Nicotinic acid availability impacts redox cofactor metabolism in Saccharomyces cerevisiae during alcoholic fermentation.

Anaerobic alcoholic fermentation, particularly in high-sugar environments, presents metabolic challenges for yeasts. Crabtree-positive yeasts, including Saccharomyces cerevisiae, prefer fermentation even in the presence of oxygen. These yeasts rely on internal NAD+ recycling and extracellular assimilation of its precursor, nicotinic acid (vitamin B3), rather than de novo NAD+ production. Surprisingly, nicotinic acid assimilation is poorly characterized, even in S. cerevisiae. This study elucidated the timing of nicotinic acid uptake during grape juice-like fermentation and its impact on NAD(H) levels, the NAD+/NADH ratio, and metabolites produced. Complete uptake of extracellular nicotinic acid occurred premid-exponential phase, thereafter small amounts of vitamin B3 were exported back into the medium. Suboptimal levels of nicotinic acid were correlated with slower fermentation and reduced biomass, disrupting redox balance and impeding NAD+ regeneration, thereby affecting metabolite production. Metabolic outcomes varied with nicotinic acid concentrations, linking NAD+ availability to fermentation efficiency. A model was proposed encompassing rapid nicotinic acid uptake, accumulation during cell proliferation, and recycling with limited vitamin B3 export. This research enhances the understanding of nicotinic acid uptake dynamics during grape juice-like fermentation. These insights contribute to advancing yeast metabolism research and have profound implications for the enhancement of biotechnological practices and the wine-making industry.

Differences in the management of intracellular redox state between wine yeast species dictate their fermentation performances and metabolite production.

The maintenance of the balance between oxidised and reduced redox cofactors is essential for the functioning of many cellular processes in all living organisms. While the electron transport chain plays a key role in maintaining this balance under respiratory conditions, its inactivity in the absence of oxygen poses a challenge that yeasts such as Saccharomyces cerevisiae overcome through the production of various metabolic end-products during alcoholic fermentation. In this study, we investigated the diversity occurring between wine yeast species in their management of redox balance and its consequences on the fermentation performances and the formation of metabolites. To this aim, we quantified the changes in NAD(H) and NADP(H) concentrations and redox status throughout the fermentation of 6 wine yeast species. While the availability of NADP and NADPH remained balanced and stable throughout the process for all the strains, important differences between species were observed in the dynamics of NAD and NADH intracellular pools. A comparative analysis of these data with the fermentation capacity and metabolic profiles of the strains revealed that Saccharomyces cerevisiae, Torulaspora delbrueckii and Lachancea thermotolerans strains were able to reoxidise NADH to NAD throughout the fermentation, mainly by the formation of glycerol. These species exhibited good fermentation capacities. Conversely, Starmerella bacillaris and Metschnikowia pulcherrima species were unable to regenerate NAD as early as one third of sugars were consumed, explaining at least in part their poor growth and fermentation performances. The Kluyveromyces marxianus strain exhibited a specific behaviour, by maintaining similar levels of NAD and NADH throughout the process. This balance between oxidised and reduced redox cofactors ensured the consumption of a large part of sugars by this species, despite a low fermentation rate. In addition, the dynamics of redox cofactors affected the production of by-products by the various strains either directly or indirectly, through the formation of precursors. Major examples are the increased formation of glycerol by S. bacillaris and M. pulcherrima strains, as a way of trying to reoxidise NADH, and the greater capacity to produce acetate and derived metabolites of yeasts capable of maintaining their redox balance. Overall, this study provided new insight into the contribution of the management of redox status to the orientation of yeast metabolism during fermentation. This information should be taken into account when developing strategies for more efficient and effective fermentation.

Characterization of Mannoprotein Structural Diversity in Wine Yeast Species.

The structure of yeast cell wall (CW) mannoproteins (MPs) influences their impact on wine properties. Yeast species produce a diverse range of MPs, but the link between properties and specific structural features has been ill-characterized. This study compared the protein and polysaccharide moieties of MP-rich preparations from four strains of four different enologically relevant yeast species, named Saccharomyces boulardii (SB62), Saccharomyces cerevisiae (SC01), Metschnikowia fructicola (MF77), and Torulaspora delbrueckii (TD70), and a commercial MP preparation. Monosaccharide determination revealed that SB62 MPs contained the highest mannose/glucose ratio followed by SC01, while polysaccharide size distribution analyses showed maximum molecular weights ranging from 1349 kDa for MF77 to 483 kDa for TD70. Protein identification analysis led to the identification of unique CW proteins in SB62, SC01, and TD70, as well as some proteins shared between different strains. This study reveals MP composition diversity within wine yeasts and paves the way toward their industrial exploitation.

