The aim of this study was to characterise and evaluate a selection of South African T. delbrueckii yeast isolates under laboratory conditions.8
The yeast population found on grape and winery equipment surfaces usually consists of Saccharomyces cerevisiae, and other species broadly referred to as non-Saccharomyces yeasts.1 While the wine yeast S. cerevisiae is the desired yeast for completion of alcoholic fermentation, the non-Saccharomyces yeasts also play a role.1,2,3 These yeasts typically include species from the genera Hanseniaspora (Kloeckera), Pichia, Torulaspora, Metschnikowia, Lachancea (Kluyveromyces), Zygosaccharomyces and Candida. One non-Saccharomyces yeast that has received much attention in wine industries is Torulaspora delbrueckii (also known as Candida colliculosa).1,4,5,6
T. delbrueckii yeast is found naturally in most wine producing regions of the world, and relatively recently commercial strains have become available. However, the use of T. delbrueckii yeast for winemaking has been suggested as far back as 1948 for musts low in sugar and acid, and for the production of red and rosé wines in Italy.7 Recent studies showed that T. delbrueckii yeasts produce lower levels of volatile acidity (VA) and acetaldehyde in comparison to S. cerevisiae, and can contribute positively to the flavour of wines.1,2,3,4 The changing taxonomic description of this species in the past, together with the natural genetic variation found amongst yeasts, means there is potential for strains with improved performance in comparison to the commercial strains.
Materials and methods
Forty-three T. delbrueckii yeast strains isolated from South African vineyards and wines were investigated. Reference yeasts included two commercial T. delbrueckii strains, the T. delbrueckii type strain and one commercial S. cerevisiae wine yeast.
The yeasts were characterised by various molecular, biochemical and physiological methods. Laboratory-scale fermentations were conducted at 22 and 15°C, respectively, in a clarified Chenin blanc/Chardonnay (1:1 ratio) grape must blend (21.5°B; total acidity: 7.0 g/ℓ; and pH: 3.69).
Results and discussion
In comparison to S. cerevisiae, the deliberate use of non-Saccharomyces yeast in wine production is new. While an estimated 200 commercial S. cerevisiae yeasts are available world-wide, only a limited number of T. delbrueckii commercial yeast strains are available to wine industries.1 Although the number of commercial T. delbrueckii yeast strains will probably never reach that of S. cerevisiae, there is undoubtedly scope for more strains for commercial winemaking.
The various characterisation methods used in this investigation made it possible to differentiate between, and to confirm the identity of the T. delbrueckii yeast strains. The South African strains could be divided into 13 groups. The groups contained 17, seven, six, three and two strains, respectively, while the remaining eight groups contained one strain each. None of the groups matched that of the profiles of the T. delbrueckii reference strains, underlining the wide genetic diversity amongst the strains.
The fermentation data of the 22°C trial showed that the T. delbrueckii yeasts fell into two distinct groups, a faster (18 strains) and a slower (25 strains) fermenting group. The two commercial T. delbrueckii strains and the S. cerevisiae reference yeast were part of the faster fermenting group. One South African strain finished the fermentation at a similar rate to that of the S. cerevisiae reference yeast, thus showing potential for use in further fermentation studies.
The fermentation data of the trial carried out at 15°C showed that the yeasts also fermented at different rates, 14 faster fermenting strains and 29 slower fermenting strains. However, the spread between the fermentation rates was greater than at 22°C. The best T. delbrueckii strain at 22°C, also performed well at 15°C. Overall, the T. delbrueckii yeasts took longer to ferment the must at 15°C (26 days) compared to 22°C (19 days). The fermentation abilities and the 13 biochemical and physiological groupings did not coincide. T. delbrueckii strains with rapid fermentation abilities were found in many of the 13 groups.
Chemical analyses performed on the resultant laboratory-scale wines showed that the VA values for most of the yeast strains were similar (0.3 – 0.5 g/ℓ) at 15°C and 22°C. One T. delbrueckii strain produced VA levels nearly double than that of the other yeasts (0.7 g/ℓ at 15°C and 1.0 g/ℓ at 22°C). Although still within the permissible limit (1.2 g/ℓ) for South African wines, the use of this particular strain would not be recommended for wine production.
The concentration of glycerol produced by most of the T. delbrueckii strains was generally higher than that of the S. cerevisiae reference yeast and ranged from 5.0 – 13 g/ℓ at 22°C, and from 5.0 – 7.9 g/ℓ at 15°C. Glycerol is one of the major products of alcoholic fermentation. It can have a notable effect on sweetness and mouth-feel in dry wines in concentrations above 5.2 g/ℓ, although it has no direct impact on the aromatic characteristics of wine.
The residual sugar levels of the must fermented at 22°C showed 10 T. delbrueckii yeasts that fermented to below 10 g/ℓ; three with residual sugars between 10 and 30 g/ℓ and the remainder with residual sugars above 30 g/ℓ. The isolate with the fastest fermentation rate was able to ferment the must to dryness (<5 g/ℓ RS) as defined by South African legislation. The residual sugar levels of the 15°C fermentation trials showed six T. delbrueckii yeasts that fermented to below 10 g/ℓ, nine with residual sugar between 10 and 30 g/ℓ and the remainder with residual sugar above 30 g/ℓ. The best performing strain at 22°C was also able to ferment the must to dryness at 15°C. The other strains would have to be co-inoculated with a S. cerevisiae to ensure dry wines.
Some of the wines had higher total SO2 levels than the reference strains, but still within the permissible limits for South African wines. High levels of SO2 are not desirable as this could negatively affect the wine quality, and inhibit sensitive co-inoculant wine yeasts and malolactic bacteria.
Characterisation of the South African T. delbrueckii strains showed a large genetic diversity. None of them matched the biochemical profiles of the commercial strains or type strain. The laboratory-scale fermentations indicated that not all of the T. delbrueckii yeast strains were suitable for wine production. However, some showed potential for use as single inoculants, or as co-inoculants with a S. cerevisiae yeast strain. Therefore, depending on the fermentation temperature, different T. delbrueckii strains would be suitable for specific wine styles. These promising yeast strains would require further studies to evaluate their performance under industry conditions.
Torulaspora delbrueckii yeast strains isolated from South African grapes and must, T. delbrueckii reference strains and a S. cerevisiae reference yeast were characterised using conventional and molecular microbiological techniques. Based on the characterisation results the T. delbrueckii strains were divided into 13 groups. The performances of these yeasts were evaluated in grape must in laboratory-scale fermentations at 15 and 22°C. The fermentation data showed that the yeasts fell into two distinct groups, a fast and a slow fermenting group. Chemical analyses of the resultant laboratory-scale wines (alcohol, volatile acidity, glycerol, total SO2 and residual sugar) showed some of the T. delbrueckii strains produced wines with acceptable chemical profiles at both temperatures. Therefore, depending on the fermentation temperature, different T. delbrueckii strains would be suitable for specific wine styles and some may be considered for single inoculations without the addition of S. cerevisiae.
The authors wish to thank Winetech, the ARC and CPUT for financial support and the ARC Infruitec-Nietvoorbij Post Harvest and Wine Technology division for the use of their facilities, as well as the Microbiology team for technical assistance.
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