Correlation between Low Temperature Adaptation and Oxidative Stress in Saccharomyces cerevisiae

Many factors, such as must composition, juice clarification, fermentation temperature, or inoculated yeast strain, strongly affect the alcoholic fermentation and aromatic profile of wine. As fermentation temperature is effectively controlled by the wine industry, low-temperature fermentation (10–15°C) is becoming more prevalent in order to produce white and “rosé” wines with more pronounced aromatic profiles. Elucidating the response to cold in Saccharomyces cerevisiae is of paramount importance for the selection or genetic improvement of wine strains. Previous research has shown the strong implication of oxidative stress response in adaptation to low temperature during the fermentation process. Here we aimed first to quantify the correlation between recovery after shock with different oxidants and cold, and then to detect the key genes involved in cold adaptation that belong to sulfur assimilation, peroxiredoxins, glutathione-glutaredoxins, and thioredoxins pathways. To do so, we analyzed the growth of knockouts from the EUROSCARF collection S. cerevisiae BY4743 strain at low and optimal temperatures. The growth rate of these knockouts, compared with the control, enabled us to identify the genes involved, which were also deleted and validated as key genes in the background of two commercial wine strains with a divergent phenotype in their low-temperature growth. We identified three genes, AHP1, MUP1, and URM1, whose deletion strongly impaired low-temperature growth.This work has been financially supported from the Spanish Government through MINECO and FEDER funds (AGL2013-47300-C3-3-R and PCIN-2015-143 grants) and from Generalitat Valenciana through PROMETEOII/2014/042 grant, awarded to JG. This study has been carried out in the context of the European Project ERA-IB “YeastTempTation” EG also thanks the Spanish government for an FPI grant BES-2011-044498.Peer reviewe

Functional analysis of lipid metabolism genes in wine yeasts during alcoholic fermentation at low temperature

11 pages, 1 table, 6 figures.Wine produced by low-temperature fermentation is mostly considered to have improved sensory qualities. However few commercial wine strains available on the market are well-adapted to ferment at low temperature (10 – 15°C). The lipid metabolism of Saccharomyces cerevisiae plays a central role in low temperature adaptation. One strategy to modify lipid composition is to alter transcriptional activity by deleting or overexpressing the key genes of lipid metabolism. In a previous study, we identified the genes of the phospholipid, sterol and sphingolipid pathways, which impacted on growth capacity at low temperature. In the present study, we aimed to determine the influence of these genes on fermentation performance and growth during low-temperature wine fermentations. We analyzed the phenotype during fermentation at the low and optimal temperature of the lipid mutant and overexpressing strains in the background of a derivative commercial wine strain. The increase in the gene dosage of some of these lipid genes, e.g., PSD1, LCB3, DPL1 and OLE1, improved fermentation activity during low-temperature fermentations, thus confirming their positive role during wine yeast adaptation to cold. Genes whose overexpression improved fermentation activity at 12°C were overexpressed by chromosomal integration into commercial wine yeast QA23. Fermentations in synthetic and natural grape must were carried out by this new set of overexpressing strains. The strains overexpressing OLE1 and DPL1 were able to finish fermentation before commercial wine yeast QA23. Only the OLE1 gene overexpression produced a specific aroma profile in the wines produced with natural grape must.This work has been financially supported by grants AGL2010-22001-C02-01 and PROMETEOII/2014/042 from the Spanish government and the Generalitat Valenciana, respectively, awarded to JMG. MLM wishes to thank the Spanish government for her FPI grant.Peer reviewe

Improved antimicrobial activity of immobilised essential oil components against representative spoilage wine microorganisms

