It is estimated that more than 20% of processed apples and grapes are discarded as waste, which is dominated by pomace rich in malic acid that could be converted to high-value organic acids or other chemicals. A total of 98 yeast strains isolated from apple, grape, and plum wastes were evaluated for their ability to degrade malic acid relative to known yeast strains. Most (94%) of the new isolates degraded malic acid efficiently (> 50%) in the presence and absence of exogenous glucose, whereas only 14% of the known strains could do so, thus confirming the value of exploring (and exploiting) natural biodiversity. The best candidates were evaluated in synthetic media for their ability to convert malic acid to other valuable products under aerobic and oxygen-limited conditions, with two strains that produced ethanol and acetic acid as potential biorefinery products during aerobic cultivations and oxygen-limited fermentations on sterilized apple and grape pomace. Noteworthy was the identification of a Saccharomyces cerevisiae strain that is more efficient in degrading malic acid than other members of the species. This natural strain could be of value in the wine-making industry that often requires pH corrections due to excess malic acid.
Kpkt is a yeast killer toxin, naturally produced by Tetrapisispora phaffii, with possible applications in winemaking due to its antimicrobial activity on wine-related yeasts including Kloeckera/Hanseniaspora, Saccharomycodes and Zygosaccharomyces. Here, Kpkt coding gene was expressed in Komagataella phaffii (formerly Pichia pastoris) and the bioreactor production of the recombinant toxin (rKpkt) was obtained. Moreover, to produce a ready-to-use preparation of rKpkt, the cell-free supernatant of the K. phaffii recombinant killer clone was 80-fold concentrated and lyophilized. The resulting preparation could be easily solubilized in sterile distilled water and maintained its killer activity for up to six months at 4 °C. When applied to grape must, it exerted an extensive killer activity on wild wine-related yeasts while proving compatible with the fermentative activity of actively growing Saccharomyces cerevisiae starter strains. Moreover, it displayed a strong microbicidal effect on a variety of bacterial species including lactic acid bacteria and food-borne pathogens. On the contrary it showed no lethal effect on filamentous fungi and on Ceratitis capitata and Musca domestica, two insect species that may serve as non-mammalian model for biomedical research. Based on these results, bioreactor production and lyophilization represent an interesting option for the exploitation of this killer toxin that, due to its spectrum of action, may find application in the control of microbial contaminations in the wine and food industries.
- MeSH
- antiinfekční látky metabolismus farmakologie MeSH
- bioreaktory mikrobiologie MeSH
- fermentace MeSH
- killer faktory kvasinek biosyntéza farmakologie MeSH
- kvasinky účinky léků MeSH
- lyofilizace MeSH
- mikrobiální viabilita MeSH
- potravinářská mikrobiologie MeSH
- potravinářský průmysl MeSH
- rekombinantní proteiny biosyntéza farmakologie MeSH
- víno mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
The genus Candida is well known for its significant pathogenesis with which it radically affects the medical practice nowadays. C. albicans, above all, belongs to severe pathogens and it developed high resistance to a variety of antimicrobials. Undesirable effects of this species cast a shadow upon the entire group of these microorganisms, favourable metabolic qualities of which are thus often omitted. These properties find their usage in various fields of biotechnology. The present contribution is specifically focused on the application of genus Candida in traditional or recombinant biotechnology. Its pathogenesis is presented in contrast to the potential use of these yeasts in the industry.
Undesirable effects of the pathogen Botrytis cinerea include reduced quality and quantity of wine grapes. Winemaking is also complicated by the formation of a protein haze in white wines and oxidative browning of red wines. We analyzed proteins in experimental Moravian white wines characterized by their instability and haze formation in bottles during storage despite prior bentonite treatment. To study the relationship of wine proteins and haze, we carried out proteomics on hazy and clear white wines produced with partly or largely botrytized grapes and standard reference wines. Wine proteins were identified after their extraction, electrophoresis, and tryptic digestion by reversed-phase liquid chromatography of peptides, coupled with tandem mass spectrometry. Plant defense proteins, yeast glycoproteins, and various enzymes from Botrytis, particularly hydrolases, were found. As the content of the known haze-active thaumatin-like proteins and chitinases was visually low on stained gels (missing bands) compared to previous studies with unfined wines, other proteins are discussed in terms of the haze formation. As the main novelty, this work reveals the role of high proline-containing proteins in the propensity of white wines to turbidity following prior Botrytis damage of grapes.
