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Grapes have been cultivated for wine production for millennia. Wine production involves a complex biochemical process where sugars in grapes must be converted into alcohol and other compounds by microbial fermentation, primarily by the yeast Saccharomyces cerevisiae. Commercially available S. cerevisiae strains are often used in winemaking, but indigenous (native) strains are gaining attention for their potential to contribute unique flavors. Recent advancements in high-throughput DNA sequencing have revolutionized our understanding of microbial communities during wine fermentation. Indeed, transcriptomic analysis of S. cerevisiae during wine fermentation has revealed a core gene expression program and provided insights into how this yeast adapts to fermentation conditions. Here, we assessed how the age of vines impacts the grape fungal microbiome and used transcriptomics to characterize microbial functions in grape must be fermented with commercial and native S. cerevisiae. We discovered that ~130-year-old Zinfandel vines harbor higher fungal loads on their grapes compared to 20-year-old Zinfandel vines, but fungal diversity is similar. Additionally, a comparison of inoculated and uninoculated fermentations showed distinct fungal dynamics, with uninoculated fermentations harboring the yeasts Metschnikowia and Pichia. Transcriptomic analysis revealed significant differences in gene expression between fermentations inoculated and not inoculated with a commercial S. cerevisiae strain. Genes related to metabolism, stress response, and cell adhesion were differentially expressed, indicating varied functionality of S. cerevisiae in these fermentations. These findings provide insights into S. cerevisiae function during fermentation and highlight the potential for indigenous yeast to contribute to wine diversity. IMPORTANCE: Understanding microbial functions during wine fermentation, particularly the role of Saccharomyces cerevisiae, is crucial for enhancing wine quality. While commercially available S. cerevisiae strains are commonly used, indigenous strains can offer unique flavors, potentially reflecting vineyard terroir. By leveraging high-throughput DNA sequencing and transcriptomic analysis, we explored the impact of vine age on the grape mycobiome and characterized microbial functions during grape fermentation. Our findings revealed that older vines harbor higher fungal loads, but fungal diversity remains similar across vine ages. Additionally, uninoculated fermentations exhibited diverse fungal dynamics, including the beneficial wine yeasts Metschnikowia and Pichia. Transcriptomic analysis uncovered significant differences in S. cerevisiae gene expression between inoculated and uninoculated fermentations, highlighting the potential of indigenous yeast to enhance wine diversity and inform winemaking practices.

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BACKGROUND: Sophorolipids are glycolipid biosurfactants with potential antibacterial, antifungal, and anticancer applications, rendering them promising for research. Therefore, this study hypothesizes that sophorolipids may have a notable impact on disrupting membrane integrity and triggering the production of reactive oxygen species, ultimately resulting in the eradication of pathogenic microbes. RESULTS: The current study resulted in the isolation of two Metschnikowia novel yeast strains. Sophorolipids production from these strains reached maximum yields of 23.24 g/l and 21.75 g/l, respectively, at the bioreactors level. Biosurfactants sophorolipids were characterized using FTIR and LC-MS techniques and found to be a mixture of acidic and lactonic forms with molecular weights of m/z 678 and 700. Our research elucidated sophorolipids' mechanism in disrupting bacterial and fungal membranes through ROS generation, confirmed by transmission electron microscopy and FACS analysis. The results showed that these compounds disrupted the membrane integrity and induced ROS production, leading to cell death in Klebsiella pneumoniae and Fusarium solani. In addition, the anticancer properties of sophorolipids were investigated on the A549 lung cancer cell line and found that sophorolipid-11D (SL-11D) and sophorolipid-11X (SL-11X) disrupted the actin cytoskeleton, as evidenced by immunofluorescence microscopy. The A549 cells were stained with Acridine orange/Ethidium bromide, which showed that they underwent necrosis. This was confirmed by flow cytometric analysis using Annexin/PI staining. The SL-11D and SL-11X molecules exhibited low levels of haemolytic activity and in-vitro cytotoxicity in HEK293, Caco-2, and L929 cell lines. CONCLUSION: In this work, novel yeast species CIG-11DT and CIG-11XT, isolated from the bee's gut, produce significant yields of sophorolipids without needing secondary oil sources, indicating a more economical production method. Our research shows that sophorolipids disrupt bacterial and fungal membranes via ROS production. They suggest they may act as chemo-preventive agents by inducing apoptosis in lung cancer cells, offering the potential for enhancing anticancer therapies.

