Search Results (66)

Natural wines represent a new trend in winemaking without the use of preservatives and starter cultures, revealing the unique quality traits of grapes, wine, and terroir, but are susceptible to spoilage or undesirable fermentations. This study investigated the microbial populations associated with organic grapes, must, and natural wines of the Limniona red grape variety, focusing on different production stages and fermentation vessels. Samples included immature and ripe grapes, initial and fermenting must, filtered and unfiltered wines, and final bottled and filtered wines. These were analyzed in order to enumerate key groups of microorganisms and identify beneficial yeasts and bacteria of alcoholic and malolactic fermentation, respectively, as well as potential markers of off-flavors. Culture-dependent methods were used to enumerate yeasts and bacteria, while Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) and 16S rRNA sequencing provided taxonomic resolution. Beneficial fermentation microorganisms (especially Saccharomyces yeasts) were scarce in initial grapes, where other contaminants or wild yeasts were present. Gradually, as fermentation progressed, there was a prevalence of Saccharomyces cerevisiae strains of increased diversity in matured wine, as well as several lactic acid bacteria (LAB) of malolactic fermentation. Most LAB were identified as Lactobacillus and Oenococcus species. Other bacteria from environmental sources, irrelevant to alcoholic/malolactic fermentation or spoilage, like Burkholderia, were also present during the vinification process. The type of vessel affected the type of LAB that prevail, with an abundance of Oenococcus in clay vessels versus Lactobacillus species in stainless-steel vessels. Notably, Lentilactobacillus parafarraginis can be linked to off-flavors if they represent a high percentage of the wine microbiota. These findings highlight the importance of understanding, monitoring and controlling microbial succession during production stages in order to prevent sensory faults and ensure the stable quality of natural wines. Full article

Malolactic fermentation (MLF) is a bioprocess driven by lactic acid bacteria (LAB), which is desired in red and highly acidic white wines. Among all LAB, Oenococcus oeni is the main species in wine, followed by Lactiplantibacillus plantarum. The harsh conditions found in wine—not only due to the low nutrient concentration but also the presence of antimicrobial compounds such as ethanol, high acidity, SO_2, and polyphenols—can compromise MLF performance. In recent years, the use of certain non-Saccharomyces yeasts, such as Torulaspora delbrueckii or Metschnikowia pulcherrima, as starter cultures for alcoholic fermentation, has emerged as a promising strategy to improve MLF. In this study, we evaluated the effect of four different fractions from a T. delbrueckii strain on MLF performance. First, the positive impact of this strain as a starter culture on O. oeni growth was confirmed; then, yeast-derived compounds were tested in different wines. Two fractions showed the most promising results in reducing MLF duration: the inactivated yeast fraction and the autolysate fraction. Those enhanced bacterial viability and promoted mannoprotein consumption. These findings highlight the potential of T. delbrueckii-derived compounds as enological tools to support MLF under restrictive wine conditions. Full article

Previous studies have shown the positive effects of non-conventional Metschnikowia spp. yeasts in mixed cultures with Saccharomyces cerevisiae on the properties of fruit wines. In this study, we investigated the effects of using conventional S. cerevisiae and non-conventional Metschnikowia pulcherrima yeasts as starter cultures in controlled mixed fermentations of honey wort. Other non-conventional yeasts were also tested for comparison, including Wickerhamomyces anomalus, Dekkera/Bretannomyces bruxellensis, and Wickerhamomyces anomalus. We evaluated the tolerance of the tested yeasts to high sugar content and analyzed the metabolic profiles of both monocultures and mixed systems. The M. pulcherrima strain showed the highest tolerance to 30% w/v glucose. The chemical complexity of fermented honey was improved using M. pulcherrima in co-starters with S. cerevisiae. The fermented honey samples were characterized by lower ethanol content, higher glycerol level, and rich volatilomes containing higher levels of both esters (ethyl acetate, 3-methylbutyl acetate, 2-methylpropyl acetate) and aliphatic alcohols (2-methylpropan-1-ol, 3-methylbutan-1-ol, and 2-methylbutan-1-ol). Similar characteristics were obtained using mixed populations of four strains: S. cerevisiae, M. pulcherrima, D. bruxellensis, and W. anomalus. Full article