[EN] Wine, as a fermented drink, is considered a microbiologically safe beverage, but the growth of spoilage microorganisms can cause economic damage. As a new preservative process, the application of immobilised essential oil components (EOCs) is proposed in this study. EOCs were attached to the surface of three different commercial supports (silica particles, cellulose particles and cellulosic membrane) to avoid the disadvantages of using these compounds in their free form, such as volatility, low water solubility and intense aroma. The results showed that the treatment of spoilage microorganisms with antimicrobial particles (silica and cellulose) significantly reduced the viability and growth capacity of the target microorganisms. The covalent attachment of EOCs to particles led to a significant reduction in both the MIC values and viability compared with most free compounds. The enhanced antimicrobial activity of EOCs after their anchorage to a support was confirmed, resulting in MIC values of 10-90 fold lower than those of the free bioactive compounds. In addition, the filtration of microorganism suspensions through EOC-functionalised membranes showed remarkably antimicrobial activity.Authors gratefully acknowledge the financial support from the Ministerio de Economia y Competitividad and FEDER-EU (Projects AGL2015-70235-C2-1-R and AGL2016-77505-C3-1-R, granted to JMB and JMG, respectively). The authors also thank the Electron Microscopy Service at the UPV for support. Authors thank Antonio Ruiz for technical assistance.García-Ríos, E.; Ruiz Rico, M.; Guillamón Navarro, JM.; Pérez-Esteve, É.; Barat Baviera, JM. (2018). Improved antimicrobial activity of immobilised essential oil components against representative spoilage wine microorganisms. Food Control. 94:177-186. https://doi.org/10.1016/j.foodcont.2018.07.005S1771869

Generation of intra- and interspecific Saccharomyces hybrids with improved oenological and aromatic properties

Non-wine yeasts could enhance the aroma and organoleptic profile of wines. However, compared to wine strains, they have specific intolerances to winemaking conditions. To solve this problem, we generated intra- and interspecific hybrids using a non-GMO technique (rare-mating) in which non-wine strains of S. uvarum, S. kudriavzevii and S. cerevisiae species were crossed with a wine S. cerevisiae yeast. The hybrid that inherited the wine yeast mitochondrial showed better fermentation capacities, whereas hybrids carrying the non-wine strain mitotype reduced ethanol levels and increased glycerol, 2,3-butanediol and organic acid production. Moreover, all the hybrids produced several fruity and floral aromas compared to the wine yeast: β-phenylethyl acetate, isobutyl acetate, γ-octalactone, ethyl cinnamate in both varietal wines. Sc × Sk crosses produced three- to sixfold higher polyfunctional mercaptans, 4-mercapto-4-methylpentan-2-one (4MMP) and 3-mercaptohexanol (3MH). We proposed that the exceptional 3MH release observed in an S. cerevisiae × S. kudriavzevii hybrid was due to the cleavage of the non-volatile glutathione precursor (Glt-3MH) to detoxify the cell from the presence of methylglyoxal, a compound related to the high glycerol yield reached by this hybrid. In conclusion, hybrid generation allows us to obtain aromatically improved yeasts concerning their wine parent. In addition, they reduced ethanol and increased organic acids yields, which counteracts climate change effect on grapes.EEA MendozaFil: Perez, Maria Dolores. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Mendoza; Argentina.Fil: Perez, Maria Dolores. Consejo Superior de Investigación Científica (CSIC). Instituto de Agroquímica y Tecnología de los Alimentos (IATA). Departamento de Biotecnología de los Alimentos, Grupo de Biología de Sistemas en Levaduras de Interés Biotecnológico; EspañaFil: Denat, Marie. Universidad de Zaragoza. Department of Analytical Chemistry. Laboratory for Aroma Analysis and Enology. Instituto Agroalimentario de Aragón; EspañaFil: Pérez-Través, Laura. Consejo Superior de Investigación Científica (CSIC). Instituto de Agroquímica y Tecnología de los Alimentos (IATA). Departamento de Biotecnología de los Alimentos, Grupo de Biología de Sistemas en Levaduras de Interés Biotecnológico; EspañaFil: Heras, José María. Lallemand Bio S.L.; EspañaFil: Guillamón, José Manuel. Consejo Superior de Investigación Científica (CSIC). Instituto de Agroquímica y Tecnología de los Alimentos (IATA). Departamento de Biotecnología de los Alimentos, Grupo de Biología de Sistemas en Levaduras de Interés Biotecnológico; EspañaFil: Ferreira, Vicente. Universidad de Zaragoza. Department of Analytical Chemistry. Laboratory for Aroma Analysis and Enology. Instituto Agroalimentario de Aragón; EspañaFil: Querol, Amparo. Consejo Superior de Investigación Científica (CSIC). Instituto de Agroquímica y Tecnología de los Alimentos (IATA). Departamento de Biotecnología de los Alimentos, Grupo de Biología de Sistemas en Levaduras de Interés Biotecnológico; Españ