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BACKGROUND: To date, cider production has primarily relied on Saccharomyces cerevisiae. Introducing novel non-Saccharomyces yeasts can enhance the diversity of cider properties. Among these, the Metschnikowia genus stands out for its ability to produce hydrolytic enzymes that may impact the sensorial and technological properties of cider. This study focused on evaluating the impact of three Metschnikowia species - Metschnikowia koreensis (Mk), M. reukaufii (Mr), and M. pulcherrima (Mp) - which exhibit acid protease and esterase activity, on the quality enhancement of cider. RESULTS: The research findings indicate that the overall quality of cider produced through co-fermentation with these species surpassed that of cider fermented with mono-fermentation of S. cerevisiae (Sc). The cider fermented with the Sc + Mk combination exhibited the lowest levels of harsh-tasting malic acid and higher levels of softer lactic acid. Sensory array analysis also demonstrated that the Sc + Mk fermented cider exhibited high sensor response values for compounds contributing to a complex overall olfactory composition and richness. Furthermore, the Sc + Mk fermented cider exhibited the highest total quantity and variety of volatile organic compounds (VOCs). Specifically, the concentrations of phenethyl alcohol, 3-methyl-1-butanol, ethyl octanoate, and decanoic acid were notably elevated in comparison with other groups. CONCLUSION: This study illustrates that Metschnikowia species, particularly M. koreensis, show significant potential as starters for cider due to their various technological properties, including acidity modulation, aroma enhancement, and color improvement. The findings of this study provide a foundation for improving cider quality by co-fermenting S. cerevisiae with innovative starter cultures. © 2024 Society of Chemical Industry.

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Cold maceration (CM) is widely applied in winemaking to improve wine aroma and overall quality. However, more efficient CM techniques for industrial-scale winemaking are still needed. This study examined the impact of CM with indigenous cryotolerant Metschnikowia pulcherrima Mp0520 (Mp-CM) on the Muscat wine aromatic characteristics. The results demonstrated a significant divergence in the types and concentrations of aroma compounds between Mp-CM wine and the control. The Mp-CM wine exhibited a significantly higher terpenes content, resulting in a Muscat wine characterized by terpenes, compared to the control predominated by esters. Additionally, the Mp-CM wine demonstrated elevated levels of α-terpineol and terpinolene, potentially enhancing the varietal aroma stability of Muscat wine. Furthermore, Mp-CM gave Muscat wine a heightened fruity aroma and a more complex aroma. These findings suggested that the Mp-CM utilized in this study offered promising avenues for enhancing the variety aroma characteristics of Muscat wine on large scale winemaking.

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Gluconobacter oxydans (Go) and Brettanomyces bruxellensis (Bb) are detrimental micro-organisms compromising wine quality through the production of acetic acid and undesirable aromas. Non-Saccharomyces yeasts, like Metschnikowia species, offer a bioprotective approach to control spoilage micro-organisms growth. Antagonist effects of forty-six Metschnikowia strains in a co-culture with Go or Bb in commercial grape juice were assessed. Three profiles were observed against Go: no effect, complete growth inhibition, and intermediate bioprotection. In contrast, Metschnikowia strains exhibited two profiles against Bb: no effect and moderate inhibition. These findings indicate a stronger antagonistic capacity against Go compared to Bb. Four promising Metschnikowia strains were selected and their bioprotective impact was investigated at lower temperatures in Chardonnay must. The antagonistic effect against Go was stronger at 16 °C compared to 20 °C, while no significant impact on Bb growth was observed. The bioprotection impact on Saccharomyces cerevisiae fermentation has been assessed. Metschnikowia strains' presence did not affect the fermentation time, but lowered the fermentation rate of S. cerevisiae. An analysis of central carbon metabolism and volatile organic compounds revealed a strain-dependent enhancement in the production of metabolites, including glycerol, acetate esters, medium-chain fatty acids, and ethyl esters. These findings suggest Metschnikowia species' potential for bioprotection in winemaking and wine quality through targeted strain selection.