The spoilage of wine caused by Brettanomyces bruxellensis and Hanseniaspora uvarum poses a significant challenge for winemakers, necessitating the development of effective and reliable strategies to control the growth of these yeasts, such as grape must bioprotection. Despite evidence that certain microorganisms can inhibit the growth of Brettanomyces bruxellensis and Hanseniaspora uvarum, the specific mechanisms driving this inhibition remain unclear. The primary objective of this study is to elucidate the underlying mechanisms responsible for this inhibitory effect. We analyzed one Metschnikowia pulcherrima (Mp2) and two Lachancea thermotolerans (Lt29 and Lt45) strains, all of which demonstrated significant killing and inhibitory effects on Brettanomyces bruxellensis (B1 and B250) and Hanseniaspora uvarum (Hu3137) in synthetic must at pH 3.5 and 22 °C. The effectiveness of these two strains exhibited varying inhibition kinetics. The strains were monitored for growth and metabolite production (L-lactic acid, ethanol, and acetic acid) in both single and co-cultures. The low levels of these metabolites did not account for the observed bioprotective effect, indicating a different mechanism at play, especially given the different growth profiles observed with added L-lactic acid and ethanol compared to direct bioprotectant addition. Following the production, purification, and quantification of killer toxins, different concentrations of toxins were tested, showing that the semi-purified Mp2Kt, Lt29Kt, and Lt45Kt toxins controlled the growth of both spoilage yeasts in a dose-dependent manner. These bioprotectant strains also showed compatibility with Saccharomyces cerevisiae in co-cultures, suggesting their potential use alongside commercial starter cultures. Full article

Significant advances in winemaking equipment and processes, as well as a deeper understanding of the role of yeast, have significantly improved wine quality throughout history. This paper examines critical aspects related to the use of commercial wine yeast starter cultures in the fermentation of grape and fruit wines, with a focus on berry wines and blackberry wine, which is the most predominant berry wine in Croatia. While the production of grape wines remains the most significant, fruit wines are gaining importance due to their composition, which contains a variety of bioactive compounds. Although spontaneous fermentation is still preferred by some winemakers, controlled or inoculated fermentation, based on the use of wine yeast starter cultures, is predominantly employed in modern winemaking. The selection of suitable yeast strains for grape wines is easier than for fruit wines, as the broader availability of commercial yeasts for grape wines contrasts with the limited selection offered for fruit wine production due to the smaller fruit wine market. The selection of Saccharomyces and, more recently, non-Saccharomyces yeast strains with desirable characteristics are crucial for the production of high-quality wines. Selection criteria for wine yeasts have evolved to meet modern consumer preferences and focus on technological properties, secondary flavor development and health effects. Full article

The effects of spontaneous fermentation and the inoculation of grape must with a flocculant yeast starter culture, together with the supplementation of must with a commercial organic nitrogen compound (ONC), were analyzed. The microbiome during fermentation was tracked, and volatile compounds in the resulting wines were identified and quantified using gas chromatography and mass spectrometry (GC-MS). Volatile compound concentrations were then subjected to statistical analysis. No significant differences in pH, titratable and volatile acidity, and ethanol and lactic acid were observed among the four wines analyzed. However, the musts supplemented with the ONC slightly increased the fermentation rate of the flocculant yeast, and, also, this additive reduced the volume of lees in the spontaneous fermentation and flocculant yeast by 1.2% and 0.6%, respectively. The concentrations of 11 major and 28 minor volatiles were significantly influenced (p-value ≤ 0.05) by the inoculation strategy, while 8 major and 28 minor volatiles were affected by ONC supplementation. This supplementation significantly decreased the Odor Activity Values and, consequently, the values of the odorant series established in wines from spontaneous fermentation. On the contrary, those from flocculant yeast showed a significant increase in all the odorant series except for the waxy series, leading to a more balanced aroma profile. Additionally, lower scores were recorded for the green, creamy, citrus, chemical, and honey series compared to wines from spontaneous fermentation. Overall, the commercial ONC extract contributed to a content increase in volatiles that provided desirable aromatic notes to the wines made with flocculant yeast, although the organoleptic evaluation showed no significant statistical differences in the attributes evaluated at the 95% confidence level. Full article