Acetic acid bacteria in oenology

Els bacteris de l'àcid acètic han estat sempre considerats perjudicials en els àmbits enològics, com a principal font de problemes en el vi (fonamentalment per ser una font d'acidesa volàtil). El desenvolupament de noves tècniques de biologia molecular ha permès que la taxonomia i el coneixement del metabolisme d'aquests bacteris que, per les seves exigències nutricionals, són molt difícils de cultivar evolucionin ràpidament. En l'àmbit taxonòmic, s'ha produït el canvi de dos gèneres i cinc espècies el 1984 a deu gèneres i més de quaranta espècies al moment actual. De totes maneres, les poderoses eines moleculars utilitzades en taxonomia no són apropiades per a l'ús rutinari en estudis ecològics, on s'ha d'analitzar un gran nombre de mostres. Per tant, s'han desenvolupat noves tècniques moleculars que han permès la millora del seu coneixement i control a Enologia. Així mateix, han permès un avenç considerable en la producció de vinagre, procés en el qual aquests bacteris són imprescindibles, com s'ha posat de manifest en el projecte europeu WINEGAR.Acetic acid bacteria have always been considered the bad microorganisms of oenology; responsible for wine spoiling (vinegary taint). The taxonomy and our knowledge of the metabolism of acetic acid bacteria are rapidly evolving, especially as new molecular biology techniques are applied to this fastidious group of microorganisms, which are still rather difficult to work with. The dramatic change that acetic acid bacteria taxonomy has undergone can be summarized by the transformation of 2 genera and 5 species in 1984 into 10 genera and over 40 species at present. The powerful molecular tools used in taxonomy are not appropriate for frequent use in identification and ecological studies; yet new molecular tools for routine analysis have also been developed. These provide new insights and means of controlling this microbial group. Furthermore, these advances have improved vinegar production; a process where the presence of acetic acid bacteria is essential. The WINEGAR European Project is evidence of these improvements in vinegar production

Elementos del conocimiento en la ingeniería del mantenimiento industrial. Final

[ES] Todas las opciones tácticas en la ingeniería del mantenimiento industrial requieren una gran especialización, una adecuada gestión de la información, experiencia y del conocimiento generado, que implica el grado de eficiencia del propio departamento de mantenimiento, y con ello, la funcionalidad de los equipos e instalaciones, que hacen aumentar la productividad o el servicio a prestar por la propia empresa. En este artículo se analizan los elementos de la gestión y transferencia del conocimiento que están implícitos en las opciones tácticas básicas en la ingeniería del mantenimiento industrial.Cárcel Carrasco, FJ.; Grau Carrión, J.; Pascual Guillamón, M. (2016). Elementos del conocimiento en la ingeniería del mantenimiento industrial. Final. Mantenimiento en Latinoamérica. 8(1):9-14. http://hdl.handle.net/10251/102626S9148

Andreev reflection under high magnetic fields in ferromagnet-superconductor nanocontacts

We study the magnetic-field dependence of the conductance in planar ferromagnet-superconductor nanocontacts created with focused-electron/ion-beam techniques. From the fits of the differential conductance curves in high magnetic fields, we obtain the magnetic field dependences of the superconducting gap and the broadening parameter. Orbital depairing is found to be linear with magnetic field. We evaluate the magnetic field dependence of the quasiparticle density of states, and we compare it with the value obtained by scanning tunneling spectroscopy experimentsThis work was supported by the Spanish Ministry of Science (through projects MAT2008-06567-C02, including FEDER funding) and the Arag´on Regional Government (project E26). S. Sangiao acknowledges financial support from Spanish ME