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Metschnikowia persimmonesis, a novel endophytic yeast strain isolated from Diospyros kaki calyx, possesses strong antimicrobial activity. We investigated its potential use as an environmentally safe food biocontrol agent through genomics, transcriptomics, and metabolomics. Secondary metabolites were isolated from M. persimmonesis, followed by chemical structure elucidation, PUL gene cluster identification, and RNA sequencing. Pulcherrimin was isolated using 2 M NaOH, its structure was confirmed, and the yield was quantified. Biocontrol efficacy of M. persimmonesis on persimmon fruits and calyx was evaluated by assessing lesion diameter and disease incidence. Following compounds were isolated from M. persimmonesis co-culture with Botrytis cinerea and Fusarium oxysporum: fusaric acid, benzoic acid, benzeneacetic acid, 4-hydroxybenzeneacetic acid, 4-(-2-hydoxyethyl)-benzoic acid, cyclo (Leu-Leu), benzenemethanol, 4-hydroxy-benzaldehide, 2-hydroxy-4-methoxy-benzoic acid, 4-hydroxy-benzoic acid, lumichrome, heptadecanoic acid, and nonadecanoic acid. Exposing M. persimmonesis to different growth media conditions (with or without sugar) resulted in the isolation of five compounds: Tyrosol, Cyclo (Pro-Val), cyclo(L-Pro-L-Tyr), cyclo(Leu-Leu), and cyclo(l-tyrosilylicine). Differentially expressed gene analysis revealed 3264 genes that were significantly expressed (fold change ≥2 and p-value ≤0.05) during M. persimmonesis growth in different media, of which only 270 (8.27%) showed altered expression in all sample combinations with Luria-Bertani Agar as control. Minimal media with ferric ions and tween-80 triggered the most gene expression changes, with the highest levels of PUL gene expression and pulcherrimin yield (262.166 mg/L) among all media treatments. M. persimmonesis also produced a higher amount of pulcherrimin (209.733 mg/L) than Metschnikowia pulcherrima (152.8 mg/L). M. persimmonesis inhibited the growth of Fusarium oxysporum in persimmon fruit and calyx. Toxicity evaluation of M. persimmonesis extracts showed no harmful effects on the liver and mitochondria of zebrafish, and no potential risk of cardiotoxicity in hERG-HEK293 cell lines. Thus, M. persimmonesis can be commercialized as a potent and safe biocontrol agent for preserving food products.

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Although filamentous Ascomycetes may produce structures that are interpreted as male and female gametangia, ascomycetous yeasts are generally not considered to possess male and female sexes. In haplontic yeasts of the genus Metschnikowia, the sexual cycle begins with the fusion of two morphologically identical cells of complementary mating types. Soon after conjugation, a protuberance emerges from one of the conjugants, eventually maturing into an ascus. The originating cell can be regarded as an ascus mother cell, hence as female. We tested the hypothesis that the sexes, female or male, are determined by the mating types. There were good reasons to hypothesize further that mating type α cells are male. In a conceptually simple experiment, we observed the early stages of the mating reaction of mating types differentially labeled with fluorescent concanavalin A conjugates. Three large-spored Metschnikowia species, M. amazonensis, M. continentalis, and M. matae, were examined. In all three, the sexes were found to be independent of mating type, cautioning that the two terms should not be used interchangeably.

