Predictability of the community-function landscape in wine yeast ecosystems.

Predictively linking taxonomic composition and quantitative ecosystem functions is a major aspiration in microbial ecology, which must be resolved if we wish to engineer microbial consortia. Here, we have addressed this open question for an ecological function of major biotechnological relevance: alcoholic fermentation in wine yeast communities. By exhaustively phenotyping an extensive collection of naturally occurring wine yeast strains, we find that most ecologically and industrially relevant traits exhibit phylogenetic signal, allowing functional traits in wine yeast communities to be predicted from taxonomy. Furthermore, we demonstrate that the quantitative contributions of individual wine yeast strains to the function of complex communities followed simple quantitative rules. These regularities can be integrated to quantitatively predict the function of newly assembled consortia. Besides addressing theoretical questions in functional ecology, our results and methodologies can provide a blueprint for rationally managing microbial processes of biotechnological relevance.

Impact of rare yeasts in Saccharomyces cerevisiae wine fermentation performance: Population prevalence and growth phenotype of Cyberlindnera fabianii, Kazachstania unispora, and Naganishia globosa.

Saccharomyces cerevisiae is a highly fermentative species able to complete the wine fermentation. However, the interaction with other non-Saccharomyces yeasts can determine the fermentation performance of S. cerevisiae. We have characterised three rare non-Saccharomyces yeasts (Cyberlindnera fabianii, Kazachstania unispora and Naganishia globosa), studying their impact on S. cerevisiae fitness and wine fermentation performance. Using a wide meta-taxonomic dataset of wine samples, analysed through ITS amplicon sequencing, we show that about a 65.07% of wine samples contains Naganishia spp., a 27.21% contains Kazachstania spp., and only a 4.41% contains Cyberlindnera spp; in all cases with average relative abundances lower than 1% of total fungal populations. Although the studied N. globosa strain showed a limited growth capacity in wine, both K. unispora and C. fabianii showed a similar growth phenotype to that of S. cerevisiae in different fermentation conditions, highlighting the outstanding growth rate values of K. unispora. In mixed fermentations with S. cerevisiae, the three yeast species affected co-culture growth parameters and wine chemical profile (volatile compounds, polysaccharides and proteins). K. unispora DN201 strain presents an outstanding capacity to compete with S. cerevisiae strains during the first stage of wine fermentation, causing stuck fermentations in both synthetic and natural grape musts.

Ulcerative Colitis Seems to Imply Oral Microbiome Dysbiosis.

Ulcerative colitis (UC) is a recurrent pathology of complex etiology that has been occasionally associated with oral lesions, but the overall composition of the oral microbiome in UC patients and its role in the pathogenesis of the disease are still poorly understood. In this study, the oral microbiome of UC patients and healthy individuals was compared to ascertain the possible changes in the oral microbial communities associated with UC. For this, the salivary microbiota of 10 patients diagnosed with an active phase of UC and 11 healthy controls was analyzed by 16S rRNA gene sequencing (trial ref. ISRCTN39987). Metataxonomic analysis revealed a decrease in the alpha diversity and an imbalance in the relative proportions of some key members of the oral core microbiome in UC patients. Additionally, Staphylococcus members and four differential species or phylotypes were only present in UC patients, not being detected in healthy subjects. This study provides a global snapshot of the existence of oral dysbiosis associated with UC, and the possible presence of potential oral biomarkers.

Expectable diversity patterns in wine yeast communities.

Wine fermentations are dominated by Saccharomyces yeast. However, dozens of non-Saccharomyces yeast genera can be found in grape musts and in the early and intermediate stages of wine fermentation, where they co-exist with S. cerevisiae. The diversity of non-Saccharomyces species is determinant for the sensorial attributes of the resulting wines, both directly (by producing aroma impact compounds) and indirectly (modulating the performance of Saccharomyces). Many research groups worldwide are exploring the great diversity of wine yeasts to exploit their metabolic potential to improve wine flavor or to prevent wine spoilage. In this work, we share a new data set from a wide ITS amplicon survey of 272 wine samples, and we perform a preliminary exploration to build a catalogue of 242 fungal and yeast genera detectable in wine samples, estimating global figures of their prevalence and relative abundance patterns across wine samples. Thus, our mycobiome survey provides a broad measure of the yeast diversity potentially found in wine fermentations; we hope that the wine yeast research community finds it useful, and we also want to encourage further discussion on the advantages and limitations that meta-taxonomic studies may have in wine research and industry.

