Institute for Wine Biotechnology

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3-way networks : application of hypergraphs for modelling increased complexity in comparative genomics
(PLoS, 2015-03) Weighill, Deborah A.; Jacobson, Daniel A.
We present and develop the theory of 3-way networks, a type of hypergraph in which each edge models relationships between triplets of objects as opposed to pairs of objects as done by standard network models. We explore approaches of how to prune these 3-way networks, illustrate their utility in comparative genomics and demonstrate how they find relationships which would be missed by standard 2-way network models using a phylogenomic dataset of 211 bacterial genomes.
Bioengineering beans for phosphate-deficient soils in southern Africa
(Academy of Science for South Africa, 2003) Viktor, A.; Cordero-Otero, R.; Valentine, A.
APPROXIMATELY EIGHT SPECIES OF SOUTHern African legumes are currently used as sustainable food crops. Biotechnology has the potential to improve the productivity of growing these plants by small-scale farmers who cannot afford sufficient phosphate fertilizer to optimize their nitrogen fixation and hence conversion to edible protein. The metabolic adaptations that enable legumes to fix atmospheric nitrogen are currently being investigated by our group for the purpose of genetic modification to enhance crop yields. Until now, attempts at modifying host plants or symbiotic bacteria have not significantly enhanced N2 fixation. We propose, instead, to bioengineer the key enzymes that control the mechanisms involved in protein formation. This may lead to enhanced seed protein content, which would be of advantage to poor communities that rely on this source of food. We postulate that misregulation of phosphoenolpyruvate carboxylase (PEPc) could be exploited by biotechnology to improve N2 fixation and protein content. We have found that, as distinct from their roots, legume nodules are under permanent phosphate stress, even during optimal phosphate supply to the host plant, implying that the development of phosphate stress may engage different forms of PEPc to ensure continued nodule functioning.
Biogenic amines in wine : understanding the headache
(SASEV, 2008-09) Smit, A. Y.; Du Toit, W. J.; Du Toit, M.
The presence of biogenic amines in wine is becoming increasingly important to consumers and producers alike, due to the potential threats of toxicity to humans and consequent trade implications. In the scientific field, biogenic amines have the potential to be applied as indicators of food spoilage and/or authenticity. Biogenic amines can be formed from their respective amino acid precursors by various microorganisms present in the wine, at any stage of production, ageing or storage. To understand the large number of factors that could influence the formation of biogenic amines, the chemical, biochemical, enzymatic and genetic properties relating to these compounds have to be considered. Analytical and molecular methods to detect biogenic amines in wine, as well as possibilities that could enable better control over their production levels in wine will also be explored in this review.
Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae
(PLoS, 2013) Franken, Jaco; Brandt, Bianca A.; Tai, Siew L.; Bauer, Florian
Levans are fructose polymers synthesized by a broad range of micro-organisms and a limited number of plant species as non-structural storage carbohydrates. In microbes, these polymers contribute to the formation of the extracellular polysaccharide (EPS) matrix and play a role in microbial biofilm formation. Levans belong to a larger group of commercially important polymers, referred to as fructans, which are used as a source of prebiotic fibre. For levan, specifically, this market remains untapped, since no viable production strategy has been established. Synthesis of levan is catalysed by a group of enzymes, referred to as levansucrases, using sucrose as substrate. Heterologous expression of levansucrases has been notoriously difficult to achieve in Saccharomyces cerevisiae. As a strategy, this study used an invertase (Δsuc2) null mutant and two separate, engineered, sucrose accumulating yeast strains as hosts for the expression of the levansucrase M1FT, previously cloned from Leuconostoc mesenteroides. Intracellular sucrose accumulation was achieved either by expression of a sucrose synthase (Susy) from potato or the spinach sucrose transporter (SUT). The data indicate that in both Δsuc2 and the sucrose accumulating strains, the M1FT was able to catalyse fructose polymerisation. In the absence of the predicted M1FT secretion signal, intracellular levan accumulation was significantly enhanced for both sucrose accumulation strains, when grown on minimal media. Interestingly, co-expression of M1FT and SUT resulted in hyper-production and extracellular build-up of levan when grown in rich medium containing sucrose. This study presents the first report of levan production in S. cerevisiae and opens potential avenues for the production of levan using this well established industrial microbe. Furthermore, the work provides interesting perspectives when considering the heterologous expression of sugar polymerizing enzymes in yeast.
