Wine Barrel Biofilm as a Source of Yeasts with Non-Conventional Properties.

This study investigated the main microbial groups characterizing the interior surface of oak barrels from different years (1890, 1895, 1920, 1975, 2008) used in the production of vino cotto. The yeasts were characterized for the following properties: γ-aminobutyric acid (GABA) production, antioxidant activity, air-liquid interfacial biofilm formation, and anthocyanin adsorption capacity. Community-level physiological profile analysis revealed that the microbial communities inside the barrels used the tested carbon sources in different manners. The following yeast species were identified: Millerozyma farinosa, Zygosaccharomyces bisporus, Wickerhamiella versatilis, Zygosaccharomyces bailii, Starmerella lactis-condensi, and Zygosaccharomyces rouxii. All the strains were able to produce GABA, and S. lactis-condensi, Z. bisporus and Z. rouxii were the highest producers (more than 600 mg/L). The Z. rouxii and Z. bailii strains showed the highest antioxidant activity. Only seven strains out of ten M. farinosa formed air-liquid interfacial biofilm. None of the M. farinosa strains adsorbed anthocyanins on their cell wall. The other strains adsorbed anthocyanins in a strain-dependent way, and the highest adsorption was observed for the W. versatilis strains. The yeasts isolated in this study could be used to increase the functional properties and the quality of fermented foods and beverages.

Contribution of Starmerella bacillaris and Oak Chips to Trebbiano d'Abruzzo Wine Volatile and Sensory Diversity.

In this study, six fermentation trials were carried out: co-inoculation and sequential inoculation of Saccharomyces cerevisiae and Starmerella bacillaris in the presence and absence of oak chips. Moreover, Starm. bacillaris strain was attached to the oak chips and co-inoculated or sequentially inoculated with S. cerevisiae. Wines fermented with Starm. bacillaris adhered to oak chips showed a higher concentration of glycerol (more than 6 g/L) than the others (about 5 g/L). These wines also showed a higher content of polyphenols (more than 300 g/L) than the others (about 200 g/L). The addition of oak chips induced an increase of yellow color (b* value of about 3). Oak-treated wines were characterized by a higher concentration of higher alcohols, esters and terpenes. Aldehydes, phenols and lactones were detected only in these wines, independently from the inoculation strategy. Significant differences (p < 0.05) were also observed in the sensory profiles. The fruity, toasty, astringency, and vanilla sensations were perceived as more intense in wines treated with oak chips. The white flower descriptor showed a higher score in wines fermented without chips. Oak surface-adhered Starm. bacillaris cells could be a good strategy to improve the volatile and sensory profile of Trebbiano d'Abruzzo wines.

Influence of Different Aggregation States on Volatile Organic Compounds Released by Dairy Kluyveromyces marxianus Strains.

Kluyveromyces marxianus has the ability to contribute to the aroma profile of foods and beverages since it is able to produce several volatile organic compounds (VOCs). In this study, 8 dairy K. marxianus strains, previously selected for their adhesion properties, were tested for VOCs production when grown in different conditions: planktonic, biofilm-detached, and MATS forming-cells. It was shown that biofilm-detached cells were mainly able to produce higher alcohols (64.57 mg/L), while esters were mainly produced by planktonic and MATS forming-cells (117.86 and 94.90 mg/L, respectively). Moreover, K. marxianus biofilm-detached cells were able to produce VOCs with flavor and odor impacts, such as ketons, phenols, and terpenes, which were not produced by planktonic cells. In addition, specific unique compounds were associated to the different conditions tested. Biofilm-detached cells were characterized by the production of 9 unique compounds, while planktonic and MATS forming-cells by 7 and 12, respectively. The obtained results should be exploited to modulate the volatilome of foods and beverages and improve the production of certain compounds at the industrial level. Further studies will be carried out to better understand the genetic mechanisms underlying the metabolic pathways activated under different conditions.

Impact of Saccharomyces cerevisiae and non-Saccharomyces yeasts to improve traditional sparkling wines production.