Redox cofactor metabolism in Saccharomyces cerevisiae and its impact on the production of alcoholic fermentation end-products.

The alcoholic fermentation of organic carbon sources by Saccharomyces cerevisiae produces many by-products, with the most abundant originating from central carbon metabolism. The production of these metabolites involves redox reactions and largely depends on the maintenance of redox homeostasis. Despite the metabolic pathways being mostly conserved across strains of S. cerevisiae, their production of various amounts of metabolic products suggests that their intracellular concentration of redox cofactors and/or redox balance differ. This study explored the redox status dynamics and NAD(H) and NADP(H) cofactor ratios throughout alcoholic fermentation in four S. cerevisiae strains that exhibit different carbon metabolic fluxes. This study focussed on the molecular end-products of fermentation, redox cofactor ratios and the impact thereof on redox homeostasis. Strain-dependent differences were identified in the redox cofactor levels, with NADP(H) ratios and levels remaining stable while NAD(H) levels decreased drastically as the fermentation progressed. Changes in the NAD+/NADH ratio were also observed. Total levels of NAD(H) decreased drastically as the fermentation progressed despite the cells remaining viable until the end of fermentation. NAD+ was found to be favoured initially while NADH was favoured towards the end of the fermentation. The change in the NAD+/NADH redox cofactor ratio during fermentation was linked with the production of end-products. The findings in this study could steer further research in the selection of S. cerevisiae wine strains for desirable aroma contributions based on their intracellular redox balance management.

Species-Dependent Metabolic Response to Lipid Mixtures in Wine Yeasts.

Lipids are essential energy storage compounds and are the core structural elements of all biological membranes. During wine alcoholic fermentation, the ability of yeasts to adjust the lipid composition of the plasma membrane partly determines their ability to cope with various fermentation-related stresses, including elevated levels of ethanol and the presence of weak acids. In addition, the lipid composition of grape juice also impacts the production of many wine-relevant aromatic compounds. Several studies have evaluated the impact of lipids and of their metabolism on fermentation performance and aroma production in the dominant wine yeast Saccharomyces cerevisiae, but limited information is available on other yeast species. Thus, the aim of this study was to evaluate the influence of specific fatty acid and sterol mixtures on various non-Saccharomyces yeast fermentation rates and the production of primary fermentation metabolites. The data show that the response to different lipid mixtures is species-dependent. For Metschnikowia pulcherrima, a slight increase in carbon dioxide production was observed in media enriched with unsaturated fatty acids whereas Kluyveromyces marxianus fermented significantly better in synthetic media containing a higher concentration of polyunsaturated fatty acids than monounsaturated fatty acids. Torulaspora delbrueckii fermentation rate increased in media supplemented with lipids present at an equimolar concentration. The data indicate that these different responses may be linked to variations in the lipid profile of these yeasts and divergent metabolic activities, in particular the regulation of acetyl-CoA metabolism. Finally, the results suggest that the yeast metabolic footprint and ultimately the wine organoleptic properties could be optimized via species-specific lipid adjustments.

Exploring the phenotypic diversity of oenological traits in Kluyveromyces marxianus strains.

The use of non-Saccharomyces yeasts in the winemaking process may have several positive outcomes. Kluyveromyces marxianus has recently been revealed as a promising species for this industry. While the majority of studies mention the use of K. marxianus in various industries including food production (e.g. dairy and cocoa), recent studies have also shown that its aroma and pectinase production make it a suitable yeast for the wine industry. Nevertheless, only particular strain, IWBT Y885, was investigated. In this study, five different K. marxianus strains as well as one protoplast fusant (BF2020) were compared to strain Y885. These comparisons focused on various oenological traits such as fermentation performance, fermentation metabolites, hydrogen sulfide, and pectinase production. Throughout the study, variations were found between the K. marxianus strains investigated. Indeed, although common traits such as high pectinase activity appeared conserved among K. marxianus strains, a fairly large phenotypic diversity was also evident. Using cluster analysis, strain groupings emerged with strains L01, L05, Y885, and BF2020 grouping together. Similarly, strains L02 and L04 grouped together while strain L03 appearing to show the most variation between the strains investigated. Variation between strains was observed regardless of the original source of isolation.

