Physiological requirements for growth and competitiveness of Dekkera bruxellensis under oxygen-limited or anaerobic conditions

The effect of glucose and oxygen limitation on the growth and fermentation performances of Dekkera bruxellensis was investigated in order to understand which factors favour its propagation in ethanol or wine plants. Although D. bruxellensis has been described as a facultative anaerobe, no growth was observed in mineral medium under complete anaerobiosis while growth was retarded under severe oxygen limitation. In a continuous culture with no gas inflow, glucose was not completely consumed, most probably due to oxygen limitation. When an air/nitrogen mixture (O₂‐content ca. 5%) was sparged to the culture, growth became glucose‐limited. In co‐cultivations with Saccharomyces cerevisiae, ethanol yields/g consumed sugar were not affected by the co‐cultures as compared to the pure cultures. However, different population responses were observed in both systems. In oxygen‐limited cultivation, glucose was depleted within 24 h after challenging with S. cerevisiae and both yeast populations were maintained at a stable level. In contrast, the S. cerevisiae population constantly decreased to about 1% of its initial cell number in the sparged glucose‐limited fermentation, whereas the D. bruxellensis population remained constant. To identify the requirements of D. bruxellensis for anaerobic growth, the yeast was cultivated in several nitrogen sources and with the addition of amino acids. Yeast extract and most of the supplied amino acids supported anaerobic growth, which points towards a higher nutrient demand for D. bruxellensis compared to S. cerevisiae in anaerobic conditions. Copyright © 2012 John Wiley & Sons, Ltd.     

Modulation of grape wine flavor via the sequential inoculation of Williopsis saturnus and Saccharomyces cerevisiae

The effects of the inoculum ratio of Williopsis saturnus var. saturnus NCYC22 and Saccharomyces cerevisiae var. bayanus EC-1118 at 1:200 and 1:800 on the chemical and volatile compositions of grape wine were studied in sequential fermentation. The grape juice was first inoculated with Williopsis (W.) saturnus for 9 d; thereafter, Saccharomyces (S.) cerevisiae was inoculated to continue the fermentation until d 19. The cell population of W. saturnus disappeared by d 13, with S. cerevisiae dominating until the end of the fermentation in both inoculum ratios. The changes in yeast count, pH, total soluble solids, sugars, organic acids, and amino acids were similar between the two inoculum ratios. A range of volatile compounds was formed, including alcohols, esters, fatty acids, aldehydes, and terpenes. There were significant differences between both inoculum ratios for medium-chain fatty acids (C8, C10, and C12), ethyl esters of fatty acids of C6, C10, C12, and C14 as well as isoamyl octanoate, while other volatiles were statistically the same.     

Contribution of wild yeasts to the formation of volatile compounds in inoculated wine fermentations

The aim of this work was to study the contribution of wild yeasts to the volatile composition of wine in inoculated fermentations. To do so, Parellada must, sterilized and inoculated with Saccharomyces cerevisiae strain Na33 (pure inoculated fermentation), inoculated Parellada must (mixed inoculated fermentation) and Parellada must that fermented with its wild yeasts (control fermentation) were used. From the results obtained in the pure inoculated fermentation it can be seen that S. cerevisiae produced appreciable quantities of isoamyl acetate, ethyl hexanoate, ethyl octanoate, and ethyl decanoate. However, the wild yeasts also contributed to the synthesis of esters since the total concentration of these substances was higher in the mixed inoculated fermentation than in the pure inoculated fermentation. 2-Phenylethyl acetate was only synthesized by wild yeasts when they did not compete with S. cerevisiae. The concentration of total alcohols was similar in the three samples; the important production of isobutanol and 2-phenylethanol in the control fermentation is noteworthy. As regards the acids, the greatest concentration corresponded to the mixed inoculated fermentation. The wild yeasts contributed to the synthesis of these compounds to a significant extent and S. cerevisiae synthesized appreciable amounts of short-chain fatty acids.     

