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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. • Mp0520 effects the fungal microbial community in cold maceration broth. • Mp0520 shows great increase in cell wall hydrolase activities in the maceration broth. • Mp0520 increases the content of terpenes in wine. • Mp0520 highlights wine varietal characteristics and improves wine aroma quality. • Mp0520 shows wide application potential in winemaking. [ABSTRACT FROM AUTHOR]

Grass, a low-cost lignocellulosic feedstock with relatively low lignin content, serves as a potential carbon source for microbial processes due to its polymeric sugar content. However, converting carbohydrates into monomeric sugars presents challenges due to complex lignocellulosic matrix. In this study, a mixture of perennial ryegrass and white clover (RG) was pretreated with alkaline before being enzymatically hydrolysed for use as a fermentable sugar source for the oleaginous yeast Metschnikowia pulcherrima. The dilute alkaline pretreatment approach resulted in a 3 times improvement in the conversion of RG, yielding a fermentable sugar concentration of 56.5 g/L. The yeast exhibited a growth yield (Ym/m) of 0.47, producing 23.7 g/L of dry cell weight at 20°C over 140 hours, with a lipid content of 37 % with a similar composition to high oleic palm oil. These findings suggest that RG can be utilised for formulating an oleaginous yeast medium containing the necessary nutrients. [Display omitted] • Sugar production from perennial ryegrass and white clover mixture increased nearly threefold with dilute NaOH pretreatment. • Cultivation in RG hydrolysates exhibited a DCW of 23.7 g/L and FAME profile similar to that observed in a chemical medium. • Neutralisation, rather than washing, for NaOH removal reduced biomass loss during the pretreatment stage by 18.5 %. [ABSTRACT FROM AUTHOR]

This review investigates the roles of Saccharomyces sp. And non-Saccharomyces yeasts, lactic acid bacteria (LAB), and bioprotection strategies in promoting sustainable winemaking. While Saccharomyces cerevisiae has long been central to fermentation, non-Saccharomyces species like Starmerella bacillaris , Torulaspora delbrueckii , and Lachancea thermotolerans are now recognized for enhancing flavor complexity and reducing the need for chemical additives. LAB, particularly Oenococcus oeni , contribute to malolactic fermentation, improving wine stability and sensory qualities, while aiding in sustainable waste management by converting by-products into valuable materials such as biogas. Bioprotection methods using yeasts like Metschnikowia pulcherrima offer a natural alternative to chemical preservatives, reducing sulfite use and chemical inputs. These microbial strategies align with sustainability goals by minimizing synthetic additives, promoting natural fermentation, and enhancing energy efficiency. Sustainable vineyard practices, such as promoting microbial diversity, cover cropping, and organic pest management, help maintain soil health, reduce chemical fertilizers, and improve vine resilience. The integration of these practices supports both environmental and economic sustainability, reducing production costs and enhancing product quality. Additionally, advances in omics approaches enable the development of tailored microbial consortia suited to specific environmental conditions, further improving the resilience and efficiency of winemaking, especially under climate variability. This comprehensive approach meets consumer demand for natural wines while reducing the wine industry's environmental footprint and improving economic viability. [Display omitted] [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effects of using a specific glutathione-rich inactivated yeast (GIY) during the pre-fermentative stages of winemaking. The antioxidant properties of GIY were evaluated both alone and in combination with a selected strain of Metschnikowia pulcherrima , at various grape pressing stages. The assessment included measuring soluble oxygen levels and color browning in the musts. Additionally, the impact of different oenological protocols on the final aromatic profile of the wines was assessed through volatile organic compounds (VOCs) and sensory analysis. The results showed that inoculating M. pulcherrima during the pre-pressing phase, followed by GIY addition during the post-pressing stage, effectively reduced O 2 uptake (1.49 mg/L) and minimized browning (0.093 OD at 420 nm) during the pre-fermentative phase. Furthermore, the protocol involving GIY addition during pressing resulted in higher concentrations of specific compounds associated with fruity and floral aromas, such as ethyl octanoate (26.03 mg/L) and ethyl decanoate (26.87 mg/L). Sensory evaluations confirmed that all treatments had no off-odours or off-flavours, but the M. pulcherrima and GIY combination received the highest scores for smoothness and colour attributes. In conclusion, GIY treatments offer a promising alternative to enhance wine colour and mitigate oxidation effects, potentially reducing the reliance SO 2 in wine production. [Display omitted] [ABSTRACT FROM AUTHOR]