Carnitine metabolism and biosynthesis in the yeast Saccharomyces cerevisiae
(Stellenbosch : University of Stellenbosch, 2009-12) Franken, Jaco; Bauer, Florian; Strauss, Erick; University of Stellenbosch. Faculty of Agrisciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: Carnitine plays an essential role in eukaryotic metabolism by mediating the shuttling of activated acyl residues between intracellular compartments. This function of carnitine, referred to as the carnitine shuttle, is supported by the activities of carnitine acyltransferases and carnitine/acylcarnitine transporters, and is reasonably well studied and understood. While this function remains the only metabolically well established role of carnitine, several studies have been reporting beneficial effects associated with dietary carnitine supplementation, and some of those beneficial impacts appear not to be directly linked to shuttle activity. This study makes use of the yeast Saccharomyces cerevisiae as a cellular model system in order to study the impact of carnitine and of the carnitine shuttle on cellular physiology, and also investigates the eukaryotic carnitine biosynthesis pathway. The carnitine shuttle of S. cerevisiae relies on the activity of three carnitine acetyltransferases (CATs), namely Cat2p (located in the peroxisome and mitochondria), Yat1p (on the outer mitochondrial membrane) and Yat2p (in the cytosol), which catalyze the reversible transfer of activated acetyl units between CoA and carnitine. The acetylcarnitine moieties can be transferred across the intracellular membranes of the peroxisomes and mitochondria by the activity of the carnitine/acetylcarnitine translocases. The activated acetyl groups can be transferred back to free CoA-SH and further metabolised. In addition to the carnitine shuttle, yeast can also utilize the glyoxylate cycle for further metabolisation of in particular peroxisomally generated acetyl-CoA. This cycle results in the net production of succinate from two molecules of acetyl-CoA. This dicarboxylic acid can then enter the mitochondria for further metabolism. Partial disruption of the glyoxylate cycle, by deletion of the citrate synthase 2 (CIT2) gene, generates a yeast strain that is completely dependent on the activity of the carnitine shuttle and, as a consequence, on carnitine supplementation for growth on fatty acids and other non-fermentable carbon sources. In this study, we show that all three CATs are required for the function of the carnitine shuttle. Furthermore, overexpression of any of the three enzymes is unable to crosscomplement deletion of any one of the remaining two, suggesting a highly specific role for each CAT in the function of the shuttle. In addition, a role for carnitine that is independent of the carnitine shuttle is described. The data show that carnitine can influence the cellular response to oxidative stresses. Interestingly, carnitine supplementation has a protective effect against certain ROS generating oxidants, but detrimentally impacts cellular survival when combined with thiol modifying agents. Although carnitine is shown to behave like an antioxidant within a cellular context, the molecule is unable to scavenge free radicals. The protective and detrimental impacts are dependent on the general regulators of the cells protection against oxidative stress such as Yap1p and Skn7p. Furthermore, from the results of a microarray based screen, a role for the cytochrome c heme lyase (Cyc3p) in both the protective and detrimental effects of carnitine is described. The requirement of cytochrome c is suggestive of an involvement in apoptotic processes, a hypothesis that is supported by the analysis of the impact of carnitine on genome wide transcription levels. A separate aim of this project involved the cloning and expression in S. cerevisiae of the four genes encoding the enzymes from the eukaryotic carnitine biosynthesis pathway. The cloned genes, expressed from the constitutive PGK1 promoter, were sequentially integrated into the yeast genome, thereby reconstituting the pathway. The results of a plate based screen for carnitine production indicate that the engineered laboratory strains of S. cerevisiae are able to convert trimethyllysine to L-carnitine. This work forms the basis for a larger study that aims to generate carnitine producing industrial yeast strains, which could be used in commercial applications.
Carnitine requires choline to exert physiological effects in saccharomyces cerevisiae
(Frontiers Media, 2018-07-02) Du Plessis, Michelle; Franken, Jaco; Bauer, Florian; De Biase, Daniela
L-Carnitine is a key metabolite in the energy metabolism of eukaryotic cells, functioning as a shuttling molecule for activated acyl-residues between cellular compartments. In higher eukaryotes this function is essential, and defects in carnitine metabolism has severe effects on fatty acid and carbon metabolism. Carnitine supplementation has been associated with an array of mostly beneficial impacts in higher eukaryotic cells, including stress protection and regulation of redox metabolism in diseased cells. Some of these phenotypes have no obvious link to the carnitine shuttle, and suggest that carnitine has as yet unknown shuttle-independent functions. The existence of shuttle-independent functions has also been suggested in Saccharomyces cerevisiae, including a beneficial effect during hydrogen peroxide stress and a detrimental impact when carnitine is co-supplemented with the reducing agent dithiothreitol (DTT). Here we used these two distinct yeast phenotypes to screen for potential genetic factors that suppress the shuttle independent physiological effects of carnitine. Two deletion strains, Δcho2 and Δopi3, coding for enzymes that catalyze the sequential conversion of phosphatidylethanolamine to phosphatidylcholine were identified for suppressing the phenotypic effects of carnitine. Additional characterisation indicated that the suppression cannot be explained by differences in phospholipid homeostasis. The phenotypes could be reinstated by addition of extracellular choline, but show that the requirement for choline is not based on some overlapping function or the structural similarities of the two molecules. This is the first study to suggest a molecular link between a specific metabolite and carnitine-dependent, but shuttle-independent phenotypes in eukaryotes.
