Search Results (74)

This study provides a comprehensive analysis of the microbial dynamics involved in the fermentation process of traditional Musalais wine, an intangible cultural heritage of Xinjiang. Utilizing metagenomic sequencing, we identified 2894 microbial species, of which 494 persisted throughout the fermentation process. Saccharomyces cerevisiae was the dominant species, with its prevalence increasing from 97.35% in the early phase to 99.38% in the mid phase, before slightly decreasing to 98.79% in the late phase. Additionally, 24 non-Saccharomyces yeast species, including Hanseniaspora uvarum, Lachancea thermotolerans, and Torulaspora delbrueckii, were detected. Common species associated with other fermented foods, including Wickerhamomyces anomalus, Kluyveromyces marxianus, Saccharomyces eubayanus, and Zygosaccharomyces parabailii, were also identified. Notably, species not previously used in food fermentation, such as Saccharomyces jurei, Sodiomyces alkalinus, Vanrija pseudolonga, and Moesziomyces antarcticus, were also identified in this study. Furthermore, the Kyoto Encyclopedia of Genes and Genomes (KO) and Gene Ontology (GO) revealed notable variations in metabolic pathways and enriched functional genes. In addition, a total of 82 volatile compounds were detected in the final product, with higher alcohols (60.12%), esters (37.80%), and organic acids (1.80%) being the most prevalent. These results offer important insights into microbial interactions and their influence on Musalais wine quality, laying the groundwork for optimizing the fermentation process. Full article

Winemaking involves a microbial ecosystem where yeast diversity, shaped by terroir and winemaking conditions, determines wine characteristics. Understanding the microbial diversity of vineyards and spontaneous fermentation is crucial for explaining a winery’s typical wine profile. Studying and inoculating indigenous strains make it possible to produce high quality wines, reflecting the production environment. This study analyzes the yeast species involved in 16 spontaneous fermentations (8 in 2022 and 8 in 2023) from grapes of four distinct vineyards under two sets of winemaking conditions. A total of 1100 yeast colonies were identified by MALDI-TOF and DNA sequencing techniques. Saccharomyces (S.) cerevisiae and Hanseniaspora uvarum were the most prevalent species, alongside significant populations of non-Saccharomyces yeasts such as Lachancea thermotolerans and Metchnikowia pulcherrima, which were the most abundant ones. Minor yeast species, including Aureobasidium pullulans, Starmerella bacillaris, Kazachstania servazzi, and other Hanseniaspora spp., were also detected. The results demonstrated that yeast diversity in spontaneous fermentations varied according to vineyard origin and winemaking conditions. Differences between the two vintages studied indicated that annual climatic conditions significantly influenced yeast diversity, especially among non-Saccharomyces species. This substantial diversity represents a valuable source of indigenous yeasts for preserving the typicity of a winery’s wines under controlled conditions. Full article

Yeasts isolated from fermented plant-based sources—boza, pickles, and chickpeas—were evaluated for probiotic potential. Among 18 colonies, seven isolates showing no hemolytic activity were selected and identified: Pichia kudriavzevii (four isolates), Kazachstania exigua, Hanseniaspora uvarum, and Saccharomyces cerevisiae. Most isolates were able to survive under low pH and bile salt conditions, simulating the environment of the gastrointestinal tract (5.62–8.59 log CFU/mL) and tolerated NaCl concentrations up to 6–8% (w/v). All isolates exhibited antibiotic resistance; however, their susceptibility to antifungals varied. While P. kudriavzevii DD_B_M88 exhibited the highest hydrophobicity (63.07%), isolate auto-aggregation increased to 53–95% after 24 h. Most isolates showed a great capability to co-aggregate with pathogens and inhibited them significantly (up to 98.72%), except for S. cerevisiae DD_NB_M90. The selected three isolates and their cell-free supernatants, up to a certain concentration, showed no significant cytotoxicity on Caco-2 cell line. Eventually, six isolates, excluding S. cerevisiae DD_NB_M90, fulfilled the probiotic criteria, and can serve as probiotic starter cultures for alternative food production. Full article