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. • A selected S. cerevisiae strain boosted sorbitol utilization and ethanol in pear wine. • First report of P. terricola and M. pulcherrima growing in sorbitol. • P. terricola and S. cerevisiae co-fermentation elevated pear wine aroma complexity. • M. pulcherrima altered S. cerevisiae aroma, increasing phenethyl alcohol. [ABSTRACT FROM AUTHOR]

The development of new fermented foods and beverages requires more and more the use of new dehydrated yeasts species. In this context, the non- Saccharomyces (NS) yeasts Torulaspora delbrueckii , Metschnikowia pulcherrima and Lachancea thermotolerans are developed especially in winemaking as co-culture in the fermentation of the must or for the must bioprotection. However, during formulation-dehydration the yeast cells are exposed to several stresses that reduce cellular activity. Among these, the oxidative stress induced by the stabilization processes has been described as one of the main causes of cell death. In this study, we analyzed the effects of growth medium associated with two dehydration kinetics on the accumulation of reactive oxygen species (ROS) and lipid peroxidation levels. The cultivability of tested yeast strains was dependent on growth and dehydration conditions. The L. thermotolerans strain was the most sensible to dehydration when growing in nutrient-poor media, which was characterized by high levels of ROS, lipid peroxidation and reduced cultivability. In contrast, this yeast was able to restore its cultivability when growing in nutrient-rich medium before dehydration. The other NS yeast strains acquired resistance to oxidative stress similarly but in a growth-dehydration condition less dependent manner. These results showed that modulation of growing medium composition is a simple way to improve resistance to oxidative attack imposed by dehydration in NS yeasts. This was the first time that multiple quantitative and qualitative indicators of oxidative stress response in these three NS yeast strains were explored. Unlabelled Image • Oxidative stress responses were evaluated in three non- Saccharomyces (NS) wine yeasts. • Higher cultivability of NS yeasts was correlated to reduction of oxidative stress. • Lipid droplets accumulation was observed for cells grown in nutrient-poor medium. • Grown cells in nutrient-rich medium showed lower reactive oxygen species levels. • The yeast Lachancea thermotolerans was the most sensitive of all strains analyzed. [ABSTRACT FROM AUTHOR]

[Display omitted] • M. pulcherrima LI was used both as biocontrol agent on grape and as starter in wine. • M. pulcherrima LI reduced mold growth during grape withering in fruttaio and increased grape dehydration. • Multistarter fermentation with S. cerevisiae differentiated the aromatic profile of wines to a lesser extent than LI inoculation on grapes. 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. [ABSTRACT FROM AUTHOR]

Bioprotection by yeast addition is increasingly used in oenology as an alternative to sulfur dioxide (SO 2). Recent studies have also shown that it is likely to consume dissolved O 2. This ability could limit O 2 for other microorganisms and the early oxidation of the grape must. However, the ability of yeasts to consume O 2 in a context of bioprotection was poorly studied so far considering the high genetic diversity of non- Saccharomyces. The first aim of the present study was to perform an O 2 consumption rate (OCR) screening of strains from a large multi species collection found in oenology. The results demonstrate significant inter and intra species diversity with regard to O 2 consumption. In the must M. pulcherrima consumes O 2 faster than Saccharomyces cerevisiae and then other studied non-Saccharomyces species. The O 2 consumption was also evaluate in the context of a yeast mix used as industrial bioprotection (Metschnikowia pulcherrima and Torulaspora delbrueckii) in red must. These non- Saccharomyces yeasts were then showed to limit the growth of acetic acid bacteria, with a bioprotective effect comparable to that of the addition of sulfur dioxide. Laboratory experiment confirmed the negative impact of the non- Saccharomyces yeasts on Gluconobacter oxydans that may be related to O 2 consumption. This study sheds new lights on the use of bioprotection as an alternative to SO 2 and suggest the possibility to use O 2 consumption measurements as a new criteria for non- Saccharomyces strain selection in a context of bioprotection application for the wine industry. [Display omitted] • OCR is significantly different both at the inter et intra species level. • Bioprotection and cold temperatures limit the growth of acetic acid bacteria. • OCR is a new parameter for industrial bioprotection selection. [ABSTRACT FROM AUTHOR]