Carotenoid cleavage dioxygenases (CCDs) of grape
(Stellenbosch : Stellenbosch University, 2012-12) Dockrall, Samantha; Young, Philip R.; Vivier, Melane A.; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
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
Cell differentiation in response to nutrient availability : the repressor of meiosis, RME1, positively regulates invasive growth in Saccharomyces cerevisiae
(Stellenbosch : Stellenbosch University, 2003-03) Hansson, Guy Robert, 1974-; Bauer, Florian; Pretorius, I. S.; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: Yeasts, like most organisms, have to survive in highly variable and hostile environments. Survival therefore requires adaptation to the changing external conditions. On the molecular level, specific adaptation to specific environmental conditions requires the yeast to be able: (i) to sense all relevant environmental parameters; (ii) to relay the perceived signals to the interior of the cell via signal transduction networks; and (iii) to implement a specific molecular response by modifying enzyme activities and by regulating transcription of the appropriate genes. The availability of nutrients is one of the major trophic factors for all unicellular organisms, including yeast. Saccharomyces cerevisiae senses the nutritional composition of the media and implements a specific developmental choice in response to the level of essential nutrients. In conditions in which ample nutrients are available, S. cerevisiae will divide mitotically and populate the growth environment. If the nutrients are exhausted, diploid S. cerevisiae cells can undergo meiosis, which produces four ascospores encased in an ascus. These ascospores are robust and provide the yeast with a means to survive adverse environmental conditions. The ascospores can lie dormant for extended periods of time until the onset of favourable growth conditions, upon which the spores will germinate, mate and give rise to a new yeast population. However, S. cerevisiae has a third developmental option, referred to as pseudohyphal and invasive growth. In growth conditions in which nutrients are limited, but not exhausted, the yeast can undergo a morphological switch, altering its budding pattern and forming chains of elongated cells that can penetrate the growth substrate to forage for nutrients. The focus of this study was on elements of the signal transduction networks regulating invasive growth in S. cerevisiae. Some components of the signal transduction pathways are well characterised, while several transcription factors that are regulated via these pathways remain poorly studied. In this study, the RMEt gene was identified for its ability to enhance starch degradation and invasive growth when present on a multiple copy plasmid. Rme1 p had previously been identified as a repressor of meiosis and, for this reason, the literature review focuses on the regulation of the meiotic process. In particular, the review focuses on the factors governing entry into meiosis in response to nutrient starvation and ploidy. Also, the transcriptional regulation of the master initiator of meiosis, IMEt, and the action of Ime1 p are included in the review. The experimental part of the study entailed a genetic analysis of the role of Rme1 p in invasive growth and starch metabolism. Epistasis analysis was conducted of Rme1 p and elements of the MAP Kinase module, as well as of the transcription factors, Mss11p, Msn1p/Mss10p, Tec1p, Phd1p and F108p. Rme1p is known to bind to the promoter of CLN2, a G1-cyclin, and enhances its expression. Therefore, the cell cyclins CLN1 and CLN2 were included in the study. The study revealed that Rme1 p functions independently or downstream of the MAP Kinase cascade and does not require Cln1 p or Cln2p to induce invasive growth. FL011/MUC1 encodes a cell wall protein that is required for invasive growth. Like the above-mentioned factors, Rme1 p requires FL011 to induce invasive growth. We identified an Rme1 p binding site in the promoter of FL011. Overexpression of Rme1p was able to induce FL01t expression, despite deletions of mss11, msn1, ttos, tee1 and phd1. In the inverse experiment, these factors were able to induce FL011 expression in an rme1 deleted strain. This would indicate that Rme1 p does not function in a hierarchical signalling system with these factors, but could function in a more general role to modify transcription.
Characterisation of grapevine berry samples with infrared spectroscopy methods and multivariate data analyses tools
(Stellenbosch : Stellenbosch University, 2015-04) Musingarabwi, Davirai M.; Vivier, Melane A.; Nieuwoudt, Helene; Stellenbosch University. Faculty of Agrisciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: Grape quality is linked to the organoleptic properties of grapes, raisins and wine. Many advances have been made in understanding the grape components that are important in the quality of wines and other grape products. A better understanding of the compositional content of grapes entails knowing when and how the various components accumulate in the berry. Therefore, an appreciation of grape berry development is vitally important towards the understanding of how vineyard practices can be used to improve the quality of grapes and eventually, wines. The more established methods for grape berry quality assessment are based on gravimetric methods such as colorimetry, fluorescence and chromatography. These conventional methods are accurate at targeting particular components, but are typically multi-step, destructive, expensive, polluting procedures that might be technically challenging. Very often grape berries are evaluated for quality (only) at harvest. This remains a necessary exercise as it helps viticulturists and oenologists to estimate some targeted metabolite profiles that are known to greatly influence chemical and sensory profiles of wines. However, a more objective measurement of predicting grape berry quality would involve evaluation of the grapes throughout the entire development and maturation cycle right from the early fruit to the ripe fruit. To achieve this objective, the modern grape and wine industry needs rapid, reliable, simpler and cost effective methods to profile berry development. By the turn of the last millennium, developments in infrared instrumentation such as Fourier-transform infrared (FT NIR) and attenuated total reflectance Fourier-transform infrared spectroscopy (ATR FT-IR) in combination with chemometrics resulted in the development of rapid methods for evaluating the internal and external characteristics of fresh fruit, including grapes. The advancement and application of these rapid techniques to fingerprint grape compositional traits would be useful in monitoring grape berry quality. In this project an evaluation of grape berry development was investigated in a South African vineyard setting. To achieve this goal, Sauvignon blanc grape berry samples were collected and characterised at five defined stages of development: green, pre-véraison, véraison, post-véraison and ripe. Metabolically inactivated (frozen in liquid nitrogen and stored at -80oC) and fresh berries were analysed with FT-IR spectroscopy in the near infrared (NIR) and mid-infrared (MIR) ranges to provide spectral data. The spectral data were used to provide qualitative (developmental stage) and quantitative (metabolite concentration of key primary metabolites) information of the berries. High performance liquid chromatography (HPLC) was used to separate and quantify glucose, fructose, tartaric acid, malic acid and succinic acid which provided the reference data needed for quantitative analysis of the spectra. Unsupervised and supervised multivariate analyses were sequentially performed on various data blocks obtained by spectroscopy to construct qualitative and quantitative models that were used to characterise the berries. Successful treatment of data by principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) gave statistically significant chemometric models that discriminated the berries according to their stages of development. The loadings from MIR models highlighted the important discriminant variables responsible for the observed developmental stage classification. The best calibration models to predict metabolite concentrations were obtained from MIR spectra for glucose, fructose, tartaric acid and malic acid. The results showed that both NIR and MIR spectra in combination with multivariate analysis could be reliably used to evaluate Sauvignon blanc grape berry quality throughout the fruit’s development cycle. Moreover, the methods used were fast and required minimal sample processing and no metabolite extractions with organic solvent. In addition, the individual major sugar and organic acids were accurately predicted at the five stages under investigation. This study provides further proof that IR technologies are robust and suitable to explore high-throughput and in-field application of grape compound profiling.