Bioprotection in winemaking refers to the use of naturally occurring microorganisms—mainly non-Saccharomyces yeasts—to inhibit the growth of spoilage microbes and reduce the need for chemical preservatives like sulfur dioxide (SO₂). Numerous studies have demonstrated the benefits of non-Saccharomyces as bioprotectants. However, the use of Saccharomyces cerevisiae as a bioprotectant has been studied very little. Furthermore, it can offer many advantages for the production of sulfite-free wines. To test if S. cerevisiae could be used in bioprotection, we compared the ability of different strains to inhibit the growth of Brettanomyces bruxellensis and Hanseniaspora uvarum. Among the strains tested, the S. cerevisiae Sc54 strain isolated from the vineyard of the Bekaa plain was selected. To investigate its mechanisms of action, we analyzed its metabolite production, including acetic acid and ethanol. Taking into account the low levels of these metabolites and the lack of similar inhibition patterns in media supplemented with acetic acid and ethanol, it appears that other factors contribute to its antagonistic properties. Nutrient competition was ruled out as a factor, as the growth inhibition of B. bruxellensis and H. uvarum occurred rapidly within the first 24 h of co-culture. In this study, we explored the role of the S. cerevisiae killer toxin (Sc54Kt) as a bioprotective agent against H. uvarum and B. bruxellensis spoilage yeasts. Purification procedures with ethanol allowed the extraction of Sc54Kt, yielding two concentrations (0.185 and 0.5 mg/mL). Remarkably, semi-purified Sc54Kt exhibited inhibitory effects at both concentrations under winemaking conditions, effectively controlling the growth and metabolic activity of the target spoilage yeasts. Overall, these findings demonstrate that S. cerevisiae Sc54 not only exerts a strong bioprotective effect but also contributes to improving the quality of wine. The results suggest that S. cerevisiae Sc54 is a promising bioprotective agent for mitigating spoilage yeasts in winemaking, offering a natural and effective alternative to conventional antimicrobial strategies. 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

The production of strawberry wine is an effective strategy for addressing the significant economic losses caused by strawberry spoilage. In recent years, there has been an increase in consumer demand for quality and flavor diversity in fruit wines. Therefore, it is necessary to develop novel strawberry wine products. In this research, we assessed and analyzed the influences of fermentation with Hanseniaspora uvarum, Kurtzmaniella quercitrusa, and Saccharomyces cerevisiae under four fermentation conditions on the fermentation kinetics, organoleptic characteristics, chemical compositions, antioxidant capacities, and flavor profiles of strawberry wines. Strawberry wines fermented with the indigenous non-Saccharomyces yeasts H. uvarum and K. quercitrusa showed higher 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-Azinobis-(3-ethylbenzothiazoline-6-sulfonate) (ABTS) free-radical-scavenging capacities and significantly different flavor profiles compared to strawberry wines fermented with S. cerevisiae. In addition, adjusting the initial soluble solids contents of strawberry juices and fermentation temperatures positively affected the quality and flavor profiles of strawberry wines fermented with the H. uvarum and K. quercitrusa strains. Under the condition of 18 °C–20 °Brix, strawberry wine fermented with K. quercitrusa presented the highest antioxidant capacity, with enhanced flavor diversity and color intensity. It is worth noting that K. quercitrusa can be an alternative yeast for producing high-quality strawberry wine with a distinct floral aroma. Full article

It is estimated that approximately 25% of waste remains after the apple juice pressing process. Combining this waste biomass with valuable compounds creates the potential for reuse. To create a cost-efficient ecological process without any expensive steps, the aim of this research was to examine the potential of using non-Saccharomyces yeasts (Kazachstania barnettii D1, Hanseniaspora uvarum D9, and Wickerhamomyces anomalus D11) for the low-temperature valorisation of apple pomace. The scope encompassed characteristics of apple pomace and the evaluation of yeast growth and metabolic activity, including carbohydrate consumption, enzymatic activity, and the biosynthesis of volatile organic compounds. Moreover, the effect of inoculum size on biomass increases and the productivity of metabolites during the fermentation of apple pomace were evaluated. To investigate the potential intensification of the process, the experiment was performed on hydrolysed and untreated apple pomace. The obtained results indicate that yeast growth was satisfactory regardless of the inoculum size in both fermentation media. Various activities of peptidases, esterases, phosphatases, and glucosidases were observed. The yeast isolates presented metabolic activity during the process which was confirmed by the production of ethanol and acetic acid. Moreover, a significant amount of volatile organic compounds, especially esters, were synthesised, which have a positive impact on the sensory profile of fermented apple pomace. In general, the hydrolysis of apple pomace did not result in better yeast activity and the formation of aroma compounds. Full article