The community of epibiotic yeasts significantly influences the quality of Docynia delavayi (Franch.) Schneid. wine. The yeast diversity in four different Docynia delavayi (Franch.) Schneid. wines during the brewing stage was investigated using pure culture methods and high-throughput sequencing technology. A total of 229,381,292 sequencing bases were generated, yielding 323,820 valid sequences with an average length of 708 nt and identifying 93 operational taxonomic units (OTUs) from naturally fermented samples of Docynia delavayi (Franch.) Schneid. wine for classification purposes. At the early fermentation stage, Hanseniaspora sp. was identified as the dominant species, whereas at the late fermentation stage, Hanseniaspora sp., Saccharomyces sp., and Candida californica became predominant. From these samples, a total of 109 yeast strains were isolated from Docynia delavayi (Franch.) Schneid. wine. Three specific strains—LZX-76, LZX-89, and LZX-104—were further selected based on their growth characteristics along with hydrogen sulfide production, ester production, ethanol production, and tolerance levels. Through morphological examination and molecular biology techniques, these strains were identified as Pichia fermentans and Hanseniaspora spp. Additionally, a total of 29 volatile compounds were detected through simulated fermentation processes; these included 12 esters, 6 alcohols, 2 acids, 4 aldehydes, and 5 other compounds. When compared to commercial yeasts used as starters in winemaking processes, it was observed that utilizing yeast strains LZX-76, LZX-89, and LZX-104 resulted in an increased number of volatile compounds, which enhanced the aromatic profile characteristics of Docynia delavayi (Franch.) Schneid. wine by making its aroma richer and more complex. The findings from this study hold significant potential value for both the production practices and research endeavors related to Docynia delavayi (Franch.) Schneid. wine. Full article

The yeast Saccharomyces cerevisiae ensures successful fermentation in winemaking, although the persistent use of commercial strains lead to the loss of aroma complexity of wines. Hence, the research of indigenous S. cerevisiae with proper oenological features and well adapted to specific wine-growing areas become of great interest for winemakers. Here, 206 pure cultures of S. cerevisiae were isolated from two wineries during a two-year sampling campaign and bio-typed through interdelta sequences analyses with the aim to evaluate the occurrence and persistence of the S. cerevisiae wild population linked to each winery. Both wineries belong to the same Verdicchio DOC wine area (Castelli di Jesi), and never used commercial yeasts during fermentation. Results showed 19 different biotypes with a specific population of S. cerevisiae in each winery, without cross-contamination with each other and with commercial starter strains. Moreover, inside each winery a persistence of some dominant biotypes was observed over time (three biotypes in winery 1; 95% of isolates in the two years and one biotype in winery 2; 20% of isolates in the two years), indicating a sort of “winery-effect”. The evaluation of S. cerevisiae populations for the oenological characters by microfermentations showed a proper and well distinct aromatic imprinting on the resulted wines supporting the concept of “winery effect”. Full article

In view of climate change and the increasingly antagonistic wine market, the exploitation of native genetic resources is revisited in relation to sustainable wine production. ‘Sideritis’ is a late-ripening Greek grape variety, which is quite promising for counteracting wine quality issues associated with the annual temperature rise. The aim of this study was to improve the quality and to enhance the aroma of ‘Sideritis’ wine through the use of native yeasts. To improve vinification, Hanseniaspora opuntiae L1 was used along with Saccharomyces cerevisiae W7 in mixed fermentations (SQ). The addition of H. οpuntiae significantly altered the chemical profile of the wine compared to the single-inoculated fermentations with W7 (IS). H. opuntiae increased all the acetate esters, except for hexyl acetate and (Z)-3-hexen-1-ol acetate. The concentration of 2-phenylethyl acetate, which imparts flowery and sweet notes, exhibited a 2.6-fold increase in SQ as compared to IS wines. SQ also showed higher levels in several ethyl esters, including ethyl butyrate, ethyl heptanoate and ethyl 7-octenoate, which are associated with fruity notes compared to IS. H. opuntiae produced citronellol, a terpene associated with rose and green notes, and increased the overall acceptance of the wine. Present results are thus quite promising for improving ‘Sideritis’ wine quality towards a sustainable wine production in Greece in view of global warming. Full article