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This study took a novel approach to address the dual challenges of enhancing the ethanol content and aroma complexity in Laiyang pear wine. It focused on sorbitol as a pivotal element in the strategic selection of yeasts with specific sorbitol-utilization capabilities and their application in co-fermentation strategies. We selected two Saccharomyces cerevisiae strains (coded as Sc1, Sc2), two Metschnikowia pulcherrima (coded as Mp1, Mp2), and one Pichia terricola (coded as Tp) due to their efficacy as starter cultures. Notably, the Sc2 strain, alone or with Mp2, significantly increased the ethanol content (30% and 16%). Mixed Saccharomyces cerevisiae and Pichia terricola fermentation improved the ester profiles and beta-damascenone levels (maximum of 150%), while Metschnikowia pulcherrima addition enriched the phenethyl alcohol content (maximum of 330%), diversifying the aroma. This study investigated the efficacy of strategic yeast selection based on sorbitol utilization and co-fermentation methods in enhancing Laiyang pear wine quality and aroma.

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As a biological alternative to the antimicrobial action of SO2, bioprotection has been proposed to winemakers as a means to limit or prevent grape musts microbial alteration. Competition for nitrogenous nutrients and for oxygen are often cited as potential explanations for the effectiveness of bioprotection. This study analyses the effect of a bioprotective M. pulcherrima strain on the growth of one H. valbyensis strain and one H. uvarum strain. Bioprotection efficiency was observed only against H. valbyensis inoculated at the two lowest concentrations. These results indicate a potential species-dependent efficiency of the bioprotective strain and a strong impact of the initial ratio between bioprotective and apiculate yeasts. The analysis of the consumption of nitrogen compounds revealed that leucine, isoleucine, lysine and tryptophan were consumed preferentially by all three strains. The weaker assimilation percentages of these amino acids observed in H. valbyensis at 24 h growth suggest competition with M. pulcherrima that could negatively affects the growth of the apiculate yeast in co-cultures. The slowest rate of O2 consumption of H. valbyensis strain, in comparison with M. pulcherrima, was probably not involved in the bioprotective effect. Non-targeted metabolomic analyses of M. pulcherrima and H. valbyensis co-culture indicate that the interaction between both strains particularly impact lysin and tryptophan metabolisms.

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The bioprospection of indigenous microorganism strains with biotechnological potential represents a prominent trend. Metschnikowia yeasts exhibit diverse capabilities, such as ethanol reduction in winemaking, biocontrol potential, and lipid production. In this work, local Metschnikowia strains were isolated from different fruits by their ability to produce pulcherrimic acid, a molecule that has been linked to biocontrol activity and that binds iron giving colored colonies. Five strains were selected, each from one of five distinct sources. All of them were identified as M. pulcherrima. All five were able inhibit other yeasts and one M. pulcherrima, called M7, inhibited the growth of Aspergillus nidulans. The selected strains accumulated lipid bodies in stationary phase. Certain non-conventional yeasts like Hanseniaspora vineae are very sensitive to biomass drying, but cell extracts from M. pulcherrima added to the growth media as a source of antioxidant lipids increased their tolerance to drying. All strains isolated showed good stress tolerance (particularly to heat) and have nutrient requirements similar to a commercial M. pulcherrima strain. In addition, the M7 strain had a good growth in sugarcane and beet molasses and behaved like Saccharomyces cerevisiae in a growth medium derived from agricultural waste, a persimmon hydrolysate. Therefore, the isolation of local strains of Metschnikowia able to grow in a variety of substrates is a good source of biocontrol agents.