Phenotypic characterization of cell-to-cell interactions between two yeast species during alcoholic fermentation.

Microbial multispecies ecosystems are responsible for many biotechnological processes and are particularly important in food production. In wine fermentations, in addition to the natural microbiota, several commercially relevant yeast species may be co-inoculated to achieve specific outcomes. However, such multispecies fermentations remain largely unpredictable because of multilevel interactions between naturally present and/or co-inoculated species. Understanding the nature of such interactions has therefore become essential for successful implementation of such strategies. Here we investigate interactions between strains of Saccharomyces cerevisiae and Lachancea thermotolerans. Co-fermentations with both species sharing the same bioreactor (physical contact) were compared to co-fermentations with physical separation between the species in a membrane bioreactor ensuring free exchange of metabolites. Yeast culturability, viability and the production of core metabolites were monitored. The previously reported negative interaction between these two yeast species was confirmed. Physical contact greatly reduced the culturability and viability of L. thermotolerans and led to earlier cell death, compared to when these yeasts were co-fermenting without cell-cell contact. In turn, in the absence of cell-cell contact, L. thermotolerans metabolic activity led to an earlier decline in culturability in S. cerevisiae. Cell-cell contact did not result in significant differences in the major fermentation metabolites ethanol, acetic acid and lactic acid, but impacted on the production of some volatile compounds.

Comparative uptake of exogenous thiamine and subsequent metabolic footprint in Saccharomyces cerevisiae and Kluyveromyces marxianus under simulated oenological conditions.

Managed inoculation of non-Saccharomyces yeast species is regarded as a practical way to introduce new characteristics to wine. However, these yeasts struggle to survive until fermentation is complete. Kluyveromyces marxianus IWBT Y885 is one such yeast. Although it displays interesting oenological properties, a longer persistence during alcoholic fermentation would warranty a stronger impact on wine composition. A key factor for survival, growth and sustained metabolic activity of all yeasts is their nutrient requirements. Thus, identifying nutrients that are essential for maximising fermentation performance, and subsequently ensuring adequate levels of nutrients, is a means to ensure significant contribution of yeasts to wine properties. This study aimed to identify essential nutrients, other than previously studied sugars and nitrogen, for maximum impact of K. marxianus Y885, as well as to characterise the outcomes of their utilisation. A multifactorial experimental design was employed to investigate the impact of nutrient concentrations on fermentation performance with K. marxianus Y885 in synthetic must. B-complex vitamins most significantly impacted fermentation performance of K. marxianus Y885 compared to other nutrient groups investigated. Considering the well-established role of the vitamin, thiamine, for maximum fermentation performance during winemaking and the fact that it may be supplemented to wine fermentations legally, the responses to specifically exogenous thiamine concentration for K. marxianus Y885 and Saccharomyces cerevisiae EC1118 were compared in terms of population viability, fermentation rate, total sugars utilised, thiamine assimilation kinetics, and final wine composition. A saturation effect for initial thiamine concentration of K. marxianus Y885 fermentations was characterised, with a maximum fermentation rate and over 90% of available sugars utilisation obtained at 0.25 mg/L. An appreciably larger comparative increase in exponential cell growth rate, maximum population, fermentation rate and total CO2 production for K. marxianus Y885 compared to S. cerevisiae EC1118 revealed a greater necessity for thiamine to ensure maximum fermentation performance. A delayed uptake of thiamine at higher concentrations for K. marxianus Y885 suggested differential regulation of thiamine uptake compared to S. cerevisiae EC1118. In addition, different trends in metabolites produced between species suggest that thiamine concentration impacts the carbon metabolic flux differently in these two yeasts, potentially impacting final wine properties.

Optimised Extraction and Preliminary Characterisation of Mannoproteins from Non-Saccharomyces Wine Yeasts.