Grape variety, quality, geographic origins and phytopathology can influence the amount of polyphenols that accumulate in grape tissues. Polyphenols in wine not only shape their organoleptic characteristics but also significantly contribute to the positive impact that this beverage has on human health. However, during the winemaking process, the total polyphenol content is substantially reduced due to the adsorption onto yeast wall polymers and subsequent lees separation. Despite this, limited information is available regarding the influence of the yeast starter strain on the polyphenolic profile of wine. To address this issue, a population consisting of 136 Saccharomyces cerevisiae strains was analyzed to identify those with a diminished ability to adsorb polyphenols. Firstly, the reduction in concentration of polyphenolic compounds associated to each strain was studied by assaying Total Phenolic Content (TPC) and Trolox Equivalent Antioxidant Capacity (TEAC) in the wines produced by micro-scale must fermentation. A total of 29 strains exhibiting a TPC and TEAC reduction ≤ 50%, when compared to that detected in the utilized grape must were identified and the nine most-promising strains were further validated by larger-scale vinification. Physico-chemical analyses of the resulting wines led to the identification of four strains, namely ITEM6920, ITEM9500, ITEM9507 and ITEM9508 which showed, compared to the control wine, a TPC and TEAC reduction ≤ 20 in the produced wines. They were denoted by a significant (p < 0.05) increased amount of anthocyanin, quercetin and trans-coutaric acid, minimal volatile acidity (<0.2 g/L), absence of undesirable metabolites and a well-balanced volatile profile. As far as we know, this investigation represents the first clonal selection of yeast strains aimed at the identifying “functional” fermentation starters, thereby enabling the production of regional wines with enriched polyphenolic content. Full article

Malolactic fermentation (MLF) in wine is driven by the lactic acid bacterium Oenococcus oeni in most cases. Although this bacterium is resistant to wine stress conditions, it often faces difficulties completing MLF. Previous studies indicate that yeast mannoproteins may improve O. oeni growth and survival in wine. However, very little is known about this topic. This study evaluated the effect of the addition of mannoprotein extracts to culture media on O. oeni growth and its survival to stress conditions and MLF performance. Three commercial mannoprotein extracts were characterized in terms of polysaccharide and protein richness and were used for O. oeni culture media supplementation. The addition of mannoprotein extracts improved the survival of the two evaluated O. oeni strains, PSU-1 and VP41, after acid shock (pH 3.2) in comparison to that of the control. The transcriptional response of four genes involved in mannose metabolism was different depending on the strain, indicating the complexity of sugar metabolism in O. oeni. PSU-1 cells grown with two of the mannoprotein extracts performed faster MLF compared with the control condition, indicating that mannoprotein addition may improve the performance of O. oeni starter cultures, although this effect depends on the strain. Full article

This work focuses on the establishment of analytical and sensory differences between young wines obtained from the same grape must fermented using different strategies. The main purpose is to provide winemakers with objective criteria to help them to choose the best fermentation method to obtain the desired wine characteristics. The effects of four strategies were tested: a spontaneous fermentation with wild yeasts (WYs) and the addition of starter cultures of Lachancea thermotolerans (LT), Metschnikowia pulcherrima (MP), and a Saccharomyces cerevisiae strain that is an overproducer of glutathione (SC) in different batches of the same must of the Pedro Ximénez white grape. The analytical results obtained show as the LT wine has the highest glutathione content, while the lowest ethanol concentrations. The use of chemometric tools applied to the volatile compounds allowed to differentiate the four wines by a principal component analysis (PCA) and the identification of 27 key compounds. The four wines did not show statistical differences in their smell or taste attributes and only the LT wine was visually differentiated from the rest. Full article