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During breeding, some oriental river prawns (Macrobrachium nipponense, de Haan), an important aquaculture species in China, exhibit yellowish-brown body colouration, reduced appetite, and vitality. Diseased prawns revealed characteristic emulsifying disease signs, including whitened musculature, hepatopancreatic tissues, milky haemolymph, and non-coagulation. The present study investigated the causative agent of M. nipponense infection through isolation, histopathology, molecular sequencing, and infection experiments. The pathogenic strain exhibited distinctive white colonies on Bengal red medium, with microscopic examination confirming the presence of yeast cells. Histopathological analysis revealed prominent pathological alterations and yeast cell infiltration in muscles, hepatopancreas and gills. Additionally, 26S rDNA sequencing of the isolated yeast strain LNMN2022 revealed Metschnikowia bicuspidata (GenBank: OR518659) as the causative agent. This strain exhibited a 98.28% sequence homology with M. bicuspidata LNMB2021 (GenBank: OK094821) and 96.62% with M. bicuspidata LNES0119 (GenBank: OK073903). The pathogenicity test confirmed that M. bicuspidata elicited clinical signs in M. nipponense consistent with those observed in natural populations, and the median lethal concentration was determined to be 3.3 × 105 cfu/mL. This study establishes a foundation for further investigations into the host range and epidemiological characteristics of the pathogen M. bicuspidata in aquatic animals and provides an empirical basis for disease management in M. nipponense.

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Invasive species can have large effects on native communities. When native and invasive species share parasites, an epidemic in a native species could facilitate or inhibit the invasion. We sought to understand how the incidence and timing of epidemics in native species caused by a generalist parasite influenced the success and impact of an invasive species. We focused on North American native and invasive species of zooplankton (Daphnia dentifera and Daphnia lumholtzi, respectively), that can both become infected with a fungal parasite (Metschnikowia bicuspidata). In a laboratory microcosm experiment, we exposed the native species to varying parasite inocula (none, low, high) and two invasive species introduction times (before or during an epidemic in the native species). We found that the invasive species density in treatments with the parasite was higher compared to uninfected treatments, though only the early invasion, low-parasite and uninfected treatments exhibited significant pairwise differences. However, invasive resting eggs were only found in the uninfected treatments. The density of the native species was lowest with a combination of the parasite present, and the invasive species introduced during the epidemic. Native infection prevalence in these treatments (late invasion, parasite present) was also higher than prevalence in treatments where the invasive species was introduced before the epidemic. Therefore, the timing of an invasion relative to an epidemic can affect both the native and invasive species. Our results suggest that the occurrence and timing of epidemics in native species can influence the impacts of a species invasion.

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Pulcherrimin is an iron (III) chelate of pulcherriminic acid that plays a role in antagonistic microbial interactions, iron metabolism, and stress responses. Some bacteria and yeasts produce pulcherriminic acid, but so far, pulcherrimin could not be produced in Saccharomyces cerevisiae. Here, multiple integrations of the Metschnikowia pulcherrima PUL1 and PUL2 genes in the S. cerevisiae genome resulted in red colonies, which indicated pulcherrimin formation. The coloration correlated positively and significantly with the number of PUL1 and PUL2 genes. The presence of pulcherriminic acid was confirmed by mass spectrometry. In vitro competition assays with the plant pathogenic fungus Botrytis caroliana revealed inhibitory activity on conidiation by an engineered, strong pulcherrimin-producing S. cerevisiae strain. We demonstrate that the PUL1 and PUL2 genes from M. pulcherrima, in multiple copies, are sufficient to transfer pulcherrimin production to S. cerevisiae and represent the starting point for engineering and optimizing this biosynthetic pathway in the future.

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Three Metschnikowia strains marketed as bioprotection yeasts were studied to compare their antimicrobial effect on a mixture of two Hanseniaspora yeast strains in synthetic must at 12 °C, mimicking pre-fermentative maceration by combining different approaches. The growth of the different strains was monitored, their nitrogen and oxygen requirements were characterised, and their metabolomic footprint in single and co-cultures studied. Only the M. fructicola strain and one M. pulcherrima strains colonised the must and induced the rapid decline of Hanseniaspora. The efficiency of these two strains followed different inhibition kinetics. Furthermore, the initial ratio between Metschnikowia and Hanseniaspora was an important factor to ensure optimal bioprotection. Nutrient consumption kinetics showed that apiculate yeasts competed with Metschnikowia strains for nutrient accessibility. However, this competition did not explain the observed bioprotective effect, because of the considerable nitrogen content remaining on the single and co-cultures. The antagonistic effect of Metschnikowia on Hanseniaspora probably implied another form of amensalism. For the first time, metabolomic analyses of the interaction in a bioprotection context were performed after the pre-fermentative maceration step. A specific footprint of the interaction was observed, showing the strong impact of the interaction on the metabolic modulation of the yeasts, especially on the nitrogen and vitamin pathways.

