Appl. microbiol. biotechnol.

Non-Saccharomyces yeast species, naturally found in grape must, may impact wine quality positively or negatively. In this study, a mixture of five non-Saccharomyces species (Torulaspora delbrueckii, Metschnikowia spp., Starmerella bacillaris (formerly called Candida zemplinina), Hanseniaspora uvarum, Pichia kluyveri), mimicking the composition of the natural non-Saccharomyces community found in grape must, was used for alcoholic fermentation. The impact of CO2 saturation of the grape juice was studied first on this mixture alone, and then in the presence of Saccharomyces cerevisiae. Two isogenic strains of this species were used: the first with a short and the second a long fermentation lag phase. This study demonstrated that saturating grape juice with CO2 had interesting potential as an oenological technique, inhibiting undesirable species (S. bacillaris and H. uvarum) and stimulating non-Saccharomyces of interest (T. delbrueckii and P. kluyveri). This stimulating effect was particularly marked when CO2 saturation was associated with the presence of S. cerevisiae with long fermentation lag phase. The direct consequence of this association was an enhancement of 3-SH levels in the resulting wine

J Microbiol Methods

The existing methods for testing proteolytic activity are time consuming, quite difficult to perform, and do not allow real-time monitoring. Proteases have attracted considerable interest in winemaking and some yeast species naturally present in grape must, such as Metschnikowia pulcherrima, are capable of expressing this activity. In this study, a new test is proposed for measuring proteolytic activity directly in fermenting grape must, using azocasein, a chromogenic substrate. Several yeast strains were tested and differences in proteolytic activity were observed. Moreover, analysis of grape must proteins in wines revealed that protease secreted by Metschnikowia strains may be active against wine proteins

Corrigendum to “A new method for monitoring the extracellular proteolytic activity of wine yeasts during alcoholic fermentation of grape must” [J. Microbiol. Methods 119 (2015) 176–179]

Corrigendum to “A new method for monitoring the extracellular proteolytic activity of wine yeasts during alcoholic fermentation of grape must” [J. Microbiol. Methods 119 (2015) 176–179

Restriction patterns of D1/D2 amplicon generated by <i>Alu</i>I (A) or <i>Pst</i>I (B) for <i>Torulaspora</i> species.

<p>A: For <i>Alu</i>I restriction, four patterns were produced: 170 pb+160 pb+80 pb+70 pb+55 pb+40 pb+30 pb for <i>T. delbrueckii</i> and <i>T. quercuum</i>; 170 pb+160 pb+120 pb+70 pb+55 pb+30 pb for <i>T. maleeae</i> and <i>T. indica</i>; 170 pb+160 pb+95 pb+80 pb+70 pb+30 pb for <i>T. franciscae</i>, <i>T. microellipsoides</i>, <i>T. pretoriensis</i>; and 330 pb+170 pb+75 pb+30 pb for <i>T. globosa</i>. B: For <i>Pst</i>I restriction, two patterns were produced: 600pb (no restriction) for <i>T. maleeae</i>, <i>T. quercuum</i>, <i>T. indica</i>, <i>T. microellipsoides</i> and <i>T. globosa</i>; or 480 pb+120 pb for <i>T. delbrueckii</i>, <i>T. franciscae</i> and <i>T. pretoriensis</i>. Blue and pink bands represent internal upper and lower markers respectively.</p

Genetic relationships between 110 <i>T. delbrueckii</i> strains using eight microsatellite markers.

<p>A: Dendrogram tree built using Bruvo's distance and Neighbor-Joining's clustering. The robustness of the node was assessed using multiscale bootstrap resampling and approximated unbiased test (n = 1000 boots). Bootstrap results are shown only for the main nodes. B: Barplot representing structure results (K = 5). The posterior probability (y-axis) of assignment of each strain (vertical bar) to ancestral groups is shown by colors (dark green, green, blue, red and darkblue colors represent each 5 ancestral populations). Heterozygous strains, meaning strains with at least one heterozygote locus, are indicated by black stars.</p

Microsatellite loci for <i>Torulaspora delbrueckii</i> genotyping.

<p>Allele size in pb. Forward primers were tailed on 5′-end with M13 sequence (CACGACGTTGTAAAACGAC). Tm is the melting temperature used for microsatellite amplification (see Materials and Methods). CLIB230^T is synonymous of CBS 1146^T.</p

Genetic relationships between 110 <i>T. delbrueckii</i> strains using eight microsatellite markers.

<p>A: Dendrogram tree built using Bruvo's distance and Neighbor-Joining's clustering. The robustness of the node was assessed using multiscale bootstrap resampling and approximated unbiased test (n = 1000 boots). Bootstrap results are shown only for the main nodes. B: Barplot representing structure results (K = 5). The posterior probability (y-axis) of assignment of each strain (vertical bar) to ancestral groups is shown by colors (dark green, green, blue, red and darkblue colors represent each 5 ancestral populations). Heterozygous strains, meaning strains with at least one heterozygote locus, are indicated by black stars.</p

F-statistics and observed heterozygosity in <i>Torulaspora delbrueckii</i> population.

<p><i>F_IT</i> represents the total deficit of heterozygotes, <i>F_IS</i> the deficit of heterozygotes within the population, <i>F_ST</i> the fixation index. *** indicates a significant effect at 0.1%. Ho stands for observed heterozygosity, and did not fit the Hardy-Weinberg hypothesis (pval<<0.001) for all eight loci.</p

Appl Microbiol Biotechnol

Non-Saccharomyces (NS) species that are either naturally present in grape must or added in mixed fermentation with S. cerevisiae may impact the wine's chemical composition and sensory properties. NS yeasts are prevailing during prefermentation and early stages of alcoholic fermentation. However, obtaining the correct balance between S. cerevisiae and NS species is still a critical issue: if S. cerevisiae outcompetes the non-Saccharomyces, it may minimize their impact, while conversely if NS take over S. cerevisiae, it may result in stuck or sluggish fermentations. Here, we propose an original strategy to promote the non-Saccharomyces consortium during the prefermentation stage while securing fermentation completion: the use of a long lag phase S. cerevisiae. Various fermentations in a Sauvignon Blanc with near isogenic S. cerevisiae displaying short or long lag phase were compared. Fermentations were performed with or without a consortium of five non-Saccharomyces yeasts (Hanseniaspora uvarum, Candida zemplinina, Metschnikowia spp., Torulaspora delbrueckii, and Pichia kluyveri), mimicking the composition of natural NS community in grape must. The sensorial analysis highlighted the positive impact of the long lag phase on the wine fruitiness and complexity. Surprisingly, the presence of NS modified only marginally the wine composition but significantly impacted the lag phase of S. cerevisiae. The underlying mechanisms are still unclear, but it is the first time that a study suggests that the wine composition can be affected by the lag phase duration per se. Further experiments should address the suitability of the use of long lag phase S. cerevisiae in winemaking