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Pichia membranifaciens CYC 1086 secretes a killer toxin (PMKT2) that is inhibitory to a variety of spoilage yeasts and fungi of agronomical interest. The killer toxin in the culture supernatant was concentrated by ultrafiltration and purified to homogeneity by two successive steps, including native electrophoresis and HPLC gel filtration. Biochemical characterization of the toxin showed it to be a protein with an apparent molecular mass of 30 kDa and an isoelectric point of 3.7. At pH 4.5, optimal killer activity was observed at temperatures up to 20[degrees]C. Above approximately this pH, activity decreased sharply and was barely noticeable at pH 6. The toxin concentrations present in the supernatant during optimal production conditions exerted a fungicidal effect on a variety of fungal and yeast strains. The results obtained suggest that PMKT2 has different physico-chemical properties from PMKT as well as different potential uses in the biocontrol of spoilage yeasts. PMKT2 was able to inhibit Brettanomyces bruxellensis while Saccharomyces cerevisiae was fully resistant, indicating that PMKT2 could be used in wine fermentations to avoid the development of the spoilage yeast without deleterious effects on the fermentative strain. In small-scale fermentations, PMKT2, as well as P. membranifaciens CYC 1086, was able to inhibit B. bruxellensis, verifying the biocontrol activity of PMKT2 in simulated winemaking conditions.

The use of Pichia membranifaciens CYC 1106 killer toxin against Botrytis cinerea was investigated. This strain exerted a broad-specificity killing action against other yeasts and fungi. At pH 4, optimal killer activity was observed at temperatures up to 20 [degrees]C. At 25 [degrees]C the toxic effect was reduced to 70%. The killer activity was higher in acidic medium. Above about pH 4-5 activity decreased sharply and was barely noticeable at pH 6. The killer toxin protein from P. membranifaciens CYC 1106 was purified to electrophoretic homogeneity. SDS-PAGE of the purified killer protein indicated an apparent molecular mass of 18 kDa. Killer toxin production was stimulated in the presence of non-ionic detergents. The toxin concentrations present in the supernatant during optimal production conditions exerted a fungicidal effect on a strain of B. cinerea. The symptoms of infection and grey mould observed in Vitis vinifera plants treated with B. cinerea were prevented in the presence of purified P. membranifaciens killer toxin. The results obtained suggest that P. membranifaciens CYC 1106 killer toxin is of potential use in the biocontrol of B. cinerea.

Pichia membranifaciens killer toxin is discussed. P. membranifaciens CYC1106 is a yeast isolated from fermenting olive brines. It has been found that it binds mostly to the (1--6)-beta-D-glucan of the cell wall of a sensitive yeast (Candida boidinii IGC 3430). The toxin has maximum killer activity where NaCl is present.

The expression of the killer phenotype of yeasts isolated from spontaneously fermenting olive brines was analyzed to determine the effects of sodium chloride on the killer properties of the yeasts. The killer activity of culture filtrates from yeasts occurring in olive brines was significantly enhanced by the addition of sodium chloride. Furthermore, sodium chloride enlarged the activity spectra of a killer yeast against the selected target strains.

The use of commercial yeast strains is a common practice in winemaking leading to a predictable quality in wine production, avoiding stuck or sluggish fermentations. However, the use of commercial yeasts leads to a consequent reduction in autochthonous microbial diversity. In this study, 1047 isolates from three Spanish appellations of origin were checked for fingerprinting on interdelta polymorphisms and the strain composition and diversity analysed using an extensible open‐source platform for processing and analysis of an in‐house polymorphism database developed for this study. Ancient vineyards managed with organic practices showed intermediate to low levels of strains diversity indicating the existence of stable populations of Saccharomyces cerevisiae strains. A drastic reduction in the number of different S. cerevisiae strains was observed in vineyards with cellars using a selected autochthonous S. cerevisiae strain for winemaking. Contrary, the use of allochthonous commercial strains in wineries did not seem to affect the native S. cerevisiae strain composition and diversity. Significance and Impact of the Study: The aim of this study was to compare different viticulture and oenological practices to determine their influence on the composition and diversity of Saccharomyces cerevisiae strains in wine fermentations. The study shows that the use of autochthonous strains of S. cerevisiae as starters for wine fermentation could have an important incidence on S. cerevisiae strains diversity in surrounding vineyards. The use of autochthonous strains of S. cerevisiae reduced the detected number of S. cerevisiae strains, a fact that was not observed when allochthonous commercial strains were used. Furthermore, vineyards managed with organic practices showed intermediate to low levels of S. cerevisiae strain diversity, whereas conventional practices showed higher levels. [ABSTRACT FROM AUTHOR]

Examines the isolation and characterization of cell wall fractions from sensitive yeast C. boidinii IGC 3430 to determine the nature of the receptor for the killer toxin from P. membranifaciens CYC 1106. Identification of the primary receptor (1...6)-beta-D-glucan; Killer activities of P. mebranifaciens CYC 1106 and cerevisiae; Binding of killer toxins to cell wall fractions.

