Despite its promising potential, the capabilities of Saccharomyces paradoxus in commercial winemaking are still unutilized and require further investigation. In this study, the effects of fermentation by a S. paradoxus strain P01-161 on the composition of Malvazija istarska white wine in two harvest years were investigated. A range of complementary standard and metabolomics analysis approaches were applied, including OIV methods for basic parameters; HPLC-DAD-RI for organic acids, glycerol, and proteins; UPLC/MS/MS for phenolic compounds; and GC/FID, GC/MS, and GC × GC/TOF-MS for volatile compounds. The harvest year exhibited a significant impact, but many distinctive traits of S. paradoxus versus S. cerevisiae control wines were consistent across the seasons. These included reductions in malic acid and certain phenols and pathogenesis-related proteins. Saccharomyces paradoxus fermentation yielded higher levels of glycerol, volatile acidity, and specific thaumatin-like proteins. Among a total of 474 identified volatile compounds, S. paradoxus exhibited lower concentrations of several odoriferous alcohols, acids, and esters, as well as higher concentrations of β-damascenone, acetaldehyde, isobutyric acid, ethyl 2-methylbutyrate, ethyl acetate, isobutyl acetate, various esters of succinic and lactic acids, accompanied by numerous minor compounds, when compared to S. cerevisiae. These differences suggest the potential for distinct sensory profiles produced by the two yeasts, indicating that S. paradoxus could be a promising alternative for white wine production.

Climate change is strongly affecting the winemaking sector, notably by decreasing wine acidity due to lower malic acid levels in the grapes. Wine-related microorganisms can greatly affect the organic acid contained in wines as they are able to metabolise or synthesise different acids. Major advances in biochemistry, ecophysiology and molecular biology have led to numerous yeast strains being selected for that have specific oenological properties, including acidity modulation. The yeast Saccharomyces cerevisiae is the most extensively studied species, harbouring both malic acid-consuming and producing strains which are of interest in various vinification itineraries. Yeast-derived acidification of wines can indeed be achieved via malic acid production by S. cerevisiae, as well as via lactic acid production by Lachancea thermotolerans. Co-fermentations of these two species become promising tools to manage wine acidity while ensuring fermentation completion and wine quality. Deacidification of wines via malic acid consumption is relevant in cooler winemaking regions, and/or for shortening malolactic fermentation and thereby increasing wine stability. This review delivers an in-depth overview of the effect of various oenologically relevant yeasts on wine acidity, with a focus on the latest findings on novel (de)acidifying S. cerevisiae strains.

There is a lack of studies evaluating the metabolic contribution of non-Saccharomyces yeasts in early fermentation phases. This study aimed to investigate the volatile aroma profiles produced by various non-Saccharomyces yeasts just before sequential inoculation with Saccharomyces cerevisiae to provide an insight into the particular effects they induce at this stage. The grape must of Malvazija istarska was inoculated with monocultures of Torulaspora delbrueckii, Metschnikowia pulcherrima, Pichia kluyveri, Lachancea thermotolerans, and Schizosaccharomyces pombe, alongside a S. cerevisiae control. Eighty volatile compounds were quantified via headspace solid-phase microextraction and gas chromatography–mass spectrometry, and the data were statistically elaborated. Volatile profiles of non-Saccharomyces yeasts differed significantly from the S. cerevisiae control. Most treatments caused increases in linalool and β-damascenone, decreases in higher alcohols and fatty acids, and improved synthesis of odoriferous esters. Torulaspora delbrueckii and M. pulcherrima produced compounds not commonly found in S. cerevisiae fermented wines. Multivariate statistical analysis linked the investigated yeasts to specific, particularly abundant compounds. Future studies should explore to what degree these contributions persist after sequential inoculation with S. cerevisiae in diverse grape must matrices.