Global distribution of IRC7 alleles in Saccharomyces cerevisiae populations: a genomic and phenotypic survey within the wine clade.

The adaptation to the different biotic and abiotic factors of wine fermentation has led to the accumulation of numerous genomic hallmarks in Saccharomyces cerevisiae wine strains. IRC7, a gene encoding a cysteine-S-ß-lyase enzyme related volatile thiols production in wines, has two alleles: a full-length allele (IRC7F ) and a mutated one (IRC7S ), harbouring a 38 bp-deletion. Interestingly, IRC7S -encoding a less active enzyme - appears widespread amongst wine populations. Studying the global distribution of the IRC7S allele in different yeast lineages, we confirmed its high prevalence in the Wine clade and demonstrated a minority presence in other domesticated clades (Wine-PDM, Beer and Bread) while it is completely missing in wild clades. Here, we show that IRC7S -homozygous (HS) strains exhibited both fitness and competitive advantages compared with IRC7F -homozygous (HF) strains. There are some pieces of evidence of the direct contribution of the IRC7S allele to the outstanding behaviour of HS strains (i.e., improved response to oxidative stress conditions and higher tolerance to high copper levels); however, we also identified a set of sequence variants with significant co-occurrence patterns with the IRC7S allele, which can be co-contributing to the fitness and competitive advantages of HS strains in wine fermentations.

Effects on varietal aromas during wine making: a review of the impact of varietal aromas on the flavor of wine.

Although there are many chemical compounds present in wines, only a few of these compounds contribute to the sensory perception of wine flavor. This review focuses on the knowledge regarding varietal aroma compounds, which are among the compounds that are the greatest contributors to the overall aroma. These aroma compounds are found in grapes in the form of nonodorant precursors that, due to the metabolic activity of yeasts during fermentation, are transformed to aromas that are of great relevance in the sensory perception of wines. Due to the multiple interactions of varietal aromas with other types of aromas and other nonodorant components of the complex wine matrix, knowledge regarding the varietal aroma composition alone cannot adequately explain the contribution of these compounds to the overall wine flavor. These interactions and the associated effects on aroma volatility are currently being investigated. This review also provides an overview of recent developments in analytical techniques for varietal aroma identification, including methods used to identify the precursor compounds of varietal aromas, which are the greatest contributors to the overall aroma after the aforementioned yeast-mediated odor release.

Analytical impact of Metschnikowia pulcherrima in the volatile profile of Verdejo white wines.

Most wine aroma compounds, including the varietal fraction, are produced or released during wine production and derived from microbial activity. Varietal aromas, typically defined as terpenes and thiols, have been described as derived from their non-volatile precursors, released during wine fermentation by different yeast hydrolytic enzymes. The perception of these minority aroma compounds depends on the chemical matrix of the wine, especially on the presence of majority aroma compounds, such as esters or higher alcohols. Strategies aiming to reduce the production of these masking flavors are on the spotlight of enology research as a way to better define varietal standard profiles for the global market. Using a natural white must from Verdejo variety (defined as a thiol grape variety), here we describe the analytical and sensorial impact of using, in sequential inoculations, a selected strain of Metschnikowia pulcherrima, in combination with two different Saccharomyces cerevisiae strains. An increase in the levels of the thiol 4-MSP (4-methyl-4-sulfanylpentan-2-one) over its sensory threshold, together with a decrease in higher alcohol production, was observed when M. pulcherrima was used. This has an important impact on these wines, making them fruitier and fresher, always preferred by the sensory panel.

Microbial Contribution to Wine Aroma and Its Intended Use for Wine Quality Improvement.