Characterisation of non-saccharomyces yeasts using different methodologies and evaluation of their compatibility with malolactic fermentation
(South African Society for Enology and Viticulture, 2017) Du Plessis, H. W.; Du Toit, M.; Hoff, J. W.; Hart, R. S.; Ndimba, B. K.; Jolly, N. P.
Although Saccharomyces cerevisiae is the yeast species predominantly used for alcoholic fermentation, non-Saccharomyces yeast species are also important because they produce secondary metabolites that can contribute to the final flavour and taste of wines. In this study, 37 strains representing seven non-Saccharomyces species were characterised and evaluated for potential use in wine production, as well as for their effects on malolactic fermentation (MLF). Contour-clamped homogeneous electric field (CHEF) gel electrophoresis and matrix-assisted laser desorption ionisation using a time-of-flight mass spectrometer (MALDI-TOF MS) were used to verify species identity and to determine intra-species variation. Extracellular enzyme production, malic acid degradation and the fermentation kinetics of the yeasts were also investigated. CHEF karyotyping and MALDI-TOF MS were useful for identifying and typing Hanseniaspora uvarum, Lachancea thermotolerans, Candida zemplinina (synonym: Starmerella bacillaris) and Torulaspora delbrueckii strains. Only H. uvarum and Metschnikowia pulcherrima strains were found to have β-glucosidase activity. M. pulcherrima strains also had protease activity. Most of the strains showed limited malic acid degradation, and only Schizosaccharomyces pombe and the C. zemplinina strains showed mentionable degradation. In synthetic wine fermentations, C. stellata, C. zemplinina, H. uvarum, M. pulcherrima and Sc. pombe strains were shown to be slow to medium fermenters, whereas L. thermotolerans and T. delbrueckii strains were found to be medium to strong fermenters. The effect of the yeasts on MLF varied, but inhibition was strain dependent.
Characterisation, evaluation and use of non-Saccharomyces yeast strains isolated from vineyards and must
(Stellenbosch : Stellenbosch University, 2004-03) Jolly, N. P. (Neil Paul); Pretorius, I. S.; Augustyn, O. P. H.; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: Wine is the product of a complex biological and biochemical interaction between grapes and different microorganisms (fungi, yeasts, lactic acid bacteria and acetic acid bacteria, as well as the mycoviruses and bacteriophages affecting them) in which yeasts play the most important role regarding the alcoholic (primary) fermentation. These wine-associated yeasts can be divided into Saccharomyces and non-Saccharomyces yeasts. During fermentation, there is a sequence of dominance by the various non-Saccharomyces yeasts, followed by Saccharomyces cerevisiae, which then completes the fermentation. This is especially evident in spontaneously fermenting must, which has a low initial S. cerevisiae concentration. Some non- Saccharomyces yeasts can also be found throughout the fermentation. The non- Saccharomyces presence in the fermentation can affect wine quality, either positively or negatively. A positive contribution could be especially useful to improve wines produced from grape varieties with a neutral flavour profile due to non-optimal climatic conditions and/or soil types. As part of a comprehensive South African research programme, the specific objectives of this study were: the isolation of indigenous non-Saccharomyces yeasts from vineyards and musts; the identification of these isolates; the characterisation and evaluation of predominant species under winemaking conditions; and the development of a protocol for their use in enhancing wine quality. Initially, 720 isolates representing 24 different species, were isolated from grape (vineyard) and must samples taken over three vintages from four distinctly different wine producing regions. The isolates were characterised and grouped utilising biochemical profiles and DNA karyotyping, whereupon representative isolates were identified. The yeast species that had the highest incidence of predominance in the vineyard was Kloeckera apiculafa. However, some vineyard samples were characterised by low numbers or absence of this yeast, which is not according to generally accepted norms. Other species that also predominated in a few of the vineyard samples were Candida pulcherrima, Kluyveromyces thermofolerans, Rhodotorula sp. and Zygosaccharomyces bailii. Generally, there was a greater diversity of yeasts in the processed must than from the vineyard samples. Furthermore, while each sample showed a different yeast population, no pattern linking species to climatic zone was observed. Four species i.e. Candida collieulosa, Candida pulcherrima, Candida stel/ata and Kloeckera apiculata, were found to predominate in grape must samples. Representative strains consequently received further attention during laboratory and small-scale winemaking trials. A protocol was developed whereby individual species could be used in co-inoculated fermentations with S. cerevisiae in the small-scale production of wine. An improvement in wine quality was achieved and it was found that there was a link between specific species and grape cultivar. The ability of C. pulcherrima to improve Chenin blanc wine quality was investigated further. Results over three vintages showed that the wine produced by the co-inoculated fermentation was superior to that of a reference wine (produced by S. cerevisiae only). The improvement in wine quality was not linked to increased ester content nor were the standard chemical analyses adversely affected. The effects of pH and wine production parameters i.e. 802, fermentation temperature and use of di-ammonium phosphate (DAP), on this yeast followed the same pattern as that known for S. cerevisiae. This study was successfully completed and the developed protocol can be used for the improvement of Chenin blanc wine where additional aroma and quality is needed.
Characterization of Chenin Blanc wines produced by natural fermentation and skin contact : focus on application of rapid sensory profiling methods
(Stellenbosch : Stellenbosch University, 2014-12) Weightman, Jayne; Nieuwoudt, Helene; Setati, Mathabatha Evodia ; Stellenbosch University. Faculty of AgriSciences. Dept. of Institute for Wine Biotechnology.