[Display omitted] • Wine composition is impacted by must treatment. • SO 2 substitution by bioprotection did not impact classical oenological parameters. • Significant metabolomic differences were not reflected in the wine sensory profile. • Fate of amino acids changed with treatment, like melatonin enhanced by sulfites. 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. [ABSTRACT FROM AUTHOR]

Abstract During wine fermentation, yeasts produce metabolites that are known growth regulators. The relationship between certain higher alcohols derived from aromatic amino acid metabolism and yeast signalling has previously been reported. In the present work, tryptophol (TrpOH) or melatonin (MEL), which are putative growth regulators, were added to alcoholic fermentations. Fermentations were performed with three different inocula, combining Saccharomyces cerevisiae and four non- Saccharomyces yeast species, under two nitrogen conditions. The combinations tested were: (i) only S. cerevisiae ; (ii) the mixture of four non- Saccharomyces species; and (iii) the combination of all five species together. The results revealed that the TrpOH and MEL addition caused changes in fermentation kinetics, viability and species distribution during fermentation, but it was dependent on the nitrogen present in the media and the composition of the inocula. Low nitrogen condition seemed to favour the presence of non- Saccharomyces species until mid-fermentation, although at the end of fermentation the imposition of Saccharomyces was higher in this condition. The presence of high concentrations of TrpOH resulted in limited growth and a delay in fermentation, noticeably significant in fermentations performed with S. cerevisiae inocula. These effects were reversed by the presence of non- Saccharomyces yeast in the medium. Low TrpOH concentration allowed faster fermentation with mixed non- Saccharomyces and Saccharomyces inocula. Moreover, in the absence of S. cerevisiae , a low concentration of TrpOH increased the presence of Torulaspora delbrueckii during fermentation with high nitrogen availability but not under low nitrogen conditions, when the population of S. bacillaris was higher than that in the control. The effects of MEL were particularly evident at the beginning and end of the process, primarily favouring the growth of non- Saccharomyces strains, especially the first hours after inoculation. Highlights • Tryptophol and melatonin affect fermentation kinetics and population dynamics. • Nitrogen concentration modified the effect of both compounds on population dynamics. • High amounts of tryptophol delayed single S. cerevisiae fermentations. • Tryptophol changes the population dynamics in absence of S. cerevisiae. • Melatonin improves the fermentation performance of non- Saccharomyces. [ABSTRACT FROM AUTHOR]

Pectinase enzymes have shown a considerable influence in both, sensitive and technological properties of wines. They can help to improve clarification process, releasing more color and flavor compounds entrapped in grape skin, facilitating the liberation of phenolic compounds. This work aims to find yeasts that, because of their native pectinases, can be applied on combined fermentations with Saccharomyces cerevisiae obtaining significant benefits over single-inoculated traditional fermentations. 462 yeast strains isolated from wineries were identified and tested for several enzymatic activities of recognized interest for enology industry. Considering the 7 identified species, only Aureobasidium pullulans , Metschnikowia pulcherrima and Metschnikowia fructicola showed polygalacturonase activity. Because of its interest in winemaking, due to its reported incidence in wine flavor, the impact of M. pulcherrima as a source of pectinolytic enzymes was analyzed by measuring its influence in filterability, turbidity and the increase on color, anthocyanin and polyphenol content of wines fermented in combination with S. cerevisiae . Among the strains screened, M. pulcherrima NS-EM-34 was selected, due to its polygalacturonase activity, for further characterization in both, laboratory and semi-industrial scale assays. The kinetics concerning several metabolites of enological concern were followed during the entire fermentation process at microvinification scale. Improved results were obtained in the expected parameters when M. pulcherrima NS-EM-34 was used, in comparison to wines fermented with S. cerevisiae alone and combined with other pectinolytic and non-pectinolytic yeasts ( A. pullulans and Lachancea thermotolerans , respectively), even working better than commercial enzymes preparations in most parameters. Additionally, M. pulcherrima NS-EM-34 was used at a semi-industrial scale combined with three different S. cerevisiae strains, confirming its potential application for red wine improvement on the mentioned sensorial and technological properties. [ABSTRACT FROM AUTHOR]