Wine is a complex matrix that includes components with different chemical natures, the volatile compounds being responsible for wine aroma quality. The microbial ecosystem of grapes and wine, including Saccharomyces and non-Saccharomyces yeasts, as well as lactic acid bacteria, is considered by winemakers and oenologists as a decisive factor influencing wine aroma and consumer's preferences. The challenges and opportunities emanating from the contribution of wine microbiome to the production of high quality wines are astounding. This review focuses on the current knowledge about the impact of microorganisms in wine aroma and flavour, and the biochemical reactions and pathways in which they participate, therefore contributing to both the quality and acceptability of wine. In this context, an overview of genetic and transcriptional studies to explain and interpret these effects is included, and new directions are proposed. It also considers the contribution of human oral microbiota to wine aroma conversion and perception during wine consumption. The potential use of wine yeasts and lactic acid bacteria as biological tools to enhance wine quality and the advent of promising advice allowed by pioneering -omics technologies on wine research are also discussed.

Outlining the influence of non-conventional yeasts in wine ageing over lees.

During the last decade, the use of innovative yeast cultures of both Saccharomyces cerevisiae and non-Saccharomyces yeasts as alternative tools to manage the winemaking process have turned the oenology industry. Although the contribution of different yeast species to wine quality during fermentation is increasingly understood, information about their role in wine ageing over lees is really scarce. This work aims to analyse the incidence of three non-Saccharomyces yeast species of oenological interest (Torulaspora delbrueckii, Lachancea thermotolerans and Metschnikowia pulcherrima) and of a commercial mannoprotein-overproducer S. cerevisiae strain compared with a conventional industrial yeast strain during wine ageing over lees. To evaluate their incidence in mouthfeel properties of wine after 4 months of ageing, the mannoprotein content of wines was evaluated, together with other wine analytic parameters, such as colour and aroma, biogenic amines and amino acids profile. Some differences among the studied parameters were observed during the study, especially regarding the mannoprotein concentration of wines. Our results suggest that the use of T. delbrueckii lees in wine ageing is a useful tool for the improvement of overall wine quality by notably increasing mannoproteins, reaching values higher than obtained using a S. cerevisiae overproducer strain. Copyright © 2016 John Wiley & Sons, Ltd.

Dynamic analysis of physiological properties of Torulaspora delbrueckii in wine fermentations and its incidence on wine quality.

This work examines the physiology of a new commercial strain of Torulaspora delbrueckii in the production of red wine following different combined fermentation strategies. For a detailed comparison, several yeast metabolites and the strains implantation were measured over the entire fermentation period. In all fermentations in which T. delbrueckii was involved, the ethanol concentration was reduced; some malic acid was consumed; more pyruvic acid was released, and fewer amounts of higher alcohols were produced. The sensorial properties of final wines varied widely, emphasising the structure of wine in sequential fermentations with T. delbrueckii. These wines presented the maximum overall impression and were preferred by tasters. Semi-industrial assays were carried out confirming these differences at a higher scale. No important differences were observed in volatile aroma composition between fermentations. However, differences in mouthfeel properties were observed in semi-industrial fermentations, which were correlated with an increase in the mannoprotein content of red wines fermented sequentially with T. delbrueckii.

Membrane bioreactor wastewater treatment plants reveal diverse yeast and protist communities of potential significance in biofouling.

The yeast community was studied in a municipal full-scale membrane bioreactor wastewater treatment plant (MBR-WWTP). The unexpectedly high diversity of yeasts indicated that the activated sludge formed a suitable environment for them to proliferate, with cellular concentrations of 2.2 ± 0.8 × 10(3) CFU ml(-1). Sixteen species of seven genera were present in the biological reactor, with Ascomycetes being the most prevalent group (93%). Most isolates were able to grow in a synthetic wastewater medium, adhere to polyethylene surfaces, and develop biofilms of variable complexity. The relationship between yeast populations and the protists in the MBR-WWTP was also studied, revealing that some protist species preyed on and ingested yeasts. These results suggest that yeast populations may play a role in the food web of a WWTP and, to some extent, contribute to membrane biofouling in MBR systems.

Cell cycle arrest and apoptosis, two alternative mechanisms for PMKT2 killer activity.