ENGLISH ABSTRACT: Producers of South African (SA) dry and semi-dry Chenin blanc table wines are currently experimenting with winemaking techniques to modulate the flavours of the predominantly fruity styles of this genre. An important stage during wine style development is sensory profiling paired with consumer acceptance testing, before wine is produced on industrial scale. With those aforementioned goals in mind, this study was conducted in partnership with two commercial SA wine cellars. The main focus of the study was an investigation into the treatment effects of two winemaking techniques, respectively grape skin contact and natural fermentation, on the sensory profiles of experimentally produced Chenin blanc wines. Results obtained with descriptive sensory analysis (DA) of the wines were compared to those obtained by two rapid sensory profiling methods, namely projective mapping (PM) and frequency of attribute citation (FC). A consumer preference study was also done on the wines. In order to understand the treatment effects better, the dominant non-Saccharomyces yeasts that were present during the natural fermentations were identified using polymerase chain reaction (PCR), while the major volatile chemical compounds were identified with chromatography and mass spectrometry. The sensory and chemical profiles of the naturally fermented wines were significantly different from those of the inoculated wines. PCR analysis identified some of the yeasts present during alcoholic fermentation. In comparison to the inoculated fermented wines, the naturally fermented wines were generally perceived to have more intense and riper tropical fruity aromas, with enhanced sweetness and reduced intensities of sourness, bitterness and astringency. The wines fermented on the skins (FoS) had lower ester and volatile thiol concentrations than the control wines (with no skin contact) and limited skin contact wines. Sensory attributes linked to the FoS wines included aromas of dried fruit, dried grass and a vegetative character, with an increased sour and bitter taste and astringent mouthfeel. In contrast, the wines that were produced with limited skin contact (12 hours) retained their tropical fruity aromas better than the FoS wines. Limited skin contact seemed to have had a less harsh effect on the taste and mouthfeel than the FoS wines. A consumer study was done to establish a Generation Y consumer group’s (18-35 years) preference for the different treated wines. Overall, the naturally fermented wines, which were described as having a strong tropical fruit character, were preferred. The FoS wines were generally disliked by the consumers. Results obtained from the three sensory analysis methods, respectively DA, PM and FC, were similar, thereby confirming the suitability of the rapid methods PM and FC, to extract qualitative information from the sensory profiling of white wine. The results of this study made a significant contribution towards validation of rapid sensory methods for wine evaluation, which are particularly valuable in the context of sustainability and technology transfer to research and industry alike. The knowledge gained on the chemical profiles of SA Chenin blanc is novel and this is one of the first reports on the volatile thiol content of SA Chenin blanc wine.
Characterization of the Viable but Nonculturable (VBNC) State in Saccharomyces cerevisiae
(Public Library of Science, 2013-10-29) Salma, Mohammad; Rousseaux, Sandrine; Sequeira-Le Grand, Anabelle; Divol, Benoit; Alexandre, Herve
The Viable But Non Culturable (VBNC) state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to “resuscitate”. The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO2-induced VBNC state in S. cerevisiae and that the stress removal allows the “resuscitation” of VBNC cells during the VBNC state.
Coevolution of Saccharomyces cerevisiae and Lactobacillus plantarum : engineering interspecies cooperation
(Stellenbosch : Stellenbosch University, 2018-03) Du Toit, Sandra Christine; Bauer, Florian; Rossouw, Debra; Du Toit, Maret; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: Microbial interactions are ubiquitous in nature and play a vital role in economically important industrial processes like winemaking. Saccharomyces cerevisiae and Lactobacillus plantarum are important species responsible for the completion of alcoholic and malolactic fermentation (AF and MLF) respectively. Understanding how these species interact with each other and their environment is important to better manage successful completion of AF and MLF. However, the complexity of the wine matrix makes it nearly impossible to study these interactions in a natural environment and synthetic ecological systems can therefore be used to overcome these difficulties. This study was designed to establish a co-dependent, mutualistic relationship between S. cerevisiae and Lb. plantarum in order to gain insights into the cooperation between species, how pH, temperature, and inoculation dosages influences the interaction, and how the interaction evolves over time. The interaction, centered on the reciprocal exchange of amino acids, was established between the lysine auxotrophic strain S. cerevisiae THI4 and the isoleucine, alanine, valine, and methionine auxotrophic strain Lb. plantarum B038. Different combinations of amino acids were omitted from the chemically defined synthetic grape juice-like media in order to find an amino acid treatment which promoted the best growth for both microorganisms. B038 showed excellent growth when cocultured with THI4 for all the amino acid treatments, but THI4 struggled to grow under these conditions. The two treatments selected for further experiments were the Lys-Ile (lysine and isoleucine omitted) and Lys-Val (lysine and valine omitted) treatments since THI4 showed the best growth under these conditions. Lower temperature and pH conditions had a negative effect on the growth and malic acid consumption of B038, but when co-cultured with THI4 the yeast appeared to stimulate the growth of the bacteria under both selective and control conditions. THI4 continued to show poor growth performance and sugar consumption under these conditions. However, when THI4 and B038 were inoculated at cell densities with similar biomass, the growth of THI4 improved significantly. It was expected that THI4 and B038 would show poor growth when grown in the absence of their respective auxotrophic amino acids and support of their respective partner. This proved true for all the amino acid treatments except when B038 was grown in the absence of lysine and valine. B038’s ability to grow under these conditions was hypothesized to be linked to the uptake of glutamine and the production of γ-Aminobutyric acid (GABA), but further research is still required to investigate this. Over continuous rounds of fermentation, THI4 adapted to the imposed selective conditions by increasing its consumption of glucose while cell density remained the same. Whether this is linked to increased ethanol production still needs to be determined. No significant changes were observed in B038 after coevolving the strains. This study provides relevant insights into the industrially important interaction between S. cerevisiae and Lb. plantarum and also provides a basis for future work to create optimised yeast-bacteria pairings for both industrial applications in winemaking and to investigate the genetic changes involved in the establishment of cooperative interactions between species.