Dekkera bruxellensis is important for lambic beer fermentation but is considered a spoilage yeast in wine fermentation. We compared two D. bruxellensis strains isolated from wine and found that they differ in some basic properties, including osmotolerance. The genomes of both strains contain two highly similar copies of genes encoding putative glycerol-proton symporters from the STL family that are important for yeast osmotolerance. Cloning of the two DbSTL genes and their expression in suitable osmosensitive Saccharomyces cerevisiae mutants revealed that both identified genes encode functional glycerol uptake systems, but only DbStl2 has the capacity to improve the osmotolerance of S. cerevisiae cells.
- MeSH
- Dekkera genetika izolace a purifikace metabolismus fyziologie MeSH
- druhová specificita MeSH
- fungální proteiny genetika metabolismus MeSH
- genom bakteriální genetika MeSH
- glycerol metabolismus MeSH
- osmoregulace genetika MeSH
- protony MeSH
- rekombinantní proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae genetika MeSH
- symportéry genetika metabolismus MeSH
- testy genetické komplementace MeSH
- víno mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Mid-exponential cultures of two traditional biotechnological yeast species, winery Saccharomyces cerevisiae and the less ethanol tolerant bottom-fermenting brewery Saccharomyces pastorianus, were exposed to different concentrations of added ethanol (3, 5 and 8%) The degree of ethanol-induced cell stress was assessed by measuring the cellular activity of superoxide dismutase (SOD), level of lipid peroxidation products, changes in cell lipid content and fatty acid profile. The resveratrol as an antioxidant was found to decrease the ethanol-induced rise of SOD activity and suppress the ethanol-induced decrease in cell lipids. A lower resveratrol concentration (0.5 mg/l) even reduced the extent of lipid peroxidation in cells. Resveratrol also alleviated ethanol-induced changes in cell lipid composition in both species by strongly enhancing the proportion of saturated fatty acids and contributing thereby to membrane stabilization. Lower resveratrol concentrations could thus diminish the negative effects of ethanol stress on yeast cells and improve their physiological state. These effects may be utilized to enhance yeast vitality in high-ethanol-producing fermentations or to increase the number of yeast generations in brewery.
- MeSH
- antioxidancia metabolismus MeSH
- ethanol farmakologie MeSH
- fyziologický stres účinky léků MeSH
- lipidy fyziologie MeSH
- mastné kyseliny metabolismus MeSH
- metabolismus lipidů účinky léků MeSH
- peroxidace lipidů účinky léků MeSH
- Saccharomyces cerevisiae účinky léků MeSH
- stilbeny farmakologie MeSH
- superoxiddismutasa metabolismus MeSH
- víno mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
Trace amounts of the carcinogenic ethyl carbamate can appear in wine as a result of a reaction between ethanol and citrulline, which is produced from arginine degradation by some bacteria used in winemaking. In this study, arginine deiminase (ADI) pathway genes were evaluated in 44 Oenococcus oeni strains from wines originating from several locations in order to establish the relationship between the ability of a strain to degrade arginine and the presence of related genes. To detect the presence of arc genes of the ADI pathway in O. oeni, pairs of primers were designed to amplify arcA, arcB, arcC and arcD1 sequences. All strains contained these four genes. The same primers were used to confirm the organization of these genes in an arcABCD1 operon. Nevertheless, considerable variability in the ability to degrade arginine among these O. oeni strains was observed. Therefore, despite the presence of the arc genes in all strains, the expression patterns of individual genes must be strain dependent and influenced by the different wine conditions. Additionally, the presence of arc genes was also determined in the 57 sequenced strains of O. oeni available in GenBank, and the complete operon was found in 83% of strains derived from wine. The other strains were found to lack the arcB, arcC and arcD genes, but all contained sequences homologous to arcA, and some of them had also ADI activity.
