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28 pages, 3519 KB

Open AccessArticle

Impact of Grape Harvest Time on Wild Yeast Biodiversity and Its Influence on Wine Fermentation

by Mercè Sunyer-Figueres, Daniel Fernández-Vázquez, Cristina Cuesta-Martí, Inés Horcajo-Abal, Carlos Sánchez-Mateos, Alba Domènech, Enric Nart, Victoria A. Castillo-Olaya, Immaculada Andorrà and Miquel Puxeu

Microorganisms 2025, 13(12), 2836; https://doi.org/10.3390/microorganisms13122836 - 13 Dec 2025

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Abstract

Grape harvest time influences the berry composition, which impacts the organoleptic wine characteristics. Winemakers monitor technological, phenolic, and aromatic grape maturity to decide the harvest date. Little is known about the evolution of yeast ecology of grapes during the maturation period. The microbiota [...] Read more.

Grape harvest time influences the berry composition, which impacts the organoleptic wine characteristics. Winemakers monitor technological, phenolic, and aromatic grape maturity to decide the harvest date. Little is known about the evolution of yeast ecology of grapes during the maturation period. The microbiota involved in the fermentation impacts the wine composition and characteristics; therefore, changes in grape biodiversity could have an impact in fermentation kinetics and aroma compound formation. In this study, the yeast biodiversity of Grenache Noir and Carignan grapes from Montsant DO (Denomination of Origin) were analyzed at different ripening stages to assess how harvest time influences microbiota. The fermentation performance of the yeasts obtained was studied at both laboratory and pilot scales to evaluate the impact of these yeasts, both in pure and mixed cultures, on the wine’s chemical and aromatic composition as well as its sensory impact. The results indicated that early harvest favored a higher diversity of non-Saccharomyces species, whereas in more mature grapes, Saccharomyces cerevisiae species was dominant. The isolated strains of Saccharomyces differed in their fermentation performances, as well as ethanol content and acidity of wine. In general, they produced higher concentration of fermentative volatile compounds than a commercial strain. The non-Saccharomyces yeasts in mixed fermentations with the S. cerevisiae strains also impacted wine composition and characteristics, leading lower ethanol content or enhancing aromatic balance and sensory equilibrium. The results highlight that grape harvest timing influences microbial diversity and fermentative performance and thus should be considered to better determine the optimum harvest date and ensure consistent wine characteristics. Full article

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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18 pages, 4800 KB

Open AccessArticle

From Soil to Wine: Influence of Vegetative Covers on Microbial Communities and Fermentative Dynamics in Cabernet Sauvignon

by Gerardo Leal, Joan Miquel Canals, Gemma Beltran, Álvaro Peña-Neira, Carla Jara, Jaime Romero, Carolina Ramírez and René Sanz

Microorganisms 2025, 13(12), 2804; https://doi.org/10.3390/microorganisms13122804 - 9 Dec 2025

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Abstract

The implementation of vegetative cover crops in vineyards is a sustainable alternative to chemical weed control, potentially influencing both soil fertility and grape-associated microbiota. This study evaluated the impact of six groundcover management strategies under vines—white clover (Trifolium repens), red clover [...] Read more.

The implementation of vegetative cover crops in vineyards is a sustainable alternative to chemical weed control, potentially influencing both soil fertility and grape-associated microbiota. This study evaluated the impact of six groundcover management strategies under vines—white clover (Trifolium repens), red clover (Trifolium pratense), burr medic (Medicago polymorpha), lupine (Lupinus albus), spontaneous weeds, and an herbicide-treated control—on the microbial dynamics and physicochemical properties of Cabernet Sauvignon must and wine from the Maipo Valley, Chile. Amplicon sequencing of bacterial (16S rRNA) and fungal (ITS) communities was combined with spontaneous fermentation trials and chemical analyses of must and wine. Fungal and bacterial communities on grape surfaces were dominated by Ascomycota and Proteobacteria, respectively, with no significant compositional differences among treatments. During fermentation, Metschnikowia and Tatumella were the most abundant non-Saccharomyces and bacterial genera, respectively, showing dynamic shifts across fermentation stages. Legume-based covers, particularly red clover, increased wine total acidity and polyphenol index while reducing pH. Correlation analyses revealed associations between specific microbial taxa (Metschnikowia, Cohnella, Saliterribacillus) and key enological parameters. Overall, these findings demonstrate that leguminous cover crops subtly modulate vineyard microbial ecology and fermentation outcomes, offering an environmentally sustainable pathway to enhance enological differentiation in semi-arid viticultural regions. Full article

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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22 pages, 2162 KB