Pichia membranifaciens CYC 1086 secretes a unique 30kDa killer toxin (PMKT2) that inhibits a variety of spoilage yeasts and fungi of agronomical interest. The cytocidal effect of PMKT2 on Saccharomyces cerevisiae cells was studied. Metabolic events associated with the loss of S. cerevisiae viability caused by PMKT2 were qualitatively identical to those reported for K28 killer toxin activity, but different to those reported for PMKT. At higher doses, none of the cellular events accounting for the action of PMKT, the killer toxin secreted by P. membranifaciens CYC 1106, was observed for PMKT2. Potassium leakage, sodium influx and the decrease of intracellular pH were not among the primary effects of PMKT2. We report here that this protein is unable to form ion-permeable channels in liposome membranes, suggesting that channel formation is not the mechanism of cytotoxic action of PMKT2. Nevertheless, flow cytometry studies have revealed a cell cycle arrest at an early S-phase with an immature bud and pre-replicated 1n DNA content. By testing the sensitivity of cells arrested at different stages in the cell cycle, we hoped to identify the execution point for lethality more precisely. Cells arrested at the G1-phase by α-factor or arrested at G2-phase by the spindle poison methyl benzimidazol-2-yl-carbamate (MBC) were protected against the toxin. Cells released from the arrest in both cases were killed by PMKT2 at a similar rate. Nevertheless, cells released from MBC-arrest were able to grow for a short time, and then viability dropped rapidly. These findings suggest that cells released from G2-phase are initially able to divide, but die in the presence of PMKT2 after initiating the S-phase in a new cycle, adopting a terminal phenotype within that cycle. By contrast, low doses of PMKT and PMKT2 were able to generate the same cellular response. The evidence presented here shows that treating yeast with low doses of PMKT2 leads to the typical membranous, cytoplasmic, mitochondrial and nuclear markers of apoptosis, namely, the production of reactive oxygen species, DNA strand breaks, metacaspase activation and cytochrome c release.

Hypertension- and glycaemia-lowering effects of a grape-pomace-derived seasoning in high-cardiovascular risk and healthy subjects. Interplay with the gut microbiome.

Purpose: Grape pomace (GP) is a winery by-product rich in polyphenols and dietary fibre. Some recent results suggest that GP-derived extracts could be promising additives in food, specially recommended for low-salt diets. The hypothesis tested in this paper is that the regular consumption of GP-derived seasonings could help in the control of hypertension and glycaemia. Methods: A randomized intervention study (6 weeks) was performed in high-risk cardiovascular subjects (n = 17) and in healthy subjects (n = 12) that were randomly allocated into intervention (2 g day-1 of GP seasoning) or control (no seasoning consumed) groups. Blood samples, faeces, urine and blood pressure (BP) were taken at the baseline and at the end of the intervention. Faecal samples were analysed for microbiota composition (16S rRNA gene sequencing) and microbial-derived metabolites (short chain fatty acids and phenolic metabolites). Results: Among the clinical parameters studied, BP and fasting blood glucose significantly decreased (p < 0.05) after the seasoning intervention, but not for the control group. Notably, application of a novel approach based on ASV (Amplicon Sequence Variant) co-occurrence networks allowed us to identify some bacterial communities whose relative abundances were related with metadata. Conclusion: Our primary findings suggest that GP-seasoning may help in the modulation of cardiometabolic risk factors, mainly in the early stages. Furthermore, it evidences modulation of gut microbiota and functional bacterial communities by grape pomace, which might mediate the cardiometabolic effects of this by-product.

A global microbiome survey of vineyard soils highlights the microbial dimension of viticultural terroirs.