In this study the effect of a co-inoculum of S. cerevisiae (F6789) with Torulaspora delbrueckii (TB1) or Starmerella bacillaris (SB48) on the oenological and aroma characteristics of sparkling wines obtained with the Champenoise method was investigated. The autolytic outcome and the sensory profile of sparkling wines were also evaluated. The secondary fermentations were completed by all mixed and single starter cultures with the only exception of those guided by Starm.bacillaris. Sparkling wines produced with S. cerevisiae F6789+Starm.bacillaris SB48 showed the highest amounts of glycerol (6.51 g/L). The best autolytic potential was observed in sparkling wines produced with +Starm.bacillaris (81.98 mg leucin/L) and S. cerevisiae+T. delbrueckii (79.03 mg leucin/L). The lowest value was observed for sparkling wines obtained with S. cerevisiae F6789 (53.96 mg leucin/L). Sparkling wines showed different aroma and sensory profiles. Esters were mainly present in sparkling wines obtained with S. cerevisiae F6789 (88.09 mg/L) followed by those obtained with S. cerevisiae+T. delbrueckii (87.20 mg/L), S. cerevisiae +Starm.bacillaris (81.93 mg/L). The content of esters decreased over time, and that might be related to the adsorption on lees and chemical hydrolysis. The highest concentrations of higher alcohols were found in sparkling wines produced with S. cerevisiae+T. delbrueckii (27.50 mg/L). Sparkling wines obtained with S. cerevisiae +Starm.bacillaris were well differentiated from the others due to their high score for the descriptor for spicy, bread crust, freshness and floral. Tailored strains with different autolytic potential might represent an interesting strategy to improve traditional sparkling wine production and favour their differentiation.

Impact of vineyard management on grape fungal community and Montepulciano d'Abruzzo wine quality.

In this study the effect of farming practices on the diversity of grape fungal community of Vitis vinifera L. cultivar Montepulciano and the microbial metabolic activity was investigated. Small-scale vinifications were performed and wines were characterized. Agronomic management system affected the fungal community composition. Hanseniaspora, Areobasidium and Botrytis genera represented 30%, 20% and 10% of the total reads in all samples. A. pullulans, and Cladosprium cladosporioides mainly occurred on organic and biodynamic grapes. Saccharomyces and Pseudopithomyces genera were present only on organic or biodynamic grapes, respectively. The agronomic managements also influenced the potential functionality of microbial community. In fact, the metabolic function was increased in organic and biodynamic grapes. Polymers were used only by organic and biodynamic microbial communities, which also showed the highest values of Shannon's diversity index, and substrate richness. However, no significative differences were observed for the oenological parameters analyzed, with the only exception of the content of sugars which were higher on conventional grapes. The fermentation profiles showed that conventional wines had a lower residual sugars content, and a higher amount of alcohol. The differences observed in the volatile composition of the wines were both quantitative and qualitative. Conventional wines showed a lower content of esters, and a higher concentration of alcohols than organic and biodynamic ones. Biodynamic wines were characterized by the highest content of organic acids. The obtained results revealed that farming practices shape the fungal community influencing wine traits linking the wine with the viti-vinicultural area of origin.

Discovering the Influence of Microorganisms on Wine Color.

Flavor, composition and quality of wine are influenced by microorganisms present on the grapevine surface which are transferred to the must during vinification. The microbiota is highly variable with a prevalence of non-Saccharomyces yeasts, whereas Saccharomyces cerevisiae is present at low number. For wine production an essential step is the fermentation carried out by different starter cultures of S. cerevisiae alone or in mixed fermentation with non-Saccharomyces species that produce wines with significant differences in chemical composition. During vinification wine color can be influenced by yeasts interacting with anthocyanin. Yeasts can influence wine phenolic composition in different manners: direct interactions-cell wall adsorption or enzyme activities-and/or indirectly-production of primary and secondary metabolites and fermentation products. Some of these characteristics are heritable trait in yeast and/or can be strain dependent. For this reason, the stability, aroma, and color of wines depend on strain/strains used during must fermentation. Saccharomyces cerevisiae or non-Saccharomyces can produce metabolites reacting with anthocyanins and favor the formation of vitisin A and B type pyranoanthocyanins, contributing to color stability. In addition, yeasts affect the intensity and tonality of wine color by the action of ß-glycosidase on anthocyanins or anthocyanidase enzymes or by the pigments adsorption on the yeast cell wall. These activities are strain dependent and are characterized by a great inter-species variability. Therefore, they should be considered a target for yeast strain selection and considered during the development of tailored mixed fermentations to improve wine production. In addition, some lactic acid bacteria seem to influence the color of red wines affecting anthocyanins' profile. In fact, the increase of the pH or the ability to degrade pyruvic acid and acetaldehyde, as well as anthocyanin adsorption by bacterial cells are responsible for color loss during malolactic fermentation. Lactic acid bacteria show different adsorption capacity probably because of the variable composition of the cell walls. The aim of this review is to offer a critical overview of the roles played by wine microorganisms in the definition of intensity and tonality of wines' color.

Promoting Candida zemplinina adhesion on oak chips: A strategy to enhance esters and glycerol content of Montepulciano d'Abruzzo organic wines.