Effects of mediums on fermentation behaviour and aroma composition in pure and mixed culture of Saccharomyces cerevisiae with Torulaspora delbrueckii

The nutrient status and composition in mediums have a significant effect on yeast metabolism and phenotypic characteristics in wine fermentation. In this study, the effects of three frequently used mediums, including synthetic grape juice (SGJ), grape juice without grape pericarp and seeds (GJ) and grape must with grape pericarp and seeds (GMPS), on yeast fermentation behaviour and aroma compounds produced by pure and mixed culture of Saccharomyces cerevisiae T73 with Torulaspora delbrueckii TD20 were investigated after alcoholic fermentation. The results showed that high fermentation activities and cell population were always found in GJ medium irrespective of inoculated approach. More esters and higher alcohols were produced in GMPS medium fermented by pure S. cerevisiae, while SGJ medium had increased levels of fatty acids. Consistent with previous literatures, the mixed fermentation of T. delbrueckii and S. cerevisiae produced more acetate esters and fatty acids than the pure culture of S. cerevisiae, while this enological trait was only found in SGJ and GJ, not in GMPS. Our results highlighted that more attention should be paid to the fermentation medium when evaluating the enological and aromatic properties of selected yeasts used in industrial winemaking. In this regard, the combined use of GJ and GMPS medium might be a suitable choice.     

Fermentation Characteristics and Aromatic Profile of Plum Wines Produced with Indigenous Microbiota and Pure Cultures of Selected Yeast

The aim of this study was to assess and compare fermentation characteristics and aromatic profile of plum wines produced with indigenous microbiota and pure cultures of different selected yeast. Experiments were carried out with plum (Prunus domestica L.) varieties of different fruit ripening times (?a?anska rana, ?a?anska lepotica, and Po?ega?a). Wine fermentations were conducted by the activity of indigenous microbiota, commercially available Saccharomyces cerevisiae and Saccharomyces bayanus yeast strains and joint activity of Schizosaccharomyces pombe and S. cerevisiae (sequential inoculation). Statistically significant differences in fermentative characteristics and the content of certain volatile compounds were observed as a result of metabolic activity of various indigenous and/or selected yeasts during fermentation of plum pomace. Minimal duration of fermentation (4 to 5 d) and fastest ethanol production rate (from 12.3 to 15.5 g/L/d) were the characteristics of the studied S. cerevisiae strains. Isobutanol, 3‐methyl‐1‐butanol, 1‐heptanol, and 1‐octanol were the most prevalent higher alcohols in the tested plum wine samples. The predominant ester in plum wines was ethyl acetate, ethyl lactate, amyl acetate, isoamyl acetate, and ethyl palmitate, esters responsible for the floral and fruity olfactory tones, were also present in large amounts. Also, the use of S. cerevisiae strains resulted in the production of plum wines with better sensory characteristics than ones produced with other investigated yeasts. Obtained results are significant since there is limited data on the compounds responsible for the unique flavor of plum wine, as well as on the impact of different yeast starter cultures application on the overall quality of fruit wines.     

Fermentation Process and Grain Structure of Baked Breads from Frozen Dough Using Freeze-Tolerant Yeasts

The fermentation process for frozen doughs using freeze‐sensitive (Saccharomyces cerevisiae, Kyowa for sweet bread; S. cerevisiae, FC for white bread) and freeze‐tolerant (S. cerevisiae, YF for sweet bread) yeasts was traced by magnetic resonance imaging (MRI). Grain network structures of baked breads were also visualized by MRI. Prefermentation before freezing, punching and remolding, or resheeting and molding treatments increased loaf volume by 10 to 110% for the baked breads using freeze‐tolerant yeast, while these treatments decreased loaf volume by 70% using freeze‐sensitive yeast. The first fermentation before freezing and the second fermentation with punching or resheeting after thawing are useful for obtaining good quality breads from frozen dough using freeze‐tolerant yeast.     

Alginate beads and apple pieces as carriers for Saccharomyces cerevisiae var. boulardii,as representative of yeast functional starter cultures

This study focused on two different carriers for Saccharomyces cerevisiae var. boulardii for its targeted release as a starter or as a probiotic: the microencapsulation into alginate beads and the immobilisation on apple pieces. Beads were studied in relation to encapsulation yield (EY), viability of cells throughout the storage, kinetic of cell release, survival under conditions that mimic the transit into the gut and in vivo application (fermentation of grape juice). Concerning the microencapsulation, EY was very high (ca. 90%), cells survived for a long period (upon to 90 days), and the beads assured cell survival and their release under conditions that mimic the gut; moreover, the capsules could be used up to ten times to start a model fermentation of grape juice. Apple pieces were a good system for the immobilisation of S. boulardii; they could be proposed successfully as a reusable carrier for a starter culture, as they assured the start of the fermentation of grape juice for at least seven times.     