Construction of recombinant Saccharomyces cerevisiae strains for starch utilisation
(Stellenbosch : Stellenbosch University, 2002-12) Eksteen, Jeremy Michael; Pretorius, I. S.; Van Rensburg, P.; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology .
ENGLISH ABSTRACT: Starch-containing agricultural crops are widely available as feedstocks for the production of fuel ethanol, potable spirits or beer, single-cell protein (animal feed) and high-fructose corn syrups (sweeteners). Starch-rich crops, such as maize, rye, barley and wheat, are usually used for the production of whisky. One of the first steps in the production of whisky is to boil the raw starch at temperatures exceeding 100°C. This gelatinisation step is performed to disrupt and solubilise the starch granules to make them more accessible for enzymatic hydrolysis. After this cooking process, the starch is liquefied by a-amylase and then saccharified by glucoamylase and a debranching enzyme. Lipomyces kononenkoae and Saccharomycopsis fibuligera secrete highly effective a-amylases and glucoamylases, making them two of the most efficient raw-starchdegrading yeasts known. However, L. kononenkoae and S. fibuligera cannot be used in existing industrial fermentations because of their low ethanol tolerance, slow growth rate, catabolite repression, poorly characterised genetics and lack of GRAS (Generally Regarded As Safe) status. This study is divided into two sections. The aim of the first section was to clone a gene (LKA2) encoding a novel starch-degrading enzyme, a second a-amylase (Lka2p) from L. kononenkoae. LKA2 was cloned into a multicopy plasmid, the yeast episomal plasmid, YEp352, under the control of the phosphoglycerate kinase promoter (PGK1 p) and terminator (PGKh) expression cassette. This recombinant plasmid was designated pJUL3 and transformed into a laboratory strain of S. cerevisiae, I1278b. Plate and liquid assays revealed that the recombinant yeast secreted active a-amylase into the medium. The optimum pH for Lka2p was pH 3.5 and the optimum temperature 60°C. The aim of the second part of the study was to construct recombinant strains of S. cerevisiae secreting a-amylase and/or glucoamylase. The individual genes were cloned into a yeast-integrating plasmid, Ylp5, under the control of the PGK1p-PGK1.,-expression cassette. Two indigenous yeasts were selected on the basis of their ability to utilise raw starch, L. kononenkoae and S. fibuligera, as gene donors. Eight constructs containing the L. kononenkoae a-amylase genes, LKA 1 and LKA2, and the S. fibuligera a-amylase (SFA 1) and glucoamylase (SFG1) genes were prepared: four single-cassette plasmids expressing the individual coding sequences under the control of the PGK1 p-PGK1.,- expression cassette, resulting in plPLKA 1, pIPLKA2, plPSFA 1 and pIPSFG1, respectively; two double-cassette plasm ids (expressing both LKA 1 and LKA2 under the control of the PGK1p-PGK1 .,-expression cassette, and SFA 1 and SFG1 under their respective native promoters and terminators), resulting in pIPLKA1/2 and pIPSFAG, respectively, and two single-cassette plasmids expressing SFA 1 and SFG1 with their native promoters and terminators, resulting in pSFA 1 and pSFG1, respectively. The respective constructs were transformed into a laboratory strain of S. cere visiae , L1278b. By homologous recombination, each plasmid was integrated into the yeast genome at the ura3 locus. S. cerevisiae L:1278b that had been transformed with plPLKA 1/2, LKA 1 and LKA2 under the control of the PGK1 rrPGK1,expression cassette resulted in the highest levels of a-amylase activity when assayed for amylolytic activity in a liquid medium. This recombinant strain resulted in the most efficient starch utilisation in batch fermentations, consuming 80% of starch and producing 6 gIL of ethanol after 156 hours of fermentation. The strain expressing SFG1 under the control of the PGK1rrPGK1,expression cassette gave the highest levels of glucoamylase activity.' These results confirmed that co-expression of a-amylase and/or glucoamylase synergistically enhance starch degradation. This study paves the way for the development of efficient starch-degrading strains of S. cerevisiae for the production of whisky, beer and biofuel ethanol.
Consumer sensory perception of South African Chenin blanc wine
(Stellenbosch : Stellenbosch University, 2018-03) Mapheleba, Andiswa; Nieuwoudt, Helene; Muller, Nina; Pentz, Chris; Oertle, Ivan; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: In the food and beverage industries, the understanding of consumer behaviour and the sensory characteristics of products are important to producers as they aid in the implementation of communication and marketing strategies. South African Chenin blanc is a white wine variety that is characterised by diverse sensory profiles, an attribute that is considered as a strength of the variety from a technical wine production perspective. Accordingly, the wine industry has become highly interested in gaining knowledge about the perceptions of consumers regarding Chenin blanc styles in particular. The main aim of this study was to investigate the sensory and overall perceptions of Chenin blanc wine among South African consumers. Sensory analysis was conducted on three different Chenin style wines, namely fresh, complex unwooded and complex wooded wines using descriptive analysis (DA) with a trained panel. Among consumers, free listing and CATA were carried out to explore their sensory perceptions through encouraging them to freely describe their tasting experience and the selection of the perceived of aroma attributes obtained from DA. Thereafter, an exploratory study with the aid of a survey questionnaire was conducted to obtain insights of the consumers’ consumption, purchase behaviour and general perceptions about Chenin blanc wine.. Industry professionals from different disciplines of the wine industry, participated in the study by providing their viewpoints on aspects regarding the position of Chenin blanc in the SA wine industry through a self-completed questionnaire. The sensory attributes that were mostly provided by the consumers in free listing were fruit, citrus, tropical, honey, wood, earthy, nutty and fresh. A significantly lower number of attributes was perceived by consumers with CATA. The important non-sensory attributes listed by the consumers included their emotions, context of usage (occasion) and pleasure. They generally displayed an inclination towards buying wines with simple sensory label descriptors. According to the industry professionals, wine labelling and initiatives by the SA wine industry should be simple and relatable. Furthermore, Chenin blanc would highly benefit from cultivar-specific shows, due to its wide wine style spectrum. This study contributed towards a better understanding of the perceptions of SA wine consumers about Chenin blanc wine of the two respondent groups to aid in the understanding of the market and the wine industry, as well as their prospective roles in the future of Chenin blanc in SA.