The aim of this study was a reliable intra-species discrimination and strain biodiversity in Oenococcus oeni populations of two different Aglianico wineries by molecular, biochemical, and physiological characterization. Pulsed field gel electrophoresis (PFGE) analysis revealed a high polymorphism related to the origin (winery) of strains, while differential display PCR (DD-PCR) allowed a further discrimination of strains from the same winery. Moreover, the heterogeneity of these natural populations was investigated by capillary electrophoresis and enzymatic assays. A variability related to a different surface charge distribution was observed among strains, linked to their origin. Malolactic activity study evidenced strain-specific differences in malic acid degradation, and then, only the presence of L(-)-malic acid in the medium induced the mle gene. This study provided evidences on the importance of intra-species biodiversity of malolactic bacterial populations in wine ecosystems, as each wine possess peculiar winemaking conditions and physical-chemical properties which make specific the bacterial survival and growth. This study highlighted a great biodiversity among O. oeni strains that can be also winery specific. Such biodiversity within a certain winery and winemaking area is important for selecting malolactic starters, and strain-specific trait identification is especially important to match individual strains to specific industrial process.
Five Oenococcus oeni strains, selected from spontaneous malolactic fermentation (MLF) of Patagonic Pinot noir wine, were assessed for their use as MLF starter cultures. After the individual evaluation of tolerance to some stress conditions, usually found in wine (pH, ethanol, SO2, and lysozyme), the behavior of the strains was analyzed in MLO broth with 14 % ethanol and pH 3.5 in order to test for the synergistic effect of high ethanol level and low pH and, finally, in a wine-like medium. Although the five strains were able to grow in MLO broth under low pH and/or high ethanol, they must be acclimated to grow in a wine-like medium. Additionally, glycosidase and tannase activities were evaluated, showing differences among the strains. The potential of the strains to ferment citrate was tested and two of the five strains showed the ability to metabolize this substrate. We did not detect the presence of genes encoding histidine, tyrosine descarboxylase, and putrescine carbamoyltransferase. All the strains tested exhibited good growth capacity and ability to consume L-malic acid in a wine-like medium after cell acclimation, and each of them showed a particular enzyme profile, which might confer different organoleptic properties to the wine.
Many different yeast species have been isolated from grapes and musts worldwide. The diversity and frequency of yeasts depend on a number of factors such as the grape variety, the physical damage of the grapes, the weather conditions and the chemical composition of must. A total of 366 isolates were associated with the three grape cultivars: Blue Frankish, Green Veltliner and Sauvignon blanc over four consecutive years. Yeast cultures were isolated from the grapes and from the fermenting musts after the first and seventh days. The ascomycetous yeasts of the genera Aureobasidium, Candida, Hanseniaspora, Metschnikowia, Pichia, Saccharomyces and Saccharomycopsis together with basidiomycetous yeasts of the genera Cryptococcus, Dioszegia, Filobasidium, Rhodotorula and Sporidiobolus were associated with the three grape varieties. Hanseniaspora uvarum, Metschnikowia pulcherrima, Pichia kluyveri, Pichia kudriavzevii and Sporidiobolus pararoseus were found on the berries in significant amounts. P. kluyveri and P. kudriavzevii were more associated with the damaged grapes, whereas Sp. pararoseus with intact ones. H. uvarum and M. pulcherrima were present on both types of grapes almost equally. The yeast composition and quantitative representation of yeast species varied over the grape varieties and the years examined. Although the basidiomycetous species formed a significant proportion of the yeast population in some individual grape variety/year combinations, the ascomycetous species were dominant.
- MeSH
- biodiverzita * MeSH
- DNA fungální chemie genetika MeSH
- fungální RNA genetika MeSH
- fylogeneze MeSH
- geny rRNA MeSH
- houby klasifikace genetika izolace a purifikace MeSH
- ribozomální DNA chemie genetika MeSH
- RNA ribozomální genetika MeSH
- sekvenční analýza DNA MeSH
- víno mikrobiologie MeSH
- Vitis mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Slovenská republika MeSH