Use of PCR‐DHPLC (Polymerase Chain Reaction—Denaturing High Performance Liquid Chromatography) for the Rapid Differentiation of Industrial Saccharomyces pastorianus and Saccharomyces cerevisiae Strains

Strain specific detection and control of Saccharomyces pastorianus and Saccharomyces cerevisiae starter cultures is of great importance for the fermentation industry. The preconditions of strain specific fermentation characteristics can be ensured by periodic analysis and confirmation of the strain identity. With regard to industrial S. pastorianus and S. cerevisiae strains and a focus on brewing strains, the differentiation methods most available are time‐consuming and not very discriminative. In this work PCR‐DHPLC analysis was investigated as a novel approach for the differentiation of industrially used S. pastorianus and S. cerevisiae strains. The PCR‐DHPLC‐system was specific for S. cerevisiae strains and S. pastorianus hybrid strains that contain IGS2 rDNA, which originates from the S. cerevisiae ancestor. For the DNA of 177 strains of 41 non‐target species, which are typical for beverage and fermentation surroundings, the absence of PCR‐amplificates could be confirmed by DHPLC analysis. It was shown that single strains of S. cerevisiae and S. pastorianus could be differentiated. A strain specific differentiation within the group of top‐fermenting Saccharomyces cerevisiae strains could also be performed. For the group of bottom fermenting S. pastorianus brewing strains, strain‐to‐strain specific differences in the DHPLC chromatograms could be observed which can be used to differentiate and to compare two single strains with each other, although the comparison of chromatograms of an unknown S. pastorianus strain with a set of known S. pastorianus chromatograms could only reveal tendencies towards grouping into types. The differential DHPLC chromatogram characteristics (fluorescence intensities, number of peaks/side‐peaks/peak‐shoulders) within S. pastorianus are present, but not as distinctive as for S. cerevisiae. Additionally PCR‐DHPLC has advantages compared to other differentiation methods, such as species specificity, speed (2.5 h for one sample) and precision with the described limits.     

类胡萝卜素降解对枸杞酒特征香气的影响

为探究类胡萝卜素降解后产生的降异戊二烯类化合物对枸杞酒特征香气的影响,采用一株可降解类胡萝卜素的库特氏菌进行实验。分别在枸杞渣中接种库特氏菌,枸杞清汁中接入工业酿酒酵母,发酵结束后混合;在枸杞汁中加入库特氏菌酶制剂,辅助工业酿酒酵母发酵;将高压灭菌的枸杞渣加入枸杞清汁后接种工业酿酒酵母发酵,并以正常发酵工艺酿制的枸杞酒作为对照。采用顶空固相微萃取及气相色谱质谱联用（GC-MS）法,对枸杞酒香气成分进行检测,并通过感官评分法评价3种降解类胡萝卜素方式对枸杞酒香气的影响。结果表明,库特氏菌酶制剂辅助发酵组枸杞酒香气和口感均为最佳,高压灭菌后发酵组澄清度最好。因此,在枸杞酒酿造过程中,可以采用库特氏菌酶制剂对枸杞汁进行降解处理,在此之前对枸杞渣可适度配合使用高压灭菌,从而改进枸杞酒的酿造工艺,提升枸杞酒香气。     

Composition of Saccharomyces cerevisiae strains in spontaneous fermentations of Pinot Noir and Chardonnay

There is a lack of knowledge about the composition of Saccharomyces cerevisiae strains in spontaneous fermentations of Pinot Noir and Chardonnay cultivars. The objectives were to determine the relative abundance of indigenous and commercial S. cerevisiae strains in spontaneous fermentations at three wineries from the two cultivars and to compare the composition of the S. cerevisiae strains between cultivars and wineries. Three fermentation vessels were sampled at three stages of fermentation for each cultivar at each winery. Isolates were identified to the strain level using seven microsatellite loci. Commercial S. cerevisiae strains were isolated at a frequency higher than that of the indigenous strains at each winery for both cultivars. The composition of S. cerevisiae strains was different for each cultivar and at each winery. Our results illustrate the clear influence of inoculated commercial active dry yeast strains on the composition of S. cerevisiae strains in spontaneous fermentations at wineries conducting both inoculated and spontaneous fermentations.     

Characteristics of wines fermented with different Saccharomyces cerevisiae strains isolated from the La Mancha region

One hundred and seventy-four strains of Saccharomyces cerevisiae from musts in fermentation were characterized using pulsed-field gel electrophoresis. Of these, 13 strains were selected for their good oenological properties and were used as inocula in laboratory-scale fermentation of an Airén must. Volatile compound and amino acid content of the resulting wines was analysed. Factorial principal component analysis of the data revealed differences between wines according to the strain used in the fermentation. Strains that produce more volatile compound and have an adecuated nitrogen assimilation are recommended to obtain quality wines.     