Deconstructing wine grape cell walls with enzymes during winemaking : new insights from glycan microarray technology
(MDPI, 2019-01-04) Gao, Yu; Zietsman, Anscha J. J.; Vivier, Melane A.; Moore, John P.
Enzyme-aid maceration is carried out in most modern winemaking industries with a range of positive impacts on wine production. However, inconsistencies in enzyme efficiency are an issue complicated by unclear targets (limited information available on berry cell wall architecture of different cultivars) and the complex wine environment (i.e., fermenting must). Recent studies have been performed to develop a clearer picture of grape cell wall structures, maceration effects, and interactions between important wine compounds and grape-derived polysaccharides. This review highlights critically important recent studies on grape berry cell wall changes during ripening, the importance of enzymes during maceration (skin contact phase) and deconstruction processes that occur during alcoholic fermentation. The novelty of the Comprehensive Microarray Polymer Profiling (CoMPP) technique using cell wall probes (e.g., antibodies) as a method for following cell wall derived polymers during different biological and biotechnological processes is discussed. Recent studies, using CoMPP together with classical analytical methods, confirmed the developmental pattern of berry cell wall changes (at the polymer level) during grape ripening. This innovative technique were also used to track enzyme-assisted depectination of grape skins during wine fermentation and determine how this influence the release of wine favourable compounds. Furthermore, polysaccharides (e.g., arabinogalactan proteins) present in the final wine could be identified. Overall, CoMPP provides a much more enriched series of datasets compared to traditional approaches. Novel insights and future studies investigating grape cell wall and polyphenol interactions, and the tailoring of enzyme cocktails for consistent, effective and “customized” winemaking is advanced and discussed.
Defining the chemical features of wine perception
(Stellenbosch : Stellenbosch University, 2018-03) Fairbairn, Samantha; Bauer, Florian; Da Silva Ferreira, A. C.; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology.
ENGLISH ABSTRACT: All wines evoke a product recognition, regardless of quality and cultivar, but what is the origin of this feature? The prevalence of this wine concept suggests that its formation occurs independent of the varietal, and ageing-related aromas, and is therefore potentially a function of yeast metabolism. Yeast utilise the nutrients present in grape must to produce biomass, and metabolites which ultimately signify the conversion of grape juice to wine. Consequently, the nutrient composition is highly influential on the aromatic outcomes of alcoholic fermentation. Synthetic grape must is widely used to evaluate all facets of the fermentation process but there remains much to learn. In this study, the impact of two nutrients, namely, amino acids and anaerobic factors, were evaluated with regard to their impact on yeast growth and aroma production under fermentative conditions. This work also examines the extent to which yeast de novo metabolism, both primary and secondary metabolism, contributes to the formation of the wine-like feature. In a single amino acid context, a linear relationship was apparent between the amino acid concentration and the production of their associated volatile products. This relationship was evaluated in more complex amino acid mixtures and as expected, this linear relationship was lost. Nonetheless, a significant degree of responsiveness between the amino acid and its catabolites remained. The impact of sterol (plant or yeast derived) or unsaturated fatty acid treatments, individually, as well as in combinations, were compared for their contributions to biomass formation and aroma production. Sterols had a greater impact on biomass development, as the fermentations treated with only unsaturated fatty acids displayed a poorer response. Moreover, they differently impacted aroma production. The unsaturated fatty acid lowered the production of acetate esters, medium chain fatty acids and their esters, whereas sterol supplementation generally bolstered the production of all compounds measured. This work highlights the importance of anaerobic factor management during winemaking. Although these nutrients certainly impact wine aroma, this study also sought to examine the degree to which these nutrients contribute to wine (product) recognition. Using a novel fermentation-based approach, Saccharomyces cerevisiae converted a synthetic grape must into a wine-like product. These synthetic products underwent sensory evaluations to rate the product’s resemblance to wine as well as to describe the aroma. This sensory data was used as a decision-making tool to decide upon treatments to be studied in subsequent fermentations. Ultimately, a wine-like character was created by altering the anaerobic factor composition of a synthetic grape must. The use of this synthetic grape must would allow for the more meaningful sensory characterisation of these synthetic products, in addition to providing a wine-like matrix used to evaluate the sensory implications of wine odorants.
The deletion and overexpression of two esterase genes, IAH1 and TIP1, in Saccharomyces cerevisiae to determine their effects on the aroma and flavour of wine and brandy
(Stellenbosch : Stellenbosch University, 2002-12) Hignett, Jason Satch; Du Toit, M.; Pretorius, I. S.; Lambrechts, M. G.; Stellenbosch University. Faculty of AgriSciences. Dept. of Viticulture and Oenology. Institute for Wine Biotechnology .