Analyzing the influence of different yeast species on several compounds with enological interest, it becomes possible to identify metabolic determinants of the incidence of yeasts on wine quality. Contrary to Saccharomyces cerevisiae, understand- ing genetic regulation, enzymatic properties and physiology of non-Saccharomyces species in enological conditions is far from being known. Because of this, the commercialization of industrial non-Saccharomyces strains on wine industry is showing a really slow pace. In order to determine the enzymatic properties of wine-related yeast species it is necessary to evaluate hundreds of yeast isolates enabling us to robustly attribute specific enzymatic activities to a specific group of yeast species. The contri- bution of yeasts to wine flavour is greatly determined by their impact on aromatic compounds release. Different glycosidases, β-lyase, pectinase, cellulase and protease activities are described as responsible for changes in wine composition, so determining inter- and intraspecific variability in these enzymatic properties in yeast species seems to be a useful tool for innovative yeast selection process. With the aim of relating enzymatic activities with a specific impact in wine properties we developed combined fermentations with non-Saccharomyces selected strains and industrial S. cerevisiae strains. The use of rational metabolomic analysis allows us to explain the physiology of non-Saccharomyces yeasts during wine fermentation and its incidence on wine quality.

Increasing food production while avoiding negative impacts on biodiversity constitutes one of the main challenges of our time. Traditional silvopastoral systems like Iberian oak savannas ("dehesas") set an example, where free-range livestock has been reared for centuries while preserving a high natural value. Nevertheless, factors decreasing productivity need to be addressed, one being acorn losses provoked by pest insects. An increased and focalized grazing by livestock on infested acorns would kill the larvae inside and decrease pest numbers, but increased livestock densities could have undesired side effects on ground arthropod communities as a whole. We designed an experimental setup including areas under trees with livestock exclosures of different ages (short-term: 1-year exclusion, long-term: 10-year exclusion), along with controls (continuous grazing), using DNA metabarcoding (mitochondrial markers COI and 16S) to rapidly assess arthropod communities' composition. Livestock removal quickly increased grass cover and arthropod taxonomic richness and diversity, which was already higher in short-term (1-year exclosures) than beneath the canopies of control trees. Interestingly, arthropod diversity was not highest at long-term exclosures (≥10 years), although their community composition was the most distinct. Also, regardless of treatment, we found that functional diversity strongly correlated with the vegetation structure, being higher at trees beneath which there was higher grass cover and taller herbs. Overall, the taxonomic diversity peak at short term exclosures would support the intermediate disturbance hypothesis, which relates it with the higher microhabitat heterogeneity at moderately disturbed areas. Thus, we propose a rotatory livestock management in dehesas: plots with increased grazing should co-exist with temporal short-term exclosures. Ideally, a few long-term excluded areas should be also kept for the singularity of their arthropod communities. This strategy would make possible the combination of biological pest control and arthropod conservation in Iberian dehesas., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Chitosan exerts a significant influence on various chemical parameters affecting the quality of wine produced using multiple strains of Lachancea thermotolerans . The impact of chitosan on these parameters varies depending on the specific strain studied. We observed that, under the influence of chitosan, the fermentation kinetics accelerated for all examined strains. The formation of lactic acid increased by 41% to 97% across the studied L. thermotolerans strains, depending on the specific strain. This effect also influenced acidity-related parameters such as total acidity, which increased by 28% to 60%, and pH, which experienced a decrease of over 0.5 units. The consumption of malic acid increased by 9% to 20% depending on the specific strain of L. thermotolerans . Nitrogen consumption also rose, as evidenced by all L. thermotolerans strains exhibiting a residual value of Primary Amino Nitrogen (PAN) of below the detection limit, and ammonia consumption increased by 90% to 100%, depending on the strain studied. However, certain parameters such as acetic acid, succinic acid, and glycerol showed contradictory results depending on the strain under investigation. In terms of volatile composition, chitosan supplementation led to increased production of i-butanol by 32% to 65%, 3-methylbutanol by 33% to 63%, and lactic acid ethyl ester by 58% to 91% across all studied strains of L. thermotolerans . Other analyzed aroma compounds exhibited varying changes depending on the specific strain of L. thermotolerans .

The study explores diverse strains of Lachancea thermotolerans in single-inoculum wine fermentation conditions using synthetic grape must. It aims to analyze the role of the species without external influences like other microorganisms or natural grape must variability. Commercial strains and selected vineyard isolates, untested together previously, are assessed. The research evaluates volatile and non-volatile chemical compounds in final wine, revealing significant strain-based variations. L. thermotolerans notably produces lactic acid and consumes malic acid, exhibiting moderate ethanol levels. The volatile profile displays strain-specific impacts, affecting higher alcohol and ester concentrations compared to S. cerevisiae . These effects vary based on the specific compounds. Using a uniform synthetic must enables direct strain comparisons, eliminating grape-related, environmental, or timing variables in the experiment, facilitating clearer insights into the behavior of L. thermotolerans in wine fermentation. The study compares for the first time all available commercial strains of L. thermotolerans ., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Santiago Benito reports financial support was provided by Spanish Ministry of Science and Innovation. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

Climate change is causing a lack of acidity during winemaking and oenologists use several solutions to cope with such a problem. Lachancea thermotolerans, which has the potential to tolerate the harsh physicochemical conditions of wine, has emerged as a promising alternative for pH management during winemaking and, currently, it is the most valuable yeast used for acidity control in wine. In this work a manageable method for L. thermotolerans genotyping based on a multiplexed microsatellite amplification in 6 different loci was developed. The proposed method was used to distinguish between 103 collection strains obtained from different geographical and isolation sources, and then challenged against a 429 L. thermotolerans isolates from several wineries and harvests. The procedure was also tested for fermentation monitoring and strain implantation. This approach was conceived to simplify the methodology available for L. thermotolerans genotyping, making it easy for applying in wine-related laboratories. This method can be applied to distinguish between L. thermotolerans strains in selection programs and to follow implantation of inoculated strains during winemaking with optimal results., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)