Yeasts Isolated from the Alcoholic Fermentation of Agave duranguensis During Mezcal Production

Mezcal is a spirit produced in some regions of México. In the state of Durango, mezcal is produced via traditional fermentation of the Agave duranguensis plant. To better understand traditional fermentation processes, it is necessary to know which yeast species are present in fermentations in different producer regions. The aim of this research was to study yeasts involved in traditional mezcal fermentation in Durango, México, and investigate the phylogeny of the native Saccharomyces cerevisiae strains involved in this process. The 5.8S-ITS genomic region was analyzed to identify strains present in the fermentation process samples in this study. To differentiate strains belonging to the genus Saccharomyces, different molecular techniques were used, including analysis of mitochondrial DNA and δ elements and sequencing of the 5.8S-ITS genomic region. Although a high diversity of microorganisms was found at the beginning of fermentation, Saccharomyces cerevisiae was the only yeast present at the end of fermentation in region I, while Torulaspora delbrueckii was found in a higher number than S. cerevisiae at the end of fermentation in the region II. Molecular techniques demonstrated that Saccharomyces cerevisiae isolated in Durango are phylogenetically independent from the strains isolated in other regions of Latin America and Europe.     

Immunization of the industrial fermentation starter culture strain of Saccharomyces cerevisiae to a contaminating killer toxin-producing Candida tropicalis

K3 killer trait was introduced into the fermentation starter strain of Saccharomyces cerevisiae BSP 1 in order to construct immune industrial strain that produces K3 type killer toxin and was resistant to Candida tropicalis (K⁺) contamination. Protoplasts of respiration-deficient Rho^o strain of S. cerevisiae NCYC 761 (K3) and S. cerevisiae BSP 1 were fused. The resulting respiration-competent hybrid with K3 type killer activity was selected on media containing a non-fermentable carbon source and by a killer zone assay in a plate test, respectively. The fusant was similar to the parent strain in its fermentation and sugar utilization patterns, growth rate, dough-raising properties and osmotolerance. The newly constructed S. cerevisiae BSP 1 (K3) inhibited the growth of C. tropicalis in a pH range from 3.5 to 5.0 and over a temperature range of 20–30°C.     

Comparative study on quality characteristics of pickled and fermented sturgeon (Acipenser sinensis) meat in retort cooking

Sturgeon (Acipenser sinensis) meat has not been fully utilised due to quality defects which were raised by Chinese sturgeon processing industry. This study was aimed to develop a high-quality sturgeon meat product and the effect of fermentation on qualities was studied. Results showed that fermentation with Saccharomyces cerevisiae could enhance ‘mushroom’, ‘fruity’ and ‘chocolate’ flavour by 1-octen-3-ol, ethyl acetate and 3-methyl-butanal; and enrich flavour through mild lipid oxidation and hydrolysis. In addition, fermentation could give an attractive colour with higher a*, b* and L* value. Texture properties were also improved with higher hardness, springiness, gumminess and chewiness. Higher free amino acids, TCA-soluble peptides and taste score in fermented sturgeon meat indicated better flavour and taste. The organoleptic evaluation suggested an overall satisfactory of wine-aroma sturgeon meat by fermentation with S. cerevisiae.     

Saccharomyces cerevisiae associated with the spontaneous fermentation of tequila agave juice

Saccharomyces cerevisiae dominates the spontaneous fermentation of blue agave juice. Because of the batch heterogeneity, the aim of this work was to determine the strain diversity of S. cerevisiae among fermentations. During January and February 2015, agave juice was sampled in triplicate from four sampling points at a tequila distillery. The heterogeneity of yeast strains and the production of carbon dioxide were assessed during fermentation, whereas the amount of ethanol produced was measured at the end of the process. The fermentation cycle times varied widely (9 to 25 days), as did fermentation efficiency (2.5–45.5%). Yeast isolates were identified at the species level by ITS‐5.8S rRNA restriction fragment length polymorphism and differentiated at the strain level by random amplified polymorphic DNA. A total of 199 isolates were obtained and identified as S. cerevisiae, showing 69 different random amplified polymorphic DNA profiles. There was no clear dominance of any strain during fermentation. However, two strains (P1 and P2) were detected in all fermentation samples, suggesting their residency in the distillery, despite the deep‐cleaning applied to the tanks after each fermentation batch. According to the RAPD profiles, the number of strains isolated from fermentation samples increased from 17 in January to 25 in February. © 2018 The Institute of Brewing & Distilling     

Variations of internal pH in typical Italian sourdough yeasts during co-fermentation with lactobacilli