ENGLISH ABSTRACT: No single chemical constituent can be accredited with giving wine and brandy their overall aroma and flavour. The aroma and flavour of wine and brandy are rather attributed to a number of chemical constituents reacting together and it is these reactions that give the beverage its character. Certain chemicals within wine and brandy do, however, make larger contributions to the flavour. These include the esters, terpenes and volatile acids, although others also exist. Esters are a large group of volatile compounds with variable aroma and flavour characteristics, including banana-like (isoamyl acetate), apple-like (ethyl caproate) and chemical/solvent-like (ethyl acetate). Esters are produced as secondary metabolites during the conversion of sugar to ethanol and are formed when an alcohol binds with a fatty acid. Chemically, ester metabolism is well documented and understood; however, much work still needs to be done on a genetic level. The yeast strain used during fermentation is one of the most important factors contributing to the type and quantity of esters produced. This is due to differences in genetic makeup. The metabolism of esters is controlled largely on a genetic level, with numerous genes being involved. The alcohol acetyltransferase genes are involved in ester anabolism, whilst esterase genes are involved in ester catabolism. Esterases have a negative effect on the overall level of esters within an alcoholic beverage, as they are capable of reducing the number of esters and are thus capable of altering the beverage's aroma and flavour profile. The IAH1 and the TIP1 gene products are believed to encode for two such esterases. The objective of this study was to investigate the contribution of the IAH1 and TIP1 genes to the level of esters in both wine and brandy. This was accomplished by using two approaches. Firstly, the above genes were disrupted using a polymerise chain reaction (PCR)-generated disruption cassette homologous to either the IAH1 or the TIP1 gene. These cassettes were integrated into the industrial wine yeast, Saccharomyces cerevisiae strain VIN13. The integrations were verified by Southern blot analysis to produce yeasts VIN13-~IAH1 and VIN13-~TIP1; however, only a single copy of each was disrupted. Secondly, the IAH1 and the TIP1 genes were cloned from S. cerevisiae using PCR into plasmid pj between the phosphoglycerate kinase gene (PGK1) promoter and terminator, producing plasmids pJ-IOE1 and pJ-TOE1. The PGK1 promoter has previously been shown to constitutively express genes at high levels. These new constructs were then used as template for PCR to produce two overexpression cassettes, one for IAH1 and the other for TlP1. These cassettes were integrated into S. cerevisiae VIN13 and verified by Southern blot analysis to produce strains VIN13-IOE1 and VIN13-TOE1. The above yeast strains including VIN13 were used for the production of wines and base wines from Colombard must. Reverse-transcriptase (RT-PCR) confirmed that the VIN13-IOE1 and VIN13-TOE1 strains overexpressed the appropriate gene at a higher level than the control VIN13 strain. The VIN13-AIAH1 disrupted strain showed no difference in expression level to that of the control strain, whilst VIN13-ATIP1 showed lower levels of expression than that of the control strain. VIN13-IOE1 behaved as expected, with a decrease of between 30% and 60% in the total ester level in the wine and base wine respectively, a 30% decrease in the total acid level and no change in the higher alcohol level. The VIN13-AIAH1 strain showed no difference to the control wine, most likely as this strain still expressed the IAH1 gene at levels consistent with the control strain. VIN13-TOE1 behaved in an unexpected manner - instead of hydrolysing esters, it appeared to produce them. This increase in the total ester level was most noticeable during distillation, when a 20% increase took place. Another unexpected occurrence was a large decline in the total acid level, with acetic acid being the most significant contributor, decreasing by up to 78%. This is a very favourable finding, as acetic acid is a known spoilage molecule and is a cause of sluggish/stuck fermentations. VIN13-ATIP1 behaved in an opposite manner to VIN13-TOE1, with higher total acid levels and slightly decreased total ester levels, especially during distillation. Neither affected the total higher alcohol levels. Sensorially, the only significant difference in the wine samples was for the fruity flavour. A panel of judges distinguished that VIN13-TOE1 was fruitier than the other wines, with VIN13-ATIP1 being the least fruity. This study again proves the significant impact that a single gene can have on the chemical makeup of wine and brandy. The relatively simple genetic alteration of an organism can drastically change and improve not only the organoleptic properties of the organism, but its viability as well. These alterations can produce more favourable organisms with more desirable characteristics for the fermenting beverage industry to produce products of higher quality and better suitability.
Determining the impact of industrial wine yeast strains on organic acid production under white and red wine-like fermentation conditions
(South African Society for Enology and Viticulture, 2015) Chidi, B. S.; Rossouw, D.; Buica, A. S.; Bauer, Florian
Organic acids are a major contributor to wine flavour and aroma. In the past, the scientific focus has mostly been on organic acids derived from grapes or on the transformation of malic acid to lactic acid by lactic acid bacteria, since these acids contribute significantly to the final total acidity of wine. However, the organic acid concentration and composition also change significantly during alcoholic fermentation, yet only limited information regarding the impact of different yeast strains on these changes has been published. Here we report on changes in organic acid (malic, tartaric, citric, succinic, acetic and pyruvic) composition during fermentation by five widely used industrial wine yeast strains in a synthetic grape must (MS300) reflecting two very different, but both wine-like, fermentation conditions. Samples were obtained from three physiological stages during fermentation, namely the exponential growth phase (day 2), early stationary phase (day 5) and late stationary phase (day 14). These different stages were selected to provide more information on acid evolution throughout fermentation, as well as on the impact of nutritional and environmental conditions during aerobic and anaerobic fermentation. Among other observations, some strains (such as VIN13 and 285) were shown to be generally higher producers of most acids in white and/ or red wine fermentation settings, while other strains (such as DV10) were generally lower acid producers. The data clearly demonstrate that different strains have different acid consumption and production patterns, and this presents a first step towards enabling winemakers to appropriately select strains for acid management during fermentation.

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