Abstract: The influence of temperature and relative humidity (RH) on the activity of three biocontrol agents—the yeast Metschnikowia pulcherrima LS16 and two strains of the yeast-like fungus Aureobasidium pullulans LS30 and AU34-2—against infection by A. carbonarius and ochratoxin A (OTA) accumulation in wine grape berries was investigated in lab-scale experiments. The presence of wounds on grape skin dramatically favored infection of berries by A. carbonarius strain A1102, since unwounded berries showed very low levels of infection at all conditions of RH and temperature tested. Artificially wounded berries pre-treated with the biocontrol agents were inoculated with the ochratoxigenic A. carbonarius strain A1102 and were incubated for 5days at two levels of RH (60% and 100%) and three different temperatures (20, 25 and 30°C). The three biocontrol agents were able to prevent infections at 60% RH and 20°C. At 60% RH and 25°C only strain AU34-2 achieved some protection on day 5, whereas at 30°C a limited biocontrol efficacy was evident only up to day 2. At 100% RH, LS16, LS30 and AU34-2 showed effective protection of grape berries at 20°C until the 5th day of incubation. The three biocontrol agents achieved significant protection at higher temperatures only until the 2nd day after the beginning of the experiment: all three strains at 25°C, and only strain LS16 at 30°C. After 5days, the three biocontrol agents were able to significantly reduce the level of OTA in berries at all the conditions tested. This occurred even when protection from infection was not significant, except at 30°C and 100% of RH for all the three strains, and at 25°C and 100% of RH for strain LS16. The biocontrol agents displayed a higher rate of colonization on grape berries at 20 and 25°C than at 30°C. The higher value of RH (100%) appeared to increase the rate of colonization, in particular at 20 and 25°C. Taken together, our results emphasize the significant influence of environmental factors on the effectiveness of biocontrol against A. carbonarius as well as on OTA contamination in wine grape berries, and the need for biocontrol agents that can cope with the environmental conditions that are conducive to attack by A. carbonarius. [Copyright &y& Elsevier]

[Display omitted] • Active films made with apple-based polymers carrying M. pulcherrima were developed. • The higher yeast viability was observed in films made with "apple residues." • MC and "apple residues" films exerted high antifungal activity in vitro. • A coating made with "apple residues" was successfully applied as a coating on apples. • Active coatings reduced the growth of P. expansum and patulin content on apples. The aim of this work has been to prevent the growth of Penicillium expansum on the surface of apples by means of the use of biocontrol agents in combination with edible coatings. For that, Metschnikowia pulcherrima yeast isolated from wild apples was incorporated in edible film matrices based on pectins, cellulose ethers with and without the addition of liquid culture medium (yeast extract and glucose) and apple pomace residues. The viability and biocontrol activity of the yeast incorporated in the films were evaluated after film drying process and after being stored for 21 days at 10 °C. Furthermore, the matrix that exhibited the best results was applied as a coating on apples artificially inoculated with P. expansum in order to evaluate its antifungal activity against P. expansum during storage and to prevent the production of its mycotoxin, the patulin. M. pulcherrima remained viable in films after drying step and after 21 days storage, but viability depended on the matrix composition, being lower in pectin films and higher in apple pomace films. The highest antimicrobial activity in vitro was observed in methylcellulose films supplemented with yeast extract and glucose (YEG), and in apple pomace residues films, matrix which was therefore selected to be applied as coatings on apples inoculated with P. expansum. The results proved that bioactive coatings significantly reduce P. expansum growth and patulin production during storage. The results obtained in this work give added value to the apple industry residues which, in combination with apple isolated M. pulcherrima can be applied as coatings on apples to provide protection against blue mold and its mycotoxin as an alternative of postharvest disease management. [ABSTRACT FROM AUTHOR]