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Microbial diseases are of major concern in vitiviniculture as they cause grape losses and wine alterations, but the prevention with chemical substances represents a risk to human health and agricultural ecosystem. A promising alternative is the biocontrol and bioprotection activity of non-Saccharomyces yeasts, such as Metschnikowia pulcherrima, which also presents positive oenological traits when used in multistarter fermentations. The aim of this study was to assess the impact of a selected M. pulcherrima strain in the post-harvest withering and vinification of Garganega grapes to produce the sweet 'passito' wine Recioto di Gambellara DOCG (Italy). M. pulcherrima was firstly inoculated on grape at the beginning of the withering process, and afterwards in must for multistarter sequential microfermentation trials with Saccharomyces cerevisiae. Microbiological, chemical, and sensory analyses were carried out to monitor the vinification of treated and control grapes. Grape bunches during withering were a suitable environment for the colonization by M. pulcherrima, which effectively prevented growth of molds. Differences in grape must composition were observed, and the diverse inoculation strategies caused noticeable variations of fermentation kinetics, main oenological parameters, wine aroma profile, and sensory perception. M. pulcherrima proved effective to protect grapes against fungal infections during withering and contribute to alcoholic fermentation generating wine with distinguished aromatic characteristics.

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BACKGROUND: Use of alternative non-Saccharomyces yeasts in wine and beer brewing has gained more attention the recent years. This is both due to the desire to obtain a wider variety of flavours in the product and to reduce the final alcohol content. Given the metabolic differences between the yeast species, we wanted to account for some of the differences by using in silico models. RESULTS: We created and studied genome-scale metabolic models of five different non-Saccharomyces species using an automated processes. These were: Metschnikowia pulcherrima, Lachancea thermotolerans, Hanseniaspora osmophila, Torulaspora delbrueckii and Kluyveromyces lactis. Using the models, we predicted that M. pulcherrima, when compared to the other species, conducts more respiration and thus produces less fermentation products, a finding which agrees with experimental data. Complex I of the electron transport chain was to be present in M. pulcherrima, but absent in the others. The predicted importance of Complex I was diminished when we incorporated constraints on the amount of enzymatic protein, as this shifts the metabolism towards fermentation. CONCLUSIONS: Our results suggest that Complex I in the electron transport chain is a key differentiator between Metschnikowia pulcherrima and the other yeasts considered. Yet, more annotations and experimental data have the potential to improve model quality in order to increase fidelity and confidence in these results. Further experiments should be conducted to confirm the in vivo effect of Complex I in M. pulcherrima and its respiratory metabolism.

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In the current situation, wine areas are affected by several problems in a context of global warming: asymmetric maturities, pH increasing, high alcohol degree and flat wines with low freshness and poor aroma profile. The use of emerging biotechnologies allows to control or manage such problems. Emerging non-Saccharomyces as Lachancea thermotolerans are very useful for controlling pH by the formation of stable lactic acid from sugars with a slight concomitant alcohol reduction. Lower pH improves freshness increasing simultaneously microbiological stability. The use of Hanseniaspora spp. (specially H. vineae and H. opuntiae) or Metschnikowia pulcherrima promotes a better aroma complexity and improves wine sensory profile by the expression of a more complex metabolic pattern and the release of extracellular enzymes. Some of them are also compatible or synergic with the acidification by L. thermotolerans, and M. pulcherrima is an interesting biotool for reductive winemaking and bioprotection. The use of bioprotection is a powerful tool in this context, allowing oxidation control by oxygen depletion, the inhibition of some wild microorganisms, improving the implantation of some starters and limiting SO2. This can be complemented with the use of reductive yeast derivatives with high contents of reducing peptides and relevant compounds such as glutathione that also are interesting to reduce SO2. Finally, the use of emerging non-thermal technologies as Ultra High-Pressure Homogenization (UHPH) and Pulsed Light (PL) increases wine stability by microbial control and inactivation of oxidative enzymes, improving the implantation of emerging non-Saccharomyces and lowering SO2 additions. GRAPHICAL ABSTRACT.