In this study cell surface hydrophobicity and the ability to adhere on abiotic surfaces (polystyrene plates, stainless steel and oak chips) of 10 Candida zemplinina strains were assessed. Moreover, the impact of C. zemplinina cells adhered on oak surface on fermentation kinetics and volatile profile of Montepulciano d'Abruzzo organic wines was evaluated. All strains showed a hydrophobic nature with a certain affinity for the apolar solvents tested (hexadecane and decane). In agreement with this data strains were able to adhere on abiotic surfaces in a strain dependent way. On polystyrene plates all strains mainly grew as planktonic cells. On stainless steel surfaces sessile cells ranged from 2.6 Log CFU/mL (SB2) to 4.1 Log CFU/mL (SB8), while on oak chips were about 2 log higher ranging from 4.3 Log CFU/mL (SB8) to 6.1 Log CFU/mL (SB10). Candida zemplinina sessile state resulted in an increase of glycerol (from 6.98 g/L to 11.92 g/L) and esters amount (from 55.47 g/L to 91.5 mg/L), and a reduction of ethanol content (from 14.13% to 9.12% v/v). As for esters, methyl vanillate, ethyl isobutyrate, and ethyl isovalerate were present only when C. zemplinina was adhered on oak chips. This study revealed that changes of concentrations in esters and glycerol content reflected the fermentation bioactivity of yeast cells attached on oak chips. Surface-adhered behaviours should be considered in the improvement of strategies for the development of high-quality organic wines and eventually obtain novel wine styles.

Oenococcus oeni Lifestyle Modulates Wine Volatilome and Malolactic Fermentation Outcome.

In this study, nine Oenococcus oeni strains were tested for their ability to adhere to polystyrene using mMRS and wine as culture media. Moreover, planktonic and biofilm-detached cells were investigated for their influence on malic acid degradation kinetics and aroma compound production. Three strains were able to adhere on polystyrene plates in a strain-dependent way. In particular, MALOBACT-T1 and ISO359 strains mainly grew as planktonic cells, while the ISO360 strain was found prevalent in sessile state. The strain-dependent adhesion ability was confirmed by confocal laser scanning microscopy. Planktonic and biofilm detached cells showed a different metabolism. In fact, biofilm-detached cells had a better malic acid degradation kinetic and influenced the aroma composition of resulting wines, acting on the final concentration of esters, higher alcohols, and organic acids. Oenococcus oeni in biofilm lifestyle seems to be a suitable tool to improve malolactic fermentation outcome, and to contribute to wine aroma. The industrial-scale application of this strategy should be implemented to develop novel wine styles.

Lactic Acid Bacteria Exopolysaccharides Producers: A Sustainable Tool for Functional Foods.

Lactic acid bacteria (LAB) used in the food industry, mainly for the production of dairy products, are able to synthetize exopolysaccharides (EPS). EPS play a central role in the assessment of rheological and sensory characteristics of dairy products since they positively influence texture and organoleptic properties. Besides these, EPS have gained relevant interest for pharmacological and nutraceutical applications due to their biocompatibility, non-toxicity and biodegradability. These bioactive compounds may act as antioxidant, cholesterol-lowering, antimicrobial and prebiotic agents. This review provides an overview of exopolysaccharide-producing LAB, with an insight on the factors affecting EPS production, their dairy industrial applications and health benefits.

Cultivable microbial ecology and aromatic profile of "mothers" for Vino cotto wine production.

The aim of the present study was to assess the cultivable microbiota of "mothers" of Vino cotto collected from production of different years 1890, 1895, 1920, 1975, 2008. A total of 73 yeasts and 81 bacteria were isolated. Starmerella lactis-condensi, Starmerella bacillaris, Hanseniaspora uvarum, Saccharomyces cerevisiae, Hanseniaspora guillermondi and Metschnikowia pulcherrima were identified. Bacteria isolates belonged to lactic acid bacteria (Lactiplantibacillus plantarum and Pediococcus pentosaceus) and acetic acid bacteria (Gluconobacter oxydans). Remarkable biodiversity was observed for Starm. bacillaris, as well as L. plantarum and G. oxydans. Organic acids and volatile compounds were also determined. Malic and succinic acids were the main ones with values ranging from 8.49 g/L to 11.76 g/L and from 4.15 g/L to 7.73 g/L respectively, while citric acid was present at low concentrations (<0.2 g/L) in all samples. Esters and higher alcohols were the main volatile compounds detected followed by alkanes. This study permits to better understand the microbial communities associated to this product and could be considered a starting point for the definition of tailored starter cultures to improve the quality of Vino cotto preserving its typical traits.