Open AccessArticle

Effect of UHPH and Sulphur Dioxide Content on Verdejo Vinification: Sensory, Chemical, and Microbiological Approach After Accelerated Aging Test

by Miquel Puxeu, Carlos Sánchez-Mateos, Inés Horcajo-Abal, Mercè Sunyer-Figueres, Victoria Castillo, Daniel Fernández-Vázquez, Alejandro Suárez, Natalia Santamaría, Enric Nart, Sergi de Lamo, Antonio Morata and Immaculada Andorrà

Microorganisms 2025, 13(11), 2623; https://doi.org/10.3390/microorganisms13112623 - 19 Nov 2025

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Abstract

Ultra-High-Pressure Homogenization (UHPH) is increasingly accepted by alimentation industries as a methodology to prevent microbial contamination with minimal impact on food organoleptic characteristics. Since 2022 the International Organization of Vine and Wine allows the use of high pressure (over 200 MPa), applied continuously, [...] Read more.

Ultra-High-Pressure Homogenization (UHPH) is increasingly accepted by alimentation industries as a methodology to prevent microbial contamination with minimal impact on food organoleptic characteristics. Since 2022 the International Organization of Vine and Wine allows the use of high pressure (over 200 MPa), applied continuously, in winemaking. While previous works reported the impact of UHPH technology on must microbiology and color; in the present work, the effect of UHPH in Verdejo is investigated, evaluated, and compared with the use of different amounts of sulphur dioxide from a sensorial, chemical, and microbiological point of view. Our findings indicate that combining low doses of sulphur dioxide with UHPH improves wine quality, increasing the floral and overall assessment and decreasing the amount of aging flavors after an accelerated aging test. This study provides new insights into the effect of UHPH on wine quality and, specifically, on how the aging potential contributes to a better understanding of the impact of UHPH technology on the shelf life of wines. Full article

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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17 pages, 5990 KB

Open AccessArticle

Brettanomyces bruxellensis Strains Display Variable Resistance to Cycloheximide: Consequences on the Monitoring of Wine

by Laura Olazabal, Quentin Dapzol, Warren Albertin, Cécile Miot-Sertier, Magali Deleris-Bou, Anita Boisramé and Marguerite Dols-Lafargue

Microorganisms 2025, 13(11), 2597; https://doi.org/10.3390/microorganisms13112597 - 14 Nov 2025

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Abstract

Brettanomyces bruxellensis is a yeast that causes spoilage in red wines due to its ability to produce volatile phenols, compounds associated with major sensory defects. Specific monitoring of low populations of this species in complex ecosystems such as wine during fermentation or aging [...] Read more.

Brettanomyces bruxellensis is a yeast that causes spoilage in red wines due to its ability to produce volatile phenols, compounds associated with major sensory defects. Specific monitoring of low populations of this species in complex ecosystems such as wine during fermentation or aging often relies on plating onto selective media supplemented with cycloheximide. However, the variability of B. bruxellensis sensitivity to this antibiotic needs to be better characterized. A collection of 175 B. bruxellensis strains was, thus, grown on YPD medium supplemented with increasing concentrations of cycloheximide (0 to 0.5 g.L⁻¹), and yeast development was monitored for 20 days by image analysis. This study revealed significant inter-strain variability, with some strains showing very late or even no growth at high cycloheximide concentrations. The cycloheximide inhibitory effect was also dose- and population-dependent. In addition, colony size was frequently reduced at high doses. Additional tests were conducted on a subset of strains grown in wines with either low pH or high alcohol content or containing sulfur dioxide and then plated in the presence of increasing concentrations of cycloheximide. This revealed a cumulative effect of wine and cycloheximide stresses that resulted in an even higher delay in yeast detection. The results confirm the huge phenotypic diversity of the species and highlight the need to adapt the plates’ incubation time, particularly when the selectivity and the doses of cycloheximide needed are high (samples taken in pre-fermentation phases or during fermentation) or in case of stressful wine analysis, to minimize the risk of false negatives. Full article

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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15 pages, 1337 KB

Open AccessArticle

Evaluation of Specific Torulaspora delbrueckii Fractions to Stimulate Malolactic Fermentation in Limiting Conditions

by Aitor Balmaseda, Paloma Toraño, Benjamin Leroux, José María Heras, Sibylle Krieger-Weber, Nicolas Rozès, Magali Deleris-Bou and Cristina Reguant

Microorganisms 2025, 13(10), 2391; https://doi.org/10.3390/microorganisms13102391 - 17 Oct 2025

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Abstract

Malolactic fermentation (MLF) is a bioprocess driven by lactic acid bacteria (LAB), which is desired in red and highly acidic white wines. Among all LAB, Oenococcus oeni is the main species in wine, followed by Lactiplantibacillus plantarum. The harsh conditions found in [...] Read more.