The exogenous application of yeast-derived mannoproteins presents many opportunities for the improvement of wine technological and oenological properties. Their isolation from the cell wall of Saccharomycescerevisiae has been well studied. However, investigations into the efficiency of extraction methods from non-Saccharomyces yeasts are necessary to explore the heterogeneity in structure and composition that varies between yeast species, which may influence wine properties such as clarity and mouthfeel. In this study, nine yeast strains were screened for cell wall mannoprotein content using fluorescence microscopy techniques. Four species were subsequently exposed to a combination of mechanical and enzymatic extraction methods to optimize mannoprotein yield. Yeast cells subjected to 4 min of ultrasound treatment applied at 80% of the maximum possible amplitude with a 50% duty cycle, followed by an enzymatic treatment of 4000 U lyticase per g dry cells weight, showed the highest mannoprotein-rich yield from all species. Furthermore, preliminary evaluation of the obtained extracts revealed differences in carbohydrate/protein ratios between species and with increased enzyme incubation time. The results obtained in this study form an important step towards further characterization of extraction treatment impact and yeast species effect on the isolated mannoproteins, and their subsequent influence on wine properties.

The metabolism of lipids in yeasts and applications in oenology.

Lipids are valuable compounds present in all living organisms, which display an array of functions related to compartmentalization, energy storage and enzyme activation. Furthermore, these compounds are an integral part of the plasma membrane which is responsible for maintaining structure, facilitating the transport of solutes in and out of the cell and cellular signalling necessary for cell survival. The lipid composition of the yeast Saccharomyces cerevisiae has been extensively investigated and the impact of lipids on S. cerevisiae cellular functions during wine alcoholic fermentation is well documented. Although other yeast species are currently used in various industries and are receiving increasing attention in winemaking, little is known about their lipid metabolism. This review article provides an extensive and critical evaluation of our knowledge on the biosynthesis, accumulation, metabolism and regulation of fatty acids and sterols in yeasts. The implications of the yeast lipid content on stress resistance as well as performance during alcoholic fermentation are discussed and a particular emphasis is given on non-Saccharomyces yeasts. Understanding lipid requirements and metabolism in non-Saccharomyces yeasts may lead to a better management of these yeast to enhance their contributions to wine properties.

Nitrogen metabolism in three non-conventional wine yeast species: A tool to modulate wine aroma profiles.

The positive impact of certain non-Saccharomyces yeasts on the aromatic profile of wines has been well documented in literature and their industrial use in association with S. cerevisiae is now recommended. Competition between non-Saccharomyces species and Saccharomyces cerevisiae for various nutrients, especially nitrogen sources, greatly impacts the production of aroma compounds. In this study, we further explored the impact of different nitrogen nutrition strategies on the production of carbon and sulphur volatile compounds of three non-Saccharomyces strains, namely Pichia burtonii, Kluyveromyces marxianus, Zygoascus meyerae sequentially inoculated with S. cerevisiae in Sauvignon blanc and Shiraz grape musts. Nitrogen additions were implemented according the specific requirement of each species. At the end of fermentation, we observed specific metabolic signatures for each strain in response to the nature of the nitrogen source suggesting strain-specific metabolic fluxes present. Overall, these results confirmed and further explored the interconnection between nitrogen sources and aroma metabolism (including that of higher alcohols, fatty acids, esters and volatile sulphur compounds), and their variations according to species and the nature of the nitrogen source. The knowledge generated provides new insights to modulate the aroma profile of wines produced with non-Saccharomyces species.

Transcriptomics unravels the adaptive molecular mechanisms of Brettanomyces bruxellensis under SO2 stress in wine condition.

Sulfur dioxide is generally used as an antimicrobial in wine to counteract the activity of spoilage yeasts, including Brettanomyces bruxellensis. However, this chemical does not exert the same effectiveness on different B. bruxellensis yeasts since some strains can proliferate in the final product leading to a negative sensory profile due to 4-ethylguaiacol and 4-ethylphenol. Thus, the capability of deciphering the general molecular mechanisms characterizing this yeast species' response in presence of SO2 stress could be considered strategic for a better management of SO2 in winemaking. A RNA-Seq approach was used to investigate the gene expression of two strains of B. bruxellensis, AWRI 1499 and CBS 2499 having different genetic backgrounds, when exposed to a SO2 pulse. Results revealed that sulphites affected yeast culturability and metabolism, but not volatile phenol production suggesting that a phenotypical heterogeneity could be involved for the SO2 cell adaptation. The transcriptomics variation in response to SO2 stress confirmed the strain-related response in B. bruxellensis and the GO analysis of common differentially expressed genes showed that the detoxification process carried out by SSU1 gene can be considered as the principal specific adaptive response to counteract the SO2 presence. However, nonspecific mechanisms can be exploited by cells to assist the SO2 tolerance; namely, the metabolisms related to sugar alcohol (polyols) and oxidative stress, and structural compounds.