Yeasts produce numerous antimicrobial agents such as killer toxins, volatile organic compounds (VOCs), and other secondary metabolites, establishing themselves in developing natural and sustainable biocontrol strategies for agriculture and food preservation. This study addressed the biocontrol potential of yeasts, isolated from spontaneous fermentations of rosehips (Rosa canina L.) and rowanberries (Sorbus aucuparia L.), focusing on their killer phenotypes and VOCs production. Yeasts were isolated using spontaneous fermentations with Hanseniaspora uvarum and Metschnikowia pulcherrima identified as the dominant species, comprising approximately 70% of the yeast population. Among 163 isolated strains, 20% demonstrated killing activity, with Saccharomyces cerevisiae exhibiting the strongest killing efficiency, as well as Pichia anomala and M. pulcherrima showing broad-spectrum antagonistic activity. This study identified dsRNA-encoded killer phenotypes in S. cerevisiae, S. paradoxus, and Torulaspora delbrueckii, revealing multiple distinct killer toxin types. The biocontrol potential of wild berry-inhabiting yeasts was demonstrated in a real food system, grape juice, where the S. cerevisiae K2-type killer strain significantly reduced fungal contaminants. The selected H. uvarum, M. pulcherrima, S. cerevisiae, and S. paradoxus yeast strains representing both berries were applied for VOC analysis and identification by gas chromatography-linked mass spectrometry. It was revealed that the patterns of emitted volatiles are yeast species-specific. Statistically significant differences between the individual VOCs were observed among killing phenotype-possessing vs. non-killer S. paradoxus yeasts, thus revealing the involvement of killer systems in multi-level biocontrol enablement. The performed studies deepen our understanding of potential yeast biocontrol mechanisms, highlight the importance of produced antimicrobials and volatiles in ensuring antagonistic efficacy, and prove the relevance of isolated biocontrol yeasts for improving food safety. Full article

This study is the first to investigate the impact of indigenous non-Saccharomyces yeasts, including Hypopichia pseudoburtonii, Metschnikowia sinensis/shanxiensis, Metschnikowia chrysoperlae, Metschnikowia pulcherrima, Lachancea thermotolerans, Hanseniaspora uvarum, Hanseniaspora guilliermondii, Hanseniaspora pseudoguilliermondii, Pichia kluyveri, and Starmerella apicola on the lipid composition of sterile Maraština grape juice and wines using the UHPLC-MS/MS method. Yeasts were tested in monoculture and sequential fermentations alongside commercial Saccharomyces cerevisiae. Indigenous non-Saccharomyces yeasts showed the potential to improve fermentation performance and enable the development of new wine styles through the biosynthesis of an unsaturated fatty acid pathway, which was identified as the most significant pathway. In monoculture fermentations, L. thermotolerans, H. uvarum, H. guilliermondii, H. pseudoguilliermondii, and P. kluyveri significantly reduced lignoceric acid, potentially influencing wine aroma through the formation of esters and higher alcohols. Hyp. pseudoburtonii, M. chrysoperlae, M. pulcherrima, P. kluyveri, and S. apicola increased the demand for lipids, such as stearic acid, which may help preserve membrane permeability by integrating into the membrane in response to ethanol shock. The most significant impact on free fatty esters was observed in fermentations with H. pseudoguilliermondii. Furthermore, L. thermotolerans in sequential fermentations significantly reduced arachidic, stearic, and palmitic acid. P. kluyveri reduced the content of erucic and linoleic acid. Full article