• 98 volatile compounds were identified from black currant juice and beverages. • 51 compounds were significant in GC-O, ethyl esters important after fermentations. • Non- Saccharomyces yeasts resulted in significantly different volatile profiles. • S. bayanus fermentation increase fatty acids, alcohols, and musty odour. • All fermented black currant beverages tasted intensely sour. Black currants (Ribes nigrum) were fermented with Saccharomyces and non- Saccharomyces yeasts without added sugar to yield low-ethanol-content beverages. The effects of yeasts on the volatile compounds and sensory characteristics were analysed by HS-SPME-GC-MS, GC-O, and generic descriptive analysis. Ninety-eight volatile compounds were identified from the black currant juice and fermented beverages. Significant increases in the contents of esters (131 %), higher alcohols (391 %), and fatty acids (not present in juice sample) compared to initial juice were observed depending on the yeasts used. GC-O analysis revealed the higher impact of esters on the sensory properties of Saccharomyces bayanus- fermented beverage compared to the Torulaspora delbrueckii- fermented beverage. In the sensory evaluation, non- Saccharomyces yeasts resulted in a higher 'black currant odour'. However, all beverages were intensely sour, which can be a significant challenge in the development of alcoholic berry beverages. [ABSTRACT FROM AUTHOR]

A mixture of two yeast antagonists, Metschnikowia pulcherrima and Cryptococcus laurentii, originally isolated from apples and exhibiting greater biocontrol activity against blue mold of apple than either yeast applied alone, were used in combination with sodium bicarbonate (SBC) in a pilot test in which treated fruit were stored under commercial controlled atmosphere (CA) storage conditions. Conidia of Penicillium expansum, antagonists cells and SBC were added to the drench solution. The treatments were applied to apples by drenching entire bins filled with apples containing 100 wounded fruit evenly distributed among five positions in each bin. The treated fruit were stored in commercial CA storages for approximately six months in the 2005–2006 and 2006–2007 storage seasons and then evaluated for incidence of decay. In both years, the treatments with the antagonist alone or in combination with SBC were equally effective and reduced blue mold incidence by 84–97% in 2005–2006 and 73–82% in 2006–2007. SBC alone significantly reduced blue mold incidence compared to the non-treated control but was less effective than the antagonist alone or in combination with SBC. This pilot test showed that the combination of these two antagonists and SBC can be an effective decay control method under commercial CA conditions, confirming results from our earlier laboratory studies using similarly treated fruit stored under CA conditions. [Copyright &y& Elsevier]

Abstract: ‘Golden Delicious’ apples were wound-inoculated with Penicillium expansum and treated with various combinations of sodium bicarbonate and two antagonists (Metschnikowia pulcherrima, Cryptococcus laurentii), and then stored in air or controlled atmosphere (CA=1.4kPa O2 and 3kPa CO2) for 2 or 4 months at 1°C. The antagonists survived and their populations increased during both air and CA storage. The antagonists alone reduced blue mold but were more effective when combined. Sodium bicarbonate tended to reduce lesion size when used with these antagonist, either when they were used alone or combined. Storage under CA conditions also increased the effectiveness of both antagonist, when used alone or in combination. The only treatment that completely eliminated P. expansum-incited decay was the combination of the two antagonists and sodium bicarbonate on fruit stored under CA conditions. The proper combination of alternative control measures can provide commercially acceptable long-term control of fruit decay and could help reduce our dependency on fungicides. [Copyright &y& Elsevier]

Abstract: ‘Golden Delicious’ apples were wound-inoculated with either Colletotrichum acutatum or Penicillium expansum and then treated with various combinations of heat (38°C) for 4 days, 2% sodium bicarbonate, and two biocontrol agents alone or combined. The fruit were stored for 4 months at 1°C and then at 20°C for 2 weeks. Either heat or the antagonists reduced decay caused by C. acutatum, but a combination of the two was required to completely eliminate decay caused by this pathogen in most cases. Sodium bicarbonate alone or in combination with the antagonists had little effect on C. acutatum. The antagonists alone reduced decay caused by P. expansum but tended to be more effective when combined. Sodium bicarbonate increased the effectiveness of decay control by each antagonist alone or in combination. All of the treatments that included heat virtually eliminated decay caused by this pathogen. The proper combination of alternative control measures can provide an effective strategy to reduce postharvest decay of apple fruit. [Copyright &y& Elsevier]

