes will be subjected to profiling and accurate cell wall analyses. / AFRIKAANSE OPSOMMING: Nekrotrofiese swamme infekteer en beskadig verskeie ekonomies belangrike gewasse. Dit lei wêreldwyd tot groot verliese vir die landbousektor. Dit is noodsaaklik om te verstaan hoe plante reageer teenoor patogene, sodat die siekteweerstand van plante verbeter kan word. Navorsingshulpbronne het beduidend bygedra tot die kennis van plantreaksies tydens patogeniese aanvalle, en het sodoende ‘n reeks van weerstandmeganismes, wat die plant inspan tydens ‘n aanval, geïdentifiseer. Verskeie patogeniese swamme skei endopoligalakturonases (endoPGs) af tydens plantinfeksie. Hierdie hidrolitiese ensieme word geïnhibeer deur poligalakturonase-inhiberende proteïene (PGIPs) wat met die plantselwand geassosieerd is. PGIPs is goed gekarakteriseerd en al hulle huidiglik bekende funksies is gekoppel aan endoPG inhibisie en die daaropvolgende opregulering van plant weerstandspaaie. Navorsing op wingerd PGIPs het gewys dat, afgesien van die feit dat PGIPs goeie antifungiese proteïene is wat lei tot beskerming van die plant teen Botrytis cinerea wanneer dit ooruitgedruk word, PGIPs ook moontlik addisionele funksies verrig wat verwant is aan selwandversterking. Hierdie werkshipotese vorm die motivering vir hierdie studie waarin ‘n sinnamiel alkohol dehidrogenase (SAD) (1.1.1.195) geen geteiken is vir funksionele analise in tabak (Nicotiana tabacum). Vorige navorsing en genetiese hulpbronne daardeur verkry is belangrik vir hierdie studie, spesifiek die gekarakteriseerde transgeniese tabaklyne wat die Vitis vinifera pgip1 (Vvpgip1) geen ooruitdruk. Hierdie lyne besit bevestigde PGIP-spesifieke weerstandsfenotipes teen B. cinerea, sowel as hoër vlakke van SAD transkripte in gesonde plante. Voorlopige analises het ook aangedui dat hierdie ooruitdrukkende lyne hoër lignien vlakke het, asook differensiële uitdrukking van verskeie ander selwandgene (in die afwesigheid van infeksie). In hierdie studie is ‘n transgeniese tabakpopulasie gegenereer wat die natuurlike tabak SAD14 geen ooruitdruk, deur middel van Agrobacterium transformasie. Die transgeen is ooruitgedruk deur die Blomkool Mosaïek Virus promoter (CaMV 35Sp). Die SAD transgeniese populasie is geanaliseer vir transgeen integrasie en uitdrukking en het aktiewe transkriptering getoon, selfs in blare waar daar normaalweg nie hoë vlakke van SAD uitgedruk word nie. Hierdie lyne, die ongetransformeerde wilde-tipe kontrole sowel as ’n verteenwoordigende transgeniese VvPGIP1 tabaklyn wat vooraf gekarakteriseerd was met hoë SAD uitdrukking, is gebruik vir alle verdere analises, spesifiek SAD aktiwiteitstoetse in stingels en blare, asook heelplantinfeksies met B. cinerea. Aktiwiteitsanalises van die SAD ensiem is gedoen op gesonde ongeinfekteerde plantlyne, sonder om natuurlike tabak SAD uitdrukking of weerstandsfenotipes te induseer (dus sonder Botrytis infeksie). SAD aktiwiteit is waargeneem in beide die blare en stingels, maar ‘n statisties betekenisvolle skeiding is slegs gevind tussen die SAD populasie en die ongetransformeerde kontrole in die stingels. Die SAD toetse het ook vorige resultate bevestig wat aangedui het dat SAD transkripsie opgereguleer word in die PGIP lyn in die afwesigheid van infeksie. Die stingels het oor die algemeen ‘n 10-voudige vermeerdering in SAD aktiwiteitsvlakke getoon in vergelyking met die blare, maar die transgeniese VvPGIP1 lyn het ‘n verdere 2-3-voudige verhoging in SAD aktiwiteit gehad in die stingels ,in vergelyking met die ongetransformeerde kontrole en die meerderheid van die SADooruitdrukkende lyne. Siekteweerstand ondersoeke deur middel van heelplantinfeksies met B. cinerea van die SAD transgeniese plante, het verminderde vatbaarheid aangedui ten opsigte van hierdie patogeen. ‘n Afname in siekte-vatbaarheid van 20 – 40% (gebaseer op wondgroottes) is waargeneem vir ‘n homoloë groep transgeniese lyne wat statisties betekenisvol geskei kon word van die ongetransformeerde kontrole plante na infeksie met Botrytis in ‘n infeksietoets wat 11 dae geduur het. Die VvPGIP1 transgeniese lyn het die mees weerstandbiedende fenotipe gehad, met ‘n afname in siekte-vatbaarheid van 47%. Die afname in plantweefselafbreking en wondgrootte was die duidelikste in die VvPGIP1 lyn in vergelyking met die SAD transgeniese plante, terwyl die SAD transgeniese plante ‘n groter afname aangedui het as die ongetransformeerde kontrole. In kombinasie het die data bevestig dat SAD opregulasie kan lei tot weerstandbiedende fenotipes. Hierdie data, in vergelyking met die vorige bevinding van opregulasie van SAD in die VvPGIP1-ooruitdrukkende lyne, bevestig dat hoër SAD aktiwiteit bydra tot die waargenome weerstandbiedende fenotipes, moontlik deur versterking van die plantselwand. Ter afsluiting, hierdie studie het ‘n gekarakteriseerde transgeniese populasie wat die SAD14 geen ooruitdruk gelewer; hierdie ooruitdrukking het bygedra tot hoër RNA transkripsie in vergelyking met die kontrole, verhoogde SAD aktiwiteit (veral in die stingels) en siekteweerstandbiedende fenotipes teenoor Botrytis. Hierdie bevindinge ondersteun die huidige werkshipotese in ons groep dat PGIPs moontlik ‘n rol speel in die voorbereiding van die plantsel teen infeksie deur spesifieke selwandveranderinge te veroorsaak. Die spesifieke meganismes is steeds onbekend en word verder ondersoek. Die transgeniese lyne wat tydens hierdie studie gegenereer is, sal baie belangrik wees in opvolgende analises waar hierdie verskillende fenotipes gebruik kan word om die profiel van selwandkomponente, maar ook die akkurate selwandsamestelling te bestudeer.