Viognier is a warm climate grape variety prone to loss of acidity and accumulation of excessive sugars. The yeast Lachancea thermotolerans can improve the stability and balance of such wines due to the partial conversion of sugars to lactic acid during alcoholic fermentation. This study compared the performance of five L. thermotolerans strains in co-inoculations and sequential inoculations with Saccharomyces cerevisiae in high sugar/pH Viognier fermentations. The results highlighted the dichotomy between the non-acidified and the bio-acidified L. thermotolerans treatments, with either comparable or up to 0.5 units lower pH relative to the S. cerevisiae control. Significant differences were detected in a range of flavour-active yeast volatile metabolites. The perceived acidity mirrored the modulations in wine pH/TA, as confirmed via "Rate-All-That-Apply" sensory analysis. Despite major variations in the volatile composition and acidity alike, the varietal aromatic expression (i.e., stone fruit aroma/flavour) remained conserved between the treatments.

The yeast Lachancea thermotolerans can produce lactic acid during alcoholic fermentation (AF) and thereby acidify wines with insufficient acidity. However, little is known about the impact of L. thermotolerans on Oenococcus oeni, the primary lactic acid bacterium used in malolactic fermentation (MLF). This study explored the impact of sequential cultures of L. thermotolerans and Saccharomyces cerevisiae on MLF performance in white and red wines. Four L. thermotolerans strains were tested in Sauvignon blanc with sequential S. cerevisiae inoculation, compared to an S. cerevisiae control and the initially un-inoculated treatments. The L. thermotolerans wines showed large differences in acidification, and progression of MLF depended on lactic acid production, even at controlled pH. The highest and lowest lactic acid producing strains were tested further in Merlot fermentations with both co-inoculated and sequentially inoculated O. oeni. The low lactic acid producing strain enabled successful MLF, even when this failed in the S. cerevisiae treatment, with dramatically quicker malic acid depletion in O. oeni co-inoculation than in sequential inoculation. In contrast, a high lactic acid producing strain inhibited MLF irrespective of the O. oeni inoculation strategy. In a follow-up experiment, increasing concentrations of exogenously added lactic acid slowed MLF and reduced O. oeni growth across different matrices, with 6 g/L of lactic acid completely inhibiting MLF. The results confirm the inhibitory effect of lactic acid on O. oeni while highlighting the potential of some L. thermotolerans strains to promote MLF and the others to inhibit it.

In Latin, 'pulcherrima' is a superlative form of an adjective that translates as beautiful. Apart from being 'the most beautiful' yeast, Metschnikowia pulcherrima has a remarkable potential in production of wines with lower ethanol content. The oenological performance of six M. pulcherrima strains was hereby tested in sequential cultures with Saccharomyces cerevisiae. The best-performing strain MP2 was further characterised in fermentations with different S. cerevisiae inoculation delays in both white grape juice and Chemically Defined Grape Juice Medium (CDGJM). The analysis of main metabolites, undertaken prior to sequential inoculations and upon fermentation completion, highlighted metabolic interactions and carbon sinks other than ethanol in MP2 treatments. Depending on the inoculation delay, MP2 white wines contained between 0.6% and 1.2% (v/v) less ethanol than the S. cerevisiae monoculture, with even larger decreases detected in the CDGJM. The MP2 treatments also contained higher concentrations of TCA cycle by-products (i.e. fumarate and succinate) and glycerol, and lower concentrations of acetic acid. The analysis of volatile compounds showed increased production of acetate esters and higher alcohols in all MP2 wines, alongside other compositional alterations arising from the S. cerevisiae inoculation delay.

A changing climate has led to winegrapes being harvested with increased sugar levels and at greater risk of berry shrivel. A suggested easy-to-adopt strategy to manage the associated rising wine alcohol levels is the pre-fermentative substitution of juice with either "green harvest wine" or water. Our study investigates the effects of this approach on Vitis vinifera L. cv. Cabernet Sauvignon wine quality attributes. Wines were also made from fruit collected at consecutive earlier harvest time points to produce wines comparable in alcohol to the substituted wines. Tannin concentrations and colour did not change significantly in the wines with modified alcohol content even at higher juice substitution rates. Differences in polysaccharide and tannin composition indicated variability in extraction dynamics according to substitution rate and type of blending component. In scenarios where berry shrivel is inevitable, the incorporation of water in particular offers much promise as part of a strategy to manage wine alcohol content.