The use of multi-starters in oenological conditions (Saccharomyces cerevisiae and non-Saccharomyces species) is becoming increasingly common. For the past ten years, the combination of Torulaspora delbrueckii and S. cerevisiae has been proposed to winemakers to improve the wine aromatic profile compared to pure inoculation with Saccharomyces cerevisiae. In this work, two commercial strains, T. delbrueckii Zymaflore^® Alpha and S. cerevisiae Zymaflore^® X5 (Laffort compagny, Floirac, France), were investigated in Sauvignon blanc must using a fermentor with a double compartment allowing for physical separation of the two yeast species. The physical separation of the two species resulted in significant differences in the growth, fermentation kinetics (maximum fermentation rate (+13%)), fermentation duration (−14%) and the production of 3SH (+35%) in comparison to mixed cultures with contact. Proteomic analysis confirmed cell–cell contact interactions, as strong differences were observed for both species between mixed cultures with and without physical contact. T. delbrueckii mortality in mixed cultures with physical contact may be explained by an oxidative stress. Indeed two proteins implicated in the oxidative stress response were found in significantly higher amounts: a cytosolic catalase T and a cytoplasmic thioredoxin isoenzyme. For S. cerevisiae, an increase in proteins involved in the respiratory chain and proton transport were found in higher amounts in pure cultures and mixed culture without physical contact. Our results confirmed that the two mixed inoculations increased certain minor esters (ethylpropanoate, ethyl dihydrocinnamate and ethyl isobutanoate) specifically produced by T. delbrueckii, 3.4-fold more compared to in the pure S. cerevisiae culture. In conclusion, these results provide new insights into the underlying mechanisms involved in cell–cell contact and confirm the benefits of using T. delbrueckii species under winemaking conditions. Full article

Exploitation of the biodiversity of native wine yeast is a means of modifying the sensory characteristics of wine. Samples from different regions in China were analysed to screen native isolates as potential starter cultures. Through morphological and molecular biological analyses, we found six species, belonging to four genera (Hanseniaspora, Saccharomyces, Rhodotorula and Metschnikowia). These species were subjected to stress tolerance assays (ethanol, glucose, SO₂ and pH), enzymatic activity tests (sulphite reductase activity, β-glucosidase activity and protease activity) and fermentation tests. Saccharomyces cerevisiae showed a high tolerance to ethanol and completed fermentation independently. Hanseniaspora demonstrated good enzymatic activity and completed sequential fermentation. The fermentation experiment showed that the PCT4 strain had the best aroma complexity. This study provides a reference for selecting new starters from the perspective of flavour enzymes and tolerance and diversifying the sensory quality of wines from the region. Full article

For oenological products, most of the intrinsic and extrinsic drivers of perceived quality are associated with specific aromatic profiles. Aromatic diversity has been recognized as a central element in perceived quality as it is able to transmit the complex interactions between grape variety, geographical characteristics, and viticultural and winemaking practices, including the fermentative process. A comprehensive characterization of flavour compounds by headspace solid-phase microextraction (HS-SPME) and gas chromatography coupled to mass spectrometric analysis is often needed in order to ascertain the quality of wine. HS-SPME requires a proper optimization that can be achieved through an adequate experimental design. Here, a HS-SPME/GC-MS based method was developed to investigate the volatile compounds of wine samples obtained by laboratory-scale fermentations. This was performed by inoculating a commercial Saccharomyces cerevisiae strain, which is used both as single starter and as mixed starter, with an indigenous Hanseniaspora osmophila strain. The experimental conditions of HS-SPME (extraction temperature and time) were optimized by applying a face-centred composite experimental design. Up to 95% of the total variance was explained by the proposed model. The optimized method allowed us to confirm the usefulness of combining the inoculation of grapes with selected yeast strains in co-culture situations in order to improve the wine bouquet. Full article