The effects of organic acids (lactic and acetic) and extracellular pH (pHex) on the intracellular pH (pHi) of Saccharomyces cerevisiae and Candida milleri during co-fermentation with lactobacilli were investigated by using Fluorescence-Ratio-Imaging-Microscopy (FRIM). Yeasts were grown in a system that partially mimics sourdough composition, using individual fermentation and combinations with lactic acid bacteria. Fermentations were carried out at 25 °C for 22 h at an initial pH of 5.3. The two yeast species grew equally well during the co-fermentations with lactobacilli. Our results reveal large differences in pHi values between the two yeast species, primarily in relation with pHex changes, while the concentration of organic acids did not seem to affect the pHi. Moreover, the pHi of S. cerevisiae seemed to be affected by maltose consumption. The pH gradient (difference between internal and external pH) of S. cerevisiae remained rather constant, ranging from 2.0 to 2.5. C. milleri instead exhibited a higher pHi, that remained constant throughout the experiments and was unaffected by pHex and/or sugar consumption. Thus, the pH gradient of C. milleri varied much more than that of S. cerevisiae, ranging from 2.3 to 3.8. Our results suggest that the two yeast species have different pHi regulation mechanisms.     

Ethanolic fermentation from red ginseng extract using <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> and <Emphasis Type="Italic">Saccharomyces carlsbergensis</Emphasis>

The pH of red ginseng extracts fermented with Saccharomyces cerevisiae and Saccharomyces carlsbergensis decreased rapidly during 3 days of fermentation, with no further significant change thereafter. After 20 days of fermentation, a relatively small difference remained in the acidity of extracts fermented with S. cerevisiae (0.54%) and S. carlsbergensis (0.58%). Reducing sugar in the S. cerevisiae and S. carlsbergensis extracts decreased from 258.6 to 45.4 and 43.2 mg/mL glucose equivalents, respectively; and ethanol contents increased from 1.5% at day 0 to 16.0 and 15.0%, respectively, at day 20. Ginsenosides Rb₁, Rb₂, Rc, Re, Rf, and Rg₁ decreased during the fermentation with S. cerevisiae, but Rd and Rg₃ increased by day 12. Ginsenosides Rb₁, Rb₂, Rc, Re, and Rg₁ decreased gradually in the extract with S. carlsbergensis, but Rd and Rg₃ were increased at day 6 and 9, respectively.     

Intraspecific Diversity of Yeast Associated to Vitis vinifera Albariño Must from Different Vineyard Ecosystems

The genetic diversity of Saccharomyces cerevisiae associated to the must (Vitis vinifera Albariño) from different geographic areas and spontaneous fermentation was studied by analysis of mitochondrial DNA restriction fragment length polymorphism (mtDNA RFLP). Colonies were isolated in musts at different stages of fermentation from three different geographic areas, O Val do Salnés, O Rosal and O Condado do Tea from Galicia (NW Spain). A total of 17 different patterns out of 273 S. cerevisiae colonies were identified using mitochondrial DNA restriction analysis. Pattern I from O Rosal, VI from O Val do Salnés and XII from O Condado do Tea were the predominant ones. Only pattern II was common to all fermentations. This molecular study provides useful information about distribution and genetic relationships among several S. cerevisiae strains associated to Vitis vinifera Albariño from different vineyard ecosystems and their influence on certain oenological properties.     

Determinants of flocculence of brewer's yeast during fermentation in wort

Ability of Saccharomyces cerevisiae MPY3 cells to flocculate during fermentation in wort was found to be triggered after growth limitation by oxygen shortage and to coincide with a sharp increase in cell surface hydrophobicity of the cells. Presence of oxygen in the pitching wort influenced final cell number, flocculence of the cells and cell surface hydrophobicity. Flocculation ability of cells grown in air-depleted pitching wort was hampered, concomitant with a decrease in final cell number and in final cell surface hydrophobicity. Addition of ergosterol and Tween 80 to air-depleted wort restored normal growth of the cells as well as flocculation ability and the increase in cell surface hydrophobicity. The same parameters increased in value after addition of ergosterol and Tween 80 to a fermentation with air-saturated pitching wort. Hydrophobicity of a non-flocculent mutant of S. cerevisiae strain MPY3, fermenting in air-saturated pitching wort, did not increase at cell division arrest. These results support the hypothesis that cell surface hydrophobicity is a major determinant for yeast cells to become flocculent, and suggest that shortage of sterols and unsaturated fatty acids precedes flocculence under brewing conditions.