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

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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16 pages, 1377 KB

Open AccessArticle

Climate Change Adaptation in Winemaking: Combined Use of Non-Saccharomyces Yeasts to Improve the Quality of Pedro Ximénez Wines

by Fernando Sánchez-Suárez, Rafael Martínez-García and Rafael A. Peinado

Microorganisms 2025, 13(8), 1908; https://doi.org/10.3390/microorganisms13081908 - 15 Aug 2025

Cited by 2 | Viewed by 987

Abstract

This study evaluates the impact of two non-Saccharomyces yeasts, Lachancea thermotolerans and Metschnikowia pulcherrima, on the oenological and sensory characteristics of Pedro Ximénez (PX) white wines produced in warm regions of southern Spain. PX wines are particularly affected by climate change, [...] Read more.

This study evaluates the impact of two non-Saccharomyces yeasts, Lachancea thermotolerans and Metschnikowia pulcherrima, on the oenological and sensory characteristics of Pedro Ximénez (PX) white wines produced in warm regions of southern Spain. PX wines are particularly affected by climate change, often exhibiting low acidity and limited aromatic complexity. Fermentations were performed using pure and sequential cultures of these yeasts and compared to a control inoculated with Saccharomyces cerevisiae. Wines fermented with L. thermotolerans showed increased titratable acidity (up to 6.83 g/L), reduced pH (down to 3.02), and higher lactic acid concentrations, contributing to improved freshness and microbial stability. The use of M. pulcherrima led to a significant rise in ester production, enhancing fruity and floral aromatic notes. Sequential fermentation using both yeasts produced wines with the highest overall aromatic complexity and superior performance in sensory evaluations. These findings support the use of L. thermotolerans and M. pulcherrima as a promising biotechnological strategy to improve white wine quality under climate change conditions. Full article

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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11 pages, 1123 KB

Open AccessArticle

Biodiversity of Yeast Species Isolated During Spontaneous Fermentation: Influence of Grape Origin, Vinification Conditions, and Year of Study

by Ana Benito-Castellanos, Beatriz Larreina, María Teresa Calvo de La Banda, Pilar Santamaría, Lucía González-Arenzana and Ana Rosa Gutiérrez

Microorganisms 2025, 13(7), 1707; https://doi.org/10.3390/microorganisms13071707 - 21 Jul 2025

Cited by 1 | Viewed by 1180

Abstract

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 [...] Read more.

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

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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14 pages, 857 KB

Open AccessArticle

Step-Wise Ethanol Adaptation Drives Cell-Wall Remodeling and ROM2/KNR4 Activation in Brettanomyces bruxellensis

by Leslie Hernandez-Cabello, Nachla Rojas-Torres, Liliana Godoy, Camila G-Poblete, Yarabi Concha, Verónica Plaza, Luis Castillo, Héctor M. Mora-Montes and María Angélica Ganga

Microorganisms 2025, 13(7), 1489; https://doi.org/10.3390/microorganisms13071489 - 26 Jun 2025

Viewed by 1866

Abstract

Brettanomyces bruxellensis has been described as the main spoilage microorganism in wines due to its ability to produce volatile phenols, which negatively impact the final product’s organoleptic properties. This yeast can grow and survive in environments that are too nutritionally poor and stressful [...] Read more.

Brettanomyces bruxellensis has been described as the main spoilage microorganism in wines due to its ability to produce volatile phenols, which negatively impact the final product’s organoleptic properties. This yeast can grow and survive in environments that are too nutritionally poor and stressful for other microorganisms, and one of the stressful conditions it can endure is the high alcohol content in wine. In this study, cell wall morphology and the expression of some genes related to its composition were characterized under increasing ethanol concentrations to establish a possible ethanol resistance mechanism. B. bruxellensis LAMAP2480 showed greater resistance to β-1,3-glucanase activity when grown in media supplemented with 5% or 10% ethanol compared with the control assay (without ethanol). Transmission electron microscopy showed no significant differences in cell wall thickness during the different adaptation stages. However, the amount of wall polysaccharides and chitin briefly increased at 1% ethanol but returned to baseline at 5% and 10%. The amount of wall-associated protein increased progressively with each increment in ethanol concentration. In addition, overexpression of the ROM2 and KNR4/SMI1 genes was observed at 10% ethanol. These results suggest that the integrity of the cell wall might play an important role in the adaptation of B. bruxellensis to an ethanol-containing medium. Full article

(This article belongs to the Special Issue Wine Fermentation Microorganisms)

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