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Flavour molecules are generated now-a-days through microbial fermentation on a commercial scale. γ-Decalactone (GDL) is an important molecule due to its long-lasting flavouring impact as buttery, coconut and peach-type. In the current study, 33 microorganisms were isolated from different fruit sources, and their screening for target GDL production was performed. Using DNA sequencing, two potential strains yielding good amounts of GDL were identified from pineapple and strawberry fruits. The identified strains were Metschnikowia vanudenii (OP954735) and Candida parapsilosis (OP954733), and further optimized by Taguchi method. The effectiveness of lactone production is influenced by the rate of microbial growth under various operating conditions. The factors such as substrate concentration, pH, temperature, cell density and rotation (rpm) with 3 levels were applied for the GDL production using M. vanudenii (OP954735) and C. parapsilosis (OP954733) strains. The results revealed that the highest molar conversion of GDL was 24.69% (115.7 mg/g quantitative yield) and 52.69% (272.0 mg/g quantitative yield) at the optimal conditions using SB-62 and PA-19 strains, respectively. The two novel strains are reported for the first time for production of γ-decalactone and overall, this study opens up the possibility of using Taguchi design for large scale up process development for producing food flavours utilising environmentally friendly natural strains.

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The reduction of chemical inputs in wine has become one of the main challenges of the wine industry. One of the alternatives to sulfites developed is bioprotection, which consists in using non-Saccharomyces strains to prevent microbial deviation. However, the impact of substituting sulfites by bioprotection on the final wine remains poorly studied. For the first time, we characterized this impact on Chardonnay wine through an integrative approach. Interestingly, physico-chemical analysis did not reveal any difference between both treatments regarding classical oenological parameters. Nevertheless, bioprotection did not seem to provide as much protection against oxidation as sulfites, as observed through phenolic compound analysis. At a deeper level, untargeted metabolomic analyses revealed substantial changes in wine composition according to must treatment. In particular, the specific footprint of each treatment revealed an impact on nitrogen-containing compounds. This observation could be related to modifications in S. cerevisiae metabolism, in particular amino acid biosynthesis and tryptophan metabolism pathways. Thus, the type of must treatment seemed to impact metabolic fluxes of yeast differently, leading to the production of different compounds. For example, we observed glutathione and melatonin, compounds with antioxidant properties, which were enhanced with sulfites, but not with bioprotection. However, despite substantial modifications in wines regarding their chemical composition, the change in must treatment did not seem to impact the sensory profile of wine. This integrative approach has provided relevant new insights on the impact of sulfite substitution by bioprotection on Chardonnay wines.

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Finding alternatives to the use of chemical inputs to preserve the sanitary and organoleptic quality of food and beverages is essential to meet public health requirements and consumer preferences. In oenology, numerous manufacturers already offer a diverse range of bio-protection yeasts to protect must against microbiological alterations and therefore limit or eliminate sulphites during winemaking. Bio-protection involves selecting non-Saccharomyces yeasts belonging to different genera and species to induce negative interactions with indigenous microorganisms, thereby limiting their development and their impact on the matrix. Although the effectiveness of bio-protection in the winemaking industry has been reported in numerous journals, the underlying mechanisms are not yet well understood. The aim of this review is to examine the current state of the art of field trials and laboratory studies that demonstrate the effects of using yeasts for bio-protection, as well as the interaction mechanisms that may be responsible for these effects. It focuses on the yeast Metschnikowia pulcherrima, particularly recommended for the bio-protection of grape musts.