Adhesion Properties, Biofilm Forming Potential, and Susceptibility to Disinfectants of Contaminant Wine Yeasts.

In this study, yeasts isolated from filter membranes used for the quality control of bottled wines were identified and tested for their resistance to some cleaning agents and potassium metabisulphite, adhesion to polystyrene and stainless-steel surfaces, and formation of a thin round biofilm, referred to as a MAT. A total of 40 strains were identified by rRNA internal transcribed spacer (ITS) restriction analysis and sequence analysis of D1/D2 domain of 26S rRNA gene. Strains belong to Pichia manshurica (12), Pichia kudriavzevii (9), Pichia membranifaciens (1), Candida sojae (6), Candida parapsilosis (3), Candida sonorensis (1), Lodderomyces elongisporus (2), Sporopachydermia lactativora (3), and Clavispora lusitaniae (3) species. Regarding the adhesion properties, differences were observed among species. Yeasts preferred planktonic state when tested on polystyrene plates. On stainless-steel supports, adhered cells reached values of about 6 log CFU/mL. MAT structures were formed only by yeasts belonging to the Pichia genus. Yeast species showed different resistance to sanitizers, with peracetic acid being the most effective and active at low concentrations, with minimum inhibitory concentration (MIC) values ranging from 0.08% (v/v) to 1% (v/v). C. parapsilosis was the most sensible species. Data could be exploited to develop sustainable strategies to reduce wine contamination and establish tailored sanitizing procedures.

Correlation between IRC7 gene expression and 4-mercapto-4-methylpentan-2-one production in Saccharomyces cerevisiae strains.

Volatile thiols are not present in must but are synthesized and released by wine yeasts during alcoholic fermentation. In this study, autochthonous and commercial Saccharomyces cerevisiae strains were characterized for the expression of the main genes involved in thiols metabolism and their production in wine. New primer sets were developed on the basis of the S288c genome to evaluate the expression of Cys3, Cys4, MET17 and IRC7 genes. Obtained data revealed the occurrence of some thiols, for example, 4-mercapto-4-methylpentan-2-one (4-MMP) and 3-mercaptohexan-1-ol (3-MH) in Pecorino white wine. All genes were upregulated, but only for IRC7 was found a correlation with 4-MMP release: strains with the highest production showed the highest transcription level. IRC7 gene could be proposed as target for the selection of S. cerevisiae strains to increase thiols content in wine.

Corrigendum: Changes in Polyphenolic Concentrations of Table Olives (cv. Itrana) Produced Under Different Irrigation Regimes During Spontaneous or Inoculated Fermentation.

[This corrects the article DOI: 10.3389/fmicb.2018.01287.].

Table Olives More than a Fermented Food.

Table olives are one of the oldest vegetable fermented foods in the Mediterranean area. Beside their economic impact, fermented table olives represent also an important healthy food in the Mediterranean diet, because of their high content of bioactive and health-promoting compounds. However, olive fermentation is still craft-based following traditional processes, which can lead to a not fully predictable final product with the risk of spontaneous alterations. Nowadays, food industries have to face consumer demands for safe and healthy products. This review offers an overview about the main technologies used for olive fermentation and the role of lactic acid bacteria and yeasts characterizing this niche during the fermentation. Particular attention is offered to the selection and use of microorganisms as starter cultures to fasten and improve the safety of table olives. The development and implementation of multifunctional starter cultures in order to obtain heath-oriented table olives is also discussed.

Characterization of Bifidobacterium species in feaces of the Egyptian fruit bat: Description of B. vespertilionis sp. nov. and B. rousetti sp. nov.

Fifteen bifidobacterial strains were obtained from faeces of Rousettus aegyptiacus; after grouping them by RAPD PCR only eight were selected and characterized. Analysis of 16S rRNA and of five housekeeping (hsp60, rpoB, clpC, dnaJ, dna G) genes revealed that these eight strains were classified into five clusters: Cluster I (RST 8 and RST 16T), Cluster II (RST 9T and RST 27), Cluster III (RST 7 and RST 11), Cluster IV (RST 19), Cluster V (RST 17) were closest to Bifidobacterium avesanii DSM 100685T (96.3%), Bifidobacterium callitrichos DSM 23973T (99.2% and 99.7%), Bifidobacterium tissieri DSM 100201T (99.7 and 99.2%), Bifidobacterium reuteri DSM 23975 T (98.9%) and Bifidobacterium myosotis DSM 100196T (99.3%), respectively. Strains in Cluster I and strain RST 9 in Cluster II could not be placed within any recognized species while the other ones were identified as known species. The average nucleotide identity values between two novel strains, RST 16T and RST 9T and their closest relatives were lower than 79% and 89%, respectively. In silico DNA-DNA hybridization values for those closest relatives were 32.5 and 42.1%, respectively. Phenotypic and genotypic tests demonstrated that strains in Cluster I and RST 9T in Cluster II represent two novel species for which the names Bifidobacterium vespertilionis sp. nov. (RST 16T=BCRC 81138T=NBRC 113380T=DSM 106025T ; RST 8=BCRC 81135=NBRC 113377) and Bifidobacterium rousetti sp. nov. (RST 9T=BCRC 81136T=NBRC 113378T=DSM 106027T) are proposed.