The yeast Lachancea thermotolerans (previously Kluyveromyces thermotolerans) is a species of large, yet underexplored, oenological potential. This study delivers comprehensive oenological phenomes of 94 L. thermotolerans strains obtained from diverse ecological niches worldwide, classified in nine genetic groups based on their pre-determined microsatellite genotypes. The strains and the genetic groups were compared for their alcoholic fermentation performance, production of primary and secondary metabolites and pH modulation in Chardonnay grape juice fermentations. The common oenological features of L. thermotolerans strains were their glucophilic character, relatively extensive fermentation ability, low production of acetic acid and the formation of lactic acid, which significantly affected the pH of the wines. An untargeted analysis of volatile compounds, used for the first time in a population-scale phenotyping of a non-Saccharomyces yeast, revealed that 58 out of 90 volatiles were affected at an L. thermotolerans strain level. Besides the remarkable extent of intra-specific diversity, our results confirmed the distinct phenotypic performance of L. thermotolerans genetic groups. Together, these observations provide further support for the occurrence of domestication events and allopatric differentiation in L. thermotolerans population.

Background and Aims: The choice of yeast strain(s) to conduct the fermentation can greatly affect wine chemical and sensory profile. Even though the use of non-Saccharomyces co-inocula to build complexity and diversify styles is increasingly in vogue, a limited number of such products are available to date, and more research is required to guide their use in the wine industry. This study evaluates the potential of commercial yeast inocula to modulate the quality of Shiraz wines at two maturity levels. Methods and Results: Vinification outcomes of eight yeast treatments were compared in earlier (24 degrees Brix) and later (29 degrees Brix) harvested Shiraz fruit. Yeast treatments included five non-Saccharomyces products with sequentially inoculated Saccharomyces cerevisiae, a commercial blend of non-Saccharomyces and S. cerevisiae strains, and a S. cerevisiae inoculum. Fermentation monitoring, and comprehensive analytical profiling in terms of basic chemistry, volatile composition, phenolic measurements and descriptive sensory analysis, allowed for the comparison of the resulting wines. Both harvest date and yeast inoculation treatments had a significant impact on a range of compositional and, in turn, sensory parameters of the wines. Conclusions: Certain non-Saccharomyces sequential inoculation treatments led to increased appeal of earlier harvest wines compared to the S. cerevisiae Control. These treatments, however, were related to an increased risk of arrested fermentation in higher ripeness conditions. Significance of the Study: This study contributes to a better understanding of yeast inoculum-derived modulation of Shiraz wine quality parameters at different maturity levels.

The yeast Lachancea thermotolerans (formerly Kluyveromyces thermotolerans) is a species with remarkable, yet underexplored, biotechnological potential. This ubiquist occupies a range of natural and anthropic habitats covering a wide geographic span. To gain an insight into L. thermotolerans population diversity and structure, 172 isolates sourced from diverse habitats worldwide were analysed using a set of 14 microsatellite markers. The resultant clustering revealed that the evolution of L. thermotolerans has been driven by the geography and ecological niche of the isolation sources. Isolates originating from anthropic environments, in particular grapes and wine, were genetically close, thus suggesting domestication events within the species. The observed clustering was further validated by several means including, population structure analysis, F-statistics, Mantel's test and the analysis of molecular variance (AMOVA). Phenotypic performance of isolates was tested using several growth substrates and physicochemical conditions, providing added support for the clustering. Altogether, this study sheds light on the genotypic and phenotypic diversity of L. thermotolerans, contributing to a better understanding of the population structure, ecology and evolution of this non-Saccharomyces yeast.

Renata Ristic, Ana Hranilovic, Sijing Li, Rocco Longo, Duc-Truc Pham, Bora Qesja, Olaf J. Schelezki, and Vladimir Jiranek