The microbial biodiversity found in different vitivinicultural regions is an important determinant of wine terroir. It should be studied and preserved, although it may, in the future, be subjected to manipulation by precision agriculture and oenology. Here, we conducted a global survey of vineyards' soil microbial communities. We analysed soil samples from 200 vineyards on four continents to establish the basis for the development of a vineyard soil microbiome's map, representing microbial biogeographical patterns on a global scale. This study describes vineyard microbial communities worldwide and establishes links between vineyard locations and microbial biodiversity on different scales: between continents, countries, and between different regions within the same country. Climate data correlates with fungal alpha diversity but not with prokaryotes alpha diversity, while spatial distance, on a global and national scale, is the main variable explaining beta-diversity in fungal and prokaryotes communities. Proteobacteria, Actinobacteria and Acidobacteria phyla, and Archaea genus Nitrososphaera dominate prokaryotic communities in soil samples while the overall fungal community is dominated by the genera Solicoccozyma, Mortierella and Alternaria. Finally, we used microbiome data to develop a predictive model based on random forest analyses to discriminate between microbial patterns and to predict the geographical source of the samples with reasonable precision.

Niche differentiation drives microbial community assembly and succession in full-scale activated sludge bioreactors.

Network models and community phylogenetic analyses are applied to assess the composition, structure, and ecological assembly mechanisms of microbial communities. Here we combine both approaches to investigate the temporal dynamics of network properties in individual samples of two activated sludge systems at different adaptation stages. At initial assembly stages, we observed microbial communities adapting to activated sludge, with an increase in network modularity and co-exclusion proportion, and a decrease in network clustering, here interpreted as a consequence of niche specialization. The selective pressure of deterministic factors at wastewater treatment plants produces this trend and maintains the structure of highly functional and specialized communities responding to seasonal environmental changes.

Local Network Properties of Soil and Rhizosphere Microbial Communities in Potato Plantations Treated with a Biological Product Are Important Predictors of Crop Yield.

Understanding the effectiveness and potential mechanism of action of agricultural biological products under different soil profiles and crops will allow more precise product recommendations based on local conditions and will ultimately result in increased crop yield. This study aimed to use bulk soil and rhizosphere microbial composition and structure to evaluate the potential effect of a Bacillus amyloliquefaciens inoculant (strain QST713) on potatoes and to explore its relationship with crop yield. We implemented next-generation sequencing (NGS) and bioinformatics approaches to assess the bacterial and fungal biodiversity in 185 soil samples, distributed over four different time points-from planting to harvest-from three different geographical locations in the United States. In addition to location and sampling time (which includes the difference between bulk soil and rhizosphere) as the main variables defining the microbiome composition, the microbial inoculant applied as a treatment also had a small but significant effect in fungal communities and a marginally significant effect in bacterial communities. However, treatment preserved the native communities without causing a detectable long-lasting effect on the alpha- and beta-diversity patterns after harvest. Using information about the application of the microbial inoculant and considering microbiome composition and structure data, we were able to train a Random Forest model to estimate if a bulk soil or rhizosphere sample came from a low- or high-yield block with relatively high accuracy (84.6%), concluding that the structure of fungal communities gives us more information as an estimator of potato yield than the structure of bacterial communities. IMPORTANCE Our results reinforce the notion that each cultivar on each location recruits a unique microbial community and that these communities are modulated by the vegetative growth stage of the plant. Moreover, inoculation of a Bacillus amyloliquefaciens strain QST713-based product on potatoes also changed the abundance of specific taxonomic groups and the structure of local networks in those locations where the product caused an increase in the yield. The data obtained, from in-field assays, allowed training a predictive model to estimate the yield of a certain block, identifying microbiome variables-especially those related to microbial community structure-even with a higher predictive power than the geographical location of the block (that is, the principal determinant of microbial beta-diversity). The methods described here can be replicated to fit new models in any other crop and to evaluate the effect of any agricultural input in the composition and structure of the soil microbiome.

Network Properties of Local Fungal Communities Reveal the Anthropogenic Disturbance Consequences of Farming Practices in Vineyard Soils.