Accumulation γ-Aminobutyric Acid and Biogenic Amines in a Traditional Raw Milk Ewe's Cheese.

The influence of calf (R1), kid (R2) and pig (R3) rennets on microbiota, biogenic amines (BAs) and γ-aminobutyric acid (GABA) accumulation in raw milk ewe's cheeses was evaluated. Cheeses were investigated at different ripening times for their microbial composition, free amino acids (FAAs), BAs and GABA content. Moreover, the expression of tyrosine (tdc) and histidine (hdc) decarboxylases genes was evaluated by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Microbial counts showed similar values in all samples. Pig rennet were cheeses were characterized by higher proteolysis and the highest values of BAs. The BAs detected were putrescine, cadaverine and tyramine, while histamine was absent. qRT-PCR confirmed this data, in fact hdc gene was not upregulated, while tdc gene expression increased over time in agreement with the increasing content of tyramine and the highest fold changes were detected in R3 cheeses. GABA showed the highest concentration in R2 cheeses reaching a value of 672 mg/kg. These results showed that the accumulation of BAs and GABA in Pecorino di Farindola is influenced by ripening time and type of coagulant. Further studies are required to develop starter cultures to reduce BAs content and improve health characteristics of raw milk ewe's cheeses.

Cell Wall Surface Properties of Kluyveromyces marxianus Strains From Dairy-Products.

Thirty-three Kluyveromyces marxianus strains were tested for the ability to form biofilm and mat structures in YPD and whey and for cell surface hydrophobicity. To identify genes potentially involved in adhesion properties, a RT-qPCR analysis was performed. Eight strains were able to adhere on polystyrene plates in both media and formed a mature mat structure. These strains showed a different level of hydrophobicity ranging from 55 to 66% in YPD and from 69 to 81% in whey. Four K. marxianus orthologs genes (FLO11, STE12, TPK3, and WSC4), known from studies in other yeast to be involved in biofilm formation, have been studied. FLO11 and STE12 showed the highest fold changes in all conditions tested and especially in whey: 15.05 and 11.21, respectively. TPK3 was upregulated only in a strain, and WSC4 in 3 strains. In YPD, fold changes were lower than in whey with STE12 and FLO11 genes showing the highest fold changes. In mat structures FLO11 and STE12 fold changes ranged from 3.6-1.3 to 2-1.17, respectively. Further studies are necessary to better understand the role of these genes in K. marxianus adhesion ability.

Chemical-nutritional quality and oxidative stability of milk and dairy products obtained from Friesian cows fed with a dietary supplementation of dried grape pomace.

BACKGROUND: The aim of the study was to evaluate the nutritional properties of milk and cheese obtained from Friesian cows fed with a diet supplemented with dried grape pomace, a by-product of the oenological industry, which is very rich in polyphenols. This approach is inspired by the increasing interest in foods containing functional ingredients that may have beneficial effects on human health. During the testing period, analyses of the chemical and nutritional properties of milk and dairy products derived from it were performed; particular attention was given to the effect of cheese ripening on the oxidative stability and fermentation process, evaluating respectively the presence of malondialdehyde and γ-aminobutyric acid. RESULTS: Dietary enrichment with grape pomace did not affect the milk composition but induced modifications in the fatty acid profiles in both milk and cheese with an increase in concentration of linoleic acid, trans-vaccenic acid, rumenic acid and total n-6 fatty acids. Moreover, after 30 days of cheese ripening, an increased oxidative stability and an increased concentration of γ-aminobutyric acid were found. CONCLUSIONS: Our results indicated a general improvement in nutritional parameters of milk and related cheese obtained from Friesian cows that received the feeding enrichment with dried grape pomace. Further analysis should be performed to improve knowledge of the chemical and microbiological mechanisms at the source of these findings. © 2019 Society of Chemical Industry.