The application of a cell suspension of the BIO126 strain of Metschnikowia pulcherrima proved to be highly effective in the control of blue and grey mould, two of the most severe postharvest diseases on apple fruit. The possibilities of integrating the application of the antagonist with chemicals such as acibenzolar-S-methyl (ASM), ethanol, or sodium bicarbonate and a heat treatment were investigated in this work. The fruit were stored at 23 °C for 5 days and at 4 °C for 20 days. The antagonist, applied at 108 cells ml-1, proved to be the key element for the control of both pathogens, being more efficient after cold storage, with a reduction of 56.6 and 97.2% in the lesion diameter of blue and grey moulds. Ethanol and acibenzolar-S-methyl permitted a partial reduction in grey mould severity. Heat treatment and sodium bicarbonate significantly improved the efficacy of the biocontrol agent against blue mould with storage at 23 °C. To study the possibility of a single application of the biocontrol agent (107 cells ml-1) with reduced dosages of sodium bicarbonate or ethanol, the viability of the micro-organism with these chemicals was studied and a new set of experiments was established. Against both pathogens, the greater reduction in lesion diameter was obtained by treating simply with the biocontrol agent. Significant results on blue mould were provided by the application of 20% ethanol or 5% sodium bicarbonate before the biocontrol agent and by the application of BIO126 in 0.1% sodium bicarbonate. The application of the cell suspension of BIO126 M. pulcherrima, preceded or not by a pre-treatment with sodium bicarbonate or ethanol, could become a successful alternative to fungicide usage in postharvest disease control of pome fruit. The fungistatic effect of ethanol or sodium bicarbonate could be associated with the effect of the biocontrol agent, being persistent on the fruit for long periods. [Copyright &y& Elsevier]

The mechanisms of action and efficacy of four isolates (GS37, GS88, GA102, and BIO126) of the yeast Metschnikowia pulcherrima against Botrytis cinerea, Penicillium expansum, Alternaria sp., and Monilia sp., all postharvest pathogens of apple fruit, were studied in vitro and on apples, in controlled and semi-commercial conditions. An application of a cell suspension (108 cells per ml) of the antagonists in artificial wounds of apples reduced growth of B. cinerea and P. expansum after storage at 23 °C. A complete suppression of the pathogen was obtained against Monilia sp., stored at 23 °C, and against B. cinerea and P. expansum, stored at 4 °C. The results against Alternaria sp. were more variable. Applications of culture filtrates and autoclaved cells of the isolates were ineffective in reducing the diameter of the lesions on the fruit, supporting the hypothesis that living cells are necessary for biocontrol. In experiments of antagonism in vitro, on different solid substrates, a reduction in the mycelium growth of the pathogens resulted, so that, at least in vitro, the antagonists could produce some diffusible toxic metabolites. In co-cultivation in vitro on a synthetic medium, B. cinerea spore (105 ml−1) germination was completely inhibited by the presence of 108 cells of the antagonists, while culture filtrates and autoclaved suspensions were not able to reduce germination. Dipping boxes of apples cv. Golden delicious in a suspension of 107 antagonist cells per ml and storing for 8 months in controlled atmosphere at 1 °C, showed levels of control against B. cinerea and P. expansum similar to those from thiabendazole. [Copyright &y& Elsevier]

Metschnikowia pulcherrima , a non-traditional oleaginous yeast, was investigated using 0.5–20 g/L of acetic, propionic, butyric and valeric acid (four model products of anaerobic acidification) respectively as carbon sources to accumulate microbial lipids so as to produce biodiesel. The results showed that acetic acid was the most efficient carbon source for improving cell growth, lipid content and biodiesel quality. After 8 cycles non-sterile batch cultivation, M. pulcherrima still dominated in bioreactor. Controlling C/P ratio, K+ and Na+ concentrations, and initial pH could effectively enhance the lipid content to 25.79 ± 0.51%. Under nitrogen starvation, M. pulcherrima performed higher cellular activity and lipid productivity than that of phosphorous limitation. The biomass yield was about twice and the lipid content was 3% higher. Besides, different from the lipid improvement strategy of traditional oleaginous yeast, acetate from ethanol served as the precursor of M. pulcherrima cytoplasmic acetyl-CoA instead of citric acid, and pentose phosphate pathway (PPP) was the main donor of reducing equivalent NADPH by transcriptomics sequencing analysis. These results demonstrate M. pulcherrima is a promising species for large-scale production of biofuel from organic wastes due to the low operating and maintenance costs of non-sterile cultivation, which is beneficial for reducing environmental pollution and energy shortages. [Display omitted] • Acetic acid was optimum VFAs type for M. pulcherrima to produce lipid in batch mode. • Regulating C/P ratio, K+ and Na+ concentration and initial pH enhanced lipid yield. • N limitation was better than P limitation for lipid accumulation of M. pulcherrima. • Repeated non-sterile batch cultivation could be used to improve lipid production. • Transcriptomics sequencing analysis suggested special lipid biosynthesis mechanism. [ABSTRACT FROM AUTHOR]