A comparison of the nitrogen metabolic networks of Kluyveromyces marxianus and Saccharomyces cerevisiae.

In grape must, nitrogen is available as a complex mixture of various compounds (ammonium and amino acids). Wine yeasts assimilate these multiple sources in order to suitably fulfil their anabolic requirements during alcoholic fermentation. Nevertheless, the order of uptake and the intracellular fate of these sources are likely to differ between strains and species. Using a two-pronged strategy of isotopic filiation and RNA sequencing, the metabolic network of nitrogen utilization and its regulation in Kluyveromyces marxianus were described, in comparison with those of Saccharomyces cerevisiae. The data highlighted differences in the assimilation of ammonium and arginine between the two species. The data also revealed that the metabolic fate of certain nitrogen sources differed, thereby resulting in the production of various amounts of key wine aroma compounds. These observations were corroborated by the gene expression analysis.

Lachancea yeast species: Origin, biochemical characteristics and oenological significance.

The genus Lachancea, first proposed in 2003, currently comprises 12 valid species, all found to have eight chromosomes. Lachancea spp. occupy a myriad of natural and anthropic habitats, and their geographic as well as ecological origin have been identified as key drivers in the genetic variations amongst strains of several of the species. Lachancea thermotolerans is the type species of the genus and also the most widely explored, especially for its role in fermentation environments. Indeed, L. thermotolerans is desired for its ability to acidify beer and wine through the production of lactic acid, and to enhance aroma and flavor through increased production of various compounds. Similarly, L. fermentati has been characterized for its potential contribution to the chemical composition of these beverages, albeit to a lesser extent, while other species have received little attention. Overall, members of the genus Lachancea form part of the microbiomes in many fermentation ecosystems and contribute directly or indirectly to the modulation of aroma and flavor of different products. The current review provides an overview of this genus, including the latest reports on the genetic and biochemical characteristics of member species, as well as their biotechnological potential.

The utilisation of nitrogenous compounds by commercial non-Saccharomyces yeasts associated with wine.

Commercial wine fermentation is commonly conducted by inoculated strains of Saccharomyces cerevisiae. However, other non-Saccharomyces yeast species have recently become popular co-inoculants. Co-inoculated yeast species compete with each other for nutrients, and such competition may impact fermentation kinetics and aroma production. Understanding the specific nutrient requirements of non-Saccharomyces strains therefore is essential to better characterize the competitive potential of each strain, and to support rational decision making for nutrient supplementation during wine making. This study investigated the nitrogen source preference of commercial non-Saccharomyces yeasts by conducting pure culture and sequential culture fermentations in synthetic grape musts with adjusted nitrogen contents. Amino acid and ammonium uptake varied between yeast species. Lachancea thermotolerans and Torulaspora delbrueckii assimilated more nitrogen at a faster rate than Pichia kluyveri and Metschnikowia pulcherrima. Significant variation in amino acid preference between species was observed. Sequential fermentations confirmed the more competitive behaviour of L. thermotolerans and T. delbrueckii, with consequences for fermentation kinetics and aroma production. Furthermore, the data suggest that declining populations of non-Saccharomyces yeasts release nitrogen and supports the activity of S. cerevisiae. The data provide the most detailed assessment of nitrogen utilisation by the investigated yeast strains in a wine environment.

Investigating the biochemical and fermentation attributes of Lachancea species and strains: Deciphering the potential contribution to wine chemical composition.