The aim of the present study was to evaluate the use of different chemical treatments of apple pomace in order to produce an economical culture medium for the propagation of two non-conventional yeast strains. An experimental design approach was used for the optimization of the hydrolysis conditions of apple pomace. Both acid and alkaline treatment conditions were tested. The optimal hydrolysis conditions to disrupt the pomace lignocellulosic structure were 1% w/v of H₃PO₄, 121 °C, and 40 min for acid treatment, while 6% w/v of NH₄OH, 20 °C, and 2 h were optimal for the alkaline condition. Saccharomyces uvarum NPCC 1420 and Saccharomyces eubayanus NPCC 1292 yeasts were able to grow in the liquid fraction obtained from both acid and alkaline treatments. However, the medium with the acid treatment was found to be more suitable for yeast growth, showing, for both strains, higher µmax and biomass production and lower td and λ than that observed for the medium with the alkaline treatment. According to the growth parameter analysis for both strains, the acid treatment was selected for further studies. By taking advantage of this agroindustrial by-product, a circular economy approach is promoted, reducing environmental impact and fostering sustainable development. Full article

Diacetyl is a flavor compound in wine with a low threshold (1–2 mg/L). It is produced during alcoholic fermentation (AF) Saccharomyces and malolactic fermentation (MLF) initiated by lactic acid bacteria (LAB). Whereas, the environment after AF suppresses the normal metabolism of LAB after AF. Researchs have shown the influence on diacetyl mechanisms of the genes ILV2, ILV6, ILV3, ILV5, BDH1, BDH2, and gene aldB from Lactobacillus plantarum in Saccharomyce uvarum WY1. While we found that the diacetyl contents produced by mutants after MLF (Co-fermentation and Seq-fermentation) were significantly improved compared to AF alone. Moreover, the genes mae1 and mae2 from S. pombe, and gene mleS from L. lactis exhibited significant effect on deacidification in our previous study, but the diacetyl of the mutants showed obvious improvement in this study. Thus the effects of association mutation of genes (ILV2, ILV6, ILV3, ILV5, BDH1, BDH2, aldB, mae1, and mleS) on deacidification, diacetyl and other flavors (organic acids, higher alcohols and esters) metabolism in S. uvarum after AF were detected in the study. Among all the mutants, strains V6AmS, V635mS, and V6B12mS showed the most favorable results. Specifically, the L-malic acid contents decreased to 1.26 g/L, 1.18 g/L, and 1.19 g/L, respectively. Concurrently, diacetyl levels were reduced by 52.56%, 61.84%, and 65.31%. The production of n-propanol increased by 18.84%, 20.89%, and 28.12%, whereas isobutanol levels decreased by 37.01%, 42.36%, and 44.04%, and isoamyl alcohol levels decreased by 19.28%, 19.79%, and 16.74%, compared to the parental strain WY1. Additionally, the concentration of lactate ester in the wine increased to 13.162 mg/L, 14.729 mg/L, and 14.236 mg/L, respectively. Full article

This study aimed to provide novel information on the impact of indigenous non-Saccharomyces yeasts, including Metschnikowia chrysoperlae, Metschnikowia sinensis/shanxiensis, Metschnikowia pulcherrima, Lachancea thermotolerans, Hanseniaspora uvarum, Hanseniaspora guilliermondii, and Pichia kluyveri, on metabolites related to the metabolism of tryptophan, phenylalanine, and tyrosine. The experiment included two fermentation practices: monoculture and sequential fermentation with commercial Saccharomyces cerevisiae, using sterile Maraština grape juice. A targeted approach through ultrahigh-resolution liquid chromatography associated with mass spectrometry was used to quantify 38 metabolites. All the indigenous yeasts demonstrated better consumption of tryptophan in monoculture than in interaction with S. cerevisiae. M. sinensis/shanxiensis was the only producer of indole-3-carboxylic acid, while its ethyl ester was detected in monoculture fermentation with H. guilliermondii. H. guilliermondii consumed the most phenylalanine among the other isolates. 5-hydroxy-L-tryptophan was detected in fermentations with M. pulcherrima and M. sinensis/shanxiensis. M. pulcherrima significantly increased tryptophol content and utilised tyrosine in monoculture fermentations. Sequential fermentation with M. sinensis/shanxiensis and S. cerevisiae produced higher amounts of N-acetyl derivatives of tryptophan and phenylalanine, while H. guilliermondii-S. cerevisiae fermentation resulted in wines with the highest concentrations of L-kynurenine and 3-hydroxyanthranilic acid. P. kluyveri produced the highest concentration of N-acetyl-L-tyrosine in monoculture fermentations. These findings highlight the different yeast metabolic pathways. Full article