Genome-wide identification of genes involved in growth and fermentation activity at low temperature in Saccharomyces cerevisiae

Fermentation at low temperatures is one of the most popular current winemaking practices because of its reported positive impact on the aromatic profile of wines. However, low temperature is an additional hurdle to develop Saccharomyces cerevisiae wine yeasts, which are already stressed by high osmotic pressure, low pH and poor availability of nitrogen sources in grape must. Understanding the mechanisms of adaptation of S. cerevisiae to fermentation at low temperature would help to design strategies for process management, and to select and improve wine yeast strains specifically adapted to this winemaking practice. The problem has been addressed by several approaches in recent years, including transcriptomic and other high-throughput strategies. In this work we used a genome-wide screening of S. cerevisiae diploid mutant strain collections to identify genes that potentially contribute to adaptation to low temperature fermentation conditions. Candidate genes, impaired for growth at low temperatures (12 °C and 18 °C), but not at a permissive temperature (28 °C), were deleted in an industrial homozygous genetic background, wine yeast strain FX10, in both heterozygosis and homozygosis. Some candidate genes were required for growth at low temperatures only in the laboratory yeast genetic background, but not in FX10 (namely the genes involved in aromatic amino acid biosynthesis). Other genes related to ribosome biosynthesis (SNU66 and PAP2) were required for low-temperature fermentation of synthetic must (SM) in the industrial genetic background. This result coincides with our previous findings about translation efficiency with the fitness of different wine yeast strains at low temperature.Funding from the Spanish Government trough MINECO and FEDER funds: MINECO AGL2012-32064 and AGL2015-63629-R grants, INIA RM2012-00007-00-00 grant, MINECO RTC-2014-2186-2 and MINECO PCIN-2015-143 grants is acknowledged.Peer reviewe

Differential proteomic analysis by SWATH-MS unravels the most dominant mechanisms underlying yeast adaptation to non-optimal temperatures under anaerobic conditions

Elucidation of temperature tolerance mechanisms in yeast is essential for enhancing cellular robustness of strains, providing more economically and sustainable processes. We investigated the differential responses of three distinct Saccharomyces cerevisiae strains, an industrial wine strain, ADY5, a laboratory strain, CEN.PK113-7D and an industrial bioethanol strain, Ethanol Red, grown at sub- and supra-optimal temperatures under chemostat conditions. We employed anaerobic conditions, mimicking the industrial processes. The proteomic profile of these strains in all conditions was performed by sequential window acquisition of all theoretical spectra-mass spectrometry (SWATH-MS), allowing the quantification of 997 proteins, data available via ProteomeXchange (PXD016567). Our analysis demonstrated that temperature responses differ between the strains; however, we also found some common responsive proteins, revealing that the response to temperature involves general stress and specific mechanisms. Overall, sub-optimal temperature conditions involved a higher remodeling of the proteome. The proteomic data evidenced that the cold response involves strong repression of translation-related proteins as well as induction of amino acid metabolism, together with components related to protein folding and degradation while, the high temperature response mainly recruits amino acid metabolism. Our study provides a global and thorough insight into how growth temperature affects the yeast proteome, which can be a step forward in the comprehension and improvement of yeast thermotolerance.Financial support is acknowledged to Project ERA-IB “YeastTempTation” (ERA-IB-2-6/0001/2014), and FCT for the strategic funding of UIDB/04469/2020 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684), and BioTecNorte operation (NORTE-01-0145-FEDER-000004). Lallemand Ibéria, SA is acknowledged for the supply of yeast strains. Te proteomic analysis was carried out in the SCSIE University of Valencia Proteomics Unit, a member of the ISCIII ProteoRed Proteomics Platform. Authors thank to Luz Valero, researcher of the Proteomic unit, for her valuable support.info:eu-repo/semantics/publishedVersio