• The effect of non- Saccharomyces on malolactic fermentation depends on the yeast. • Torulaspora delbrueckii and Metschnikowia pulcherrima were the best MLF compatible. • Hanseniaspora uvarum and Starmerella bacillaris had negative impact on MLF. Interest in using non- Saccharomyces yeasts in winemaking has increased in recent years due to their ability to improve wine quality. However, little information has been published regarding the possible effect on malolactic fermentation (MLF), carried out mostly by Oenococcus oeni. The aim of this paper is therefore to evaluate the effect of the most representative non- Saccharomyces species on O. oeni and wine MLF. Different strains of Torulaspora delbrueckii , Metschnikowia pulcherrima , Hanseniaspora uvarum , Hanseniaspora vineae and Starmerella bacillaris (syn. Candida zemplinina) were used in sequential alcoholic fermentation with Saccharomyces cerevisiae. The resulting wines were inoculated with four O. oeni strains. The action of non- Saccharomyces affected the final wine composition and the later role of O. oeni. Some of its strains could not perform MLF in H. uvarum wine due to high SO 2 concentrations. In some cases, MLF was inhibited in wines inoculated with S. bacillaris. All the H. uvarum and H. vineae strains notably increased acetic acid concentrations, thus threatening wine quality. The best conditions for MLF were provided by some T. delbruecckii and M. pulcherrima strains , which showed increased concentrations of mannoproteins – compounds described as MLF activators –, no production of SO 2 , and low consumption of L-malic acid. In conclusion, non- Saccharomyces yeasts have diverse effects on O. oeni and MLF depending on the species, with T. delbrueckii and M. pulcherrima being those that showed the best compatibility with MLF development. [ABSTRACT FROM AUTHOR]

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

Abstract Macroalgae (seaweeds) represent an emerging resource for food and the production of commodity and specialty chemicals. In this study, a single-step microwave process was used to depolymerise a range of macroalgae native to the United Kingdom, producing a growth medium suitable for microbial fermentation. The medium contained a range of mono- and polysaccharides as well as macro- and micronutrients that could be metabolised by the oleaginous yeast Metschnikowia pulcherrima. Among twelve macroalgae species, the brown seaweeds exhibited the highest fermentation potential, especially the kelp Saccharina latissima. Applying a portfolio of ten native M. pulcherrima strains, yeast growth kinetics, as well as production of lipids and 2-phenylethanol were examined, with productivity and growth rate being strain dependent. On the 2 L scale, 6.9 g L−1 yeast biomass – a yield of 0.14 g g−1 with respect to the supplied macroalgae – containing 37.2% (w/w) lipid was achieved through utilisation of the proteins, mono- and polysaccharides from S. latissima , with no additional enzymes. In addition, the yeast degraded a range of fermentation inhibitors released upon microwave processing at high temperatures and long holding times. As macroalgae can be cultured to food grade, this system offers a novel, potentially low-cost route to edible microbial oils as well as a renewable feedstock for oleochemicals. Graphical abstract Unlabelled Image Highlights • Growth media generated from twelve macroalgae in a single-step microwave process. • Oleaginous Metschnikowia pulcherrima valorised macroalgal saccharides and proteins. • 6.9 g L−1 (0.15 g g−1) yeast with 37% lipid produced from the hydrolysed S. latissima • Versatility of M. pulcherrima could become key in a marine biorefinery. [ABSTRACT FROM AUTHOR]