Despite the current appreciation of pulque as a probiotic fermented beverage, pulque has been also regarded as a poor-quality product, particularly due to the lack of sanitary control during its elaboration. To address this problem, a semi-continuous fermentation system was established, emulating the artisanal production process. Microfiltration-sterilized aguamiel was employed as the substrate, whereas a good-quality pulque was used as the fermentation inoculum. During the fermentation, the physicochemical, microbiological (lactic acid and Leuconostoc-type bacteria and yeasts) and sensory characteristics of the must were monitored. The isolated microorganisms were identified by molecular biology and MALDI-MS techniques. The sterilization of aguamiel by microfiltration did not negatively affect its physicochemical attributes. After 6–8 days of operation of the semi-continuous bioreactor, the fermentation reached a quasi-stationary state considering most of the parameters monitored during the experiment. The final fermentation product presented similar physicochemical, microbial and sensory properties to those of the pulque inoculum. The genera identified were Leuconostoc, Lentilactobacillus, Lactobacillus, Liquorilactobacillus, Fructilactobacillus and Saccharomyces. The strains Lentilactobacillus diolivorans and Liquorilactobacillus capillatus and uvarum have not been previously isolated from pulque. In conclusion, the fermentation system developed in this work was effective to standardize the quality of pulque while preserving the positive attributes of the artisanal process, thus harnessing the probiotic properties of pulque. Full article

During malolactic fermentation (MLF) of vinification, the harsh L–malic acid undergoes transformation into the milder L–lactic acid, and via decarboxylation reactions it is catalyzed by malolactic enzymes in LAB. The use of bacterial malolactic starter cultures, which usually present challenges in the industry as the suboptimal conditions after alcoholic fermentation (AF), including nutrient limitations, low temperatures, acidic pH levels, elevated alcohol, and sulfur dioxide concentrations after AF, lead to “stuck” or “sluggish” MLF and spoilage of wines. Saccharomyces uvarum has interesting oenological properties and provides a stronger aromatic intensity than Saccharomyces cerevisiae in AF. In the study, the biological pathways of deacidification were constructed in S. uvarum, which made the S. uvarum carry out the AF and MLF simultaneously, as different genes encoding malolactic enzyme (mleS or mleA), malic enzyme (MAE2), and malate permease (melP or MAE1) from Schizosaccharomyces pombe, Lactococcus lactis, Oenococcus oeni, and Lactobacillus plantarum were heterologously expressed. For further inquiry, the effect of L–malic acid metabolism on the flavor balance in wine, the related flavor substances, higher alcohols, and esters production, were detected. Of all the recombinants, the strains WYm1S_N with coexpression of malate permease gene MAE1 from S. pombe and malolactic enzyme gene mleS from L. lactis and WYm1m2 with coexpression of gene MAE1 and malate permease gene MAE2 from S. pombe could reduce the L–malic acid contents to about 1 g/L, and in which the mutant WYm1S_N exhibited the best effect on the flavor quality improvement. Full article

This study investigates innovative approaches to improve the quality and aroma characteristics of Muscat Hamburg wine production by substituting the conventional Saccharomyces cerevisiae yeast with an efficient fermentation strain of Schizosaccharomyces pombe. The typical use of S. cerevisiae in Muscat Hamburg wine often leads to uniformity and prolonged processing times, requiring subsequent malolactic fermentation to degrade excessive malic acid. The study advocates for the replacement of S. cerevisiae with a specific S. pombe strain, Sp-410, isolated from the fermented grains of sauce-flavor Baijiu, a Chinese spirit. Muscat Hamburg wine fermented with the S. pombe strain demonstrates decreased malic acid levels, offering a potential alternative to malolactic fermentation. However, exclusive S. pombe fermentation may result in an overproduction of acetic acid metabolites, leading to a monotonous taste. In response, the study proposes a mixed fermentation approach, combining the S. pombe strain with a Saccharomyces uvarum strain and a non-Saccharomyces yeast, Torulaspora delbrueckii. The optimized mixed fermentation strategies (M:SP+TD and M60SP+TD) involve specific proportions and intervals of inoculation, aiming to enhance the quality and aroma complexity of Muscat Hamburg wine. In conclusion, this research contributes to advancing the production of high-quality Muscat Hamburg wines, utilizing S. pombe as the primary yeast strain and implementing mixed fermentation methodologies. Full article