Yeasts of various genera are increasingly used alongside Saccharomyces cerevisiae to drive wine fermentations owing to their positive contribution to the organoleptic profile of the resulting wines. One such yeast species is Lachancea thermotolerans. Other species of the genus Lachancea, namely, L. fermentati and L. lanzarotensis have also been isolated from the fermentation environment, but have not received the same degree of attention as L. thermotolerans. The aim of this study was to investigate the oenological potential of these three Lachancea species, regarding their expression of oenologically relevant enzymes, their fermentation attributes and the expression and location of ß-glucosidase during fermentation of synthetic and real grape must (Muscat of Alexandria). In the current study we evaluated three species viz. L. thermotolerans (14 strains), L. fermentati (1 strain) and L. lanzarotensis (2 strains). Our data show that all the species and strains produced ß-glucosidase but with different substrate specificities. Moreover, L. theromotolerans and L. fermentati also produced ß-xylosidase. H2S production, SO2 and ethanol tolerance was variable between species and strains, with the L. lanzarotensis and L. fermentati displaying considerably high H2S production while L. thermotolerans and L. fermentati displayed higher ethanol tolerance. Furthermore, L. fermentati showed higher SO2 tolerance and could proliferate at 20 mg/L total SO2. Interestingly, an increase in ß-glucosidase activity during fermentation did not result in a significant increase in monoterpene concentrations. However, mixed-fermentations with L. fermentati and L. thermotolerans Concerto enhanced geraniol levels. The data show that this activity was mostly cell-associated and constitutively expressed. Sequential fermentations with the Lachancea spp. and S. cerevisiae resulted in wines with significantly altered chemical compositions compared to that obtained from S. cerevisiae inoculated alone. Wines produced from L. thermotolerans and L. lanzarotensis mixed culture fermentations exhibited similar volatile compound composition. Conversely, L. fermentati produced chemically distinct wines consistently associated with high isobutanol and isobutyric acid, and higher monoterpenes. In particular, linalool and geraniol had potential to make perceivable aroma contribution (OAV ≥ 1).

Kluyveromyces marxianus Secretes a Pectinase in Shiraz Grape Must That Impacts Technological Properties and Aroma Profile of Wine.

Since Saccharomyces cerevisiae strains display no to weak pectinase activity, the utilization of external pectinase is a common practice in winemaking to enhance the extraction of compounds located in the grape berry skins during maceration. In this study, the activity of the native endopolygalacturonase of a Kluyveromyces marxianus strain, isolated from grape juice, was characterized in Shiraz grape must during alcoholic fermentation with or without prefermentative cold maceration. The wines made with K. marxianus had a higher methanol concentration, more free-run wine, an altered volatile compound profile, and displayed pectinase activity in cell-free wine samples. Moreover, the results strongly suggest that K. marxianus' pectinase released polygalacturonic acid soluble fragments, unlike fungal pectinases, which mostly release monomers. Overall, this study shows that K. marxianus is an effective pectinase producer in wine with potential benefits for wine properties.

Torulaspora delbrueckii produces high levels of C5 and C6 polyols during wine fermentations.

Non-Saccharomyces yeasts impact wine fermentations and can diversify the flavor profiles of wines. However, little information is available on the metabolic networks of most of these species. Here we show that unlike the main wine yeast Saccharomyces cerevisiae, Torulaspora delbrueckii and to a lesser extent Lachancea thermotolerans produce significant concentrations of C5 and C6 polyols under wine fermentation conditions. In particular, D-arabitol, D-sorbitol and D-mannitol were produced at significant levels. Their release into the extracellular matrix started when that of glycerol ceased. The data also show that polyol production is influenced by initial sugar concentration, repressed by acetic acid and induced in ethanol supplemented media. Moreover, unlike glycerol and sorbitol, mannitol was partially re-assimilated when populations started to decline. The findings suggest that polyol synthesis is a physiological adaptation to stressful conditions characteristic of alcoholic fermentation and that these polyols may serve a similar purpose as glycerol production in S. cerevisiae, including osmoadaptation and redox balancing.

Fermentation performances and aroma production of non-conventional wine yeasts are influenced by nitrogen preferences.

Saccharomyces cerevisiae is currently the most important yeast involved in food fermentations, particularly in oenology. However, several other yeast species occur naturally in grape must that are highly promising for diversifying and improving the aromatic profile of wines. If the nitrogen requirement of S. cerevisiae has been described in detail, those of non-Saccharomyces yeasts remain poorly studied despite their increasingly widespread use in winemaking. With a view to improving the use of non-Saccharomyces yeasts in winemaking, we explored the fermentation performances, the utilisation of nitrogen sources and the volatile compound production of 10 strains of non-conventional yeasts in pure culture. Two different conditions were tested: one mimicking the grape juice's nitrogen composition and one with all the nitrogen sources at the same level. We highlighted the diversity in terms of nitrogen preference and amount consumed among the yeast strains. Some nitrogen sources (arginine, glutamate, glycine, tryptophan and γ-aminobutyric acid) displayed the largest variations between strains throughout the fermentation. Several non-Saccharomyces strains produced important aroma compounds such as higher alcohols, acetate and ethyl esters in significantly higher quantities than S. cerevisiae.