Agroecosystems are human-managed ecosystems subject to generalized ecological rules. Understanding the ecology behind the assembly and dynamics of soil fungal communities is a fruitful way to improve management practices and plant productivity. Thus, monitoring soil health would benefit from the use of metrics that arise from ecological explanations that can also be informative for agricultural management. Beyond traditional biodiversity descriptors, community-level properties have the potential of informing about particular ecological situations. Here we assess the impact of different farming practices in a survey of 350 vineyard soils from the United States and Spain by estimating network properties based on spatial associations. Our observations using traditional approaches show results concurring with previous literature: the influence of geographic and climatic factors on sample distributions, or different operational taxonomic unit (OTU) compositions depending on agricultural managements. Furthermore, using network properties, we observe that fungal communities ranged from dense arrangements of associations to a sparser structure of associations, indicating differential levels of niche specialization. We detect fungal arrangements capable of thriving in wider or smaller ranges of temperature, revealing that niche specialization may be a critical soil process impacting soil health. Low-intervention practices (organic and biodynamic managements) promoted densely clustered networks, describing an equilibrium state based on mixed collaborative communities. In contrast, conventionally managed vineyards had highly modular sparser communities, supported by a higher coexclusion proportion. Thus, we hypothesize that network properties at the community level may help to understand how the environment and land use can affect community structure and ecological processes in agroecosystems.IMPORTANCE Soil fungal communities play a key role in agroecosystem sustainability. The complexity of fungal communities, at both taxonomic and functional levels, makes it difficult to find clear patterns connecting community composition with ecosystem function and to understand the impact of biotic (interspecies interactions) and abiotic (e.g., climate or anthropogenic disturbances) factors on it. Here we combine network analysis methods and properties, proposing a novel analytical approach: to infer ecological properties from local networks, which we apply to the study of fungal communities in vineyard soils. We conclude that different levels of farming intensification may lead to different ecological strategies in soil fungal communities settled by particular association arrangements.

A multi-omics approach for understanding the effects of moderate wine consumption on human intestinal health.

The human gut is a highly diverse microbial ecosystem. Although showing a well-defined core of dominant taxa, an interindividual variability exists in microbiome arrangement patterns, and the presence and proportion of specific species, determining individual metabolic features-metabotypes-which govern the health effects of dietary interventions (i.e. polyphenol consumption). Starting with a 19-volunteer human intervention study, divided into low, medium, and high wine-polyphenol-metabolizers, we detected interindividual discrepancies on the effect of wine consumption in gut bacterial alpha-diversity, but a significant homogenization of beta-diversity among moderate wine consumers, independently of their metabotype. In addition, the abundance of key health-related taxa such as Akkermansia sp. increased after moderate wine intake in the group of high polyphenol-metabolizers. Regarding the metabolic activity, significant (p < 0.05) positive correlations in the production of SCFAs were observed after wine intake. Finally, we were able to correlate the microbiome and the metabolome of the three metabotypes, and to identify some metabolites-biomarker species, highlighting the genera Phascolarctobacterium, Pelotomaculum and Prevotella, as positively correlated with polyphenol concentration, and Prevotella, Zymophilus and Eubacterium as positively correlated with SCFAs concentration in faeces. Our results contribute to the evidence of the need of including the microbiome variable in personalized nutrition programs, as different metabotyes respond differently to dietary interventions.

The Genus Metschnikowia in Enology.

Over the last decade, several non-Saccharomyces species have been used as an alternative yeast for producing wines with sensorial properties that are distinctive in comparison to those produced using only Saccharomyces cerevisiae as the classical inoculum. Among the non-Saccharomyces wine yeasts, Metschnikowia is one of the most investigated genera due to its widespread occurrence and its impact in winemaking, and it has been found in grapevine phyllospheres, fruit flies, grapes, and wine fermentations as being part of the resident microbiota of wineries and wine-making equipment. The versatility that allows some Metschnikowia species to be used for winemaking relies on an ability to grow in combination with other yeast species, such as S. cerevisiae, during the first stages of wine fermentation, thereby modulating the synthesis of secondary metabolites during fermentation in order to improve the sensory profile of the wine. Metschnikowia exerts a moderate fermentation power, some interesting enzymatic activities involving aromatic and color precursors, and potential antimicrobial activity against spoilage yeasts and fungi, resulting in this yeast being considered an interesting tool for use in the improvement of wine quality. The abovementioned properties have mostly been determined from studies on Metschnikowia pulcherrima wine strains. However, M. fructicola and M. viticola have also recently been studied for winemaking purposes.