The sensitivity of yeasts and yeasts-like fungi to copper and sulfur could explain lower yeast biodiversity in organic vineyards.

Although differences in yeast biodiversity have often been found between vineyards subjected to organic protection or conventional protection, little is known about the effect of copper and sulfur fungicides (the only fungicides allowed in organic farming) on yeast populations. The sensitivity to copper and sulfur of 158 yeast isolates of seven different species (Aureobasidium pullulans, Hanseniaspora guilliermondii, H. uvarum, Metschnikowia sp., Pichia membranifaciens, Saccharomyces cerevisiae and Starmerella bacillaris) was evaluated. The species A. pullulans and St. bacillaris appeared to be more resistant to copper than the other species tested. The species A. pullulans, H. guilliermondii and Metschnikowia sp. had the highest sulfur resistance. Thus, only isolates of the species A. pullulans exhibited high resistance to both antifungal agents. These results may explain the lower diversity of yeasts present on berries and the strong dominance of the species A. pullulans for vineyards protected by copper or sulfur-based fungicides compared to other vineyards reported by several studies.

Adaptation of the wine bacterium Oenococcus oeni to ethanol stress: role of the small heat shock protein Lo18 in membrane integrity.

Malolactic fermentation in wine is often carried out by Oenococcus oeni. Wine is a stressful environment for bacteria because ethanol is a toxic compound that impairs the integrity of bacterial membranes. The small heat shock protein (sHsp) Lo18 is an essential actor of the stress response in O. oeni. Lo18 prevents the thermal aggregation of proteins and plays a crucial role in membrane quality control. Here, we investigated the interaction between Lo18 and four types of liposomes: one was prepared from O. oeni grown under optimal growth conditions (here, control liposomes), one was prepared from O. oeni grown in the presence of 8% ethanol (here, ethanol liposomes), one was prepared from synthetic phospholipids, and one was prepared from phospholipids from Bacillus subtilis or Lactococcus lactis. We observed the strongest interaction between Lo18 and control liposomes. The lipid binding activity of Lo18 required the dissociation of oligomeric structures into dimers. Protein protection experiments carried out in the presence of the liposomes from O. oeni suggested that Lo18 had a higher affinity for control liposomes than for a model protein. In anisotropy experiments, we mimicked ethanol action by temperature-dependent fluidization of the liposomes. Results suggest that the principal determinant of Lo18-membrane interaction is lipid bilayer phase behavior rather than phospholipid composition. We suggest a model to describe the ethanol adaptation of O. oeni. This model highlights the dual role of Lo18 in the protection of proteins from aggregation and membrane stabilization and suggests how modifications of phospholipid content may be a key factor determining the balance between these two functions.

High­throughput sequencing of amplicons for monitoring yeast biodiversity in must and during alcoholic fermentation.

We compared pyrosequencing technology with the PCR-ITS-RFLP analysis of yeast isolates and denaturing gradient gel electrophoresis (DGGE). These methods gave divergent findings for the yeast population. DGGE was unsuitable for the quantification of biodiversity and its use for species detection was limited by the initial abundance of each species. The isolates identified by PCR-ITSRFLP were not fully representative of the true population. For population dynamics, high-throughput sequencing technology yielded results differing in some respects from those obtained with other approaches. This study demonstrates that 454 pyrosequencing of amplicons is more relevant than other methods for studying the yeast community on grapes and during alcoholic fermentation. Indeed, this high-throughput sequencing method detected larger numbers of species on grapes and identified species present during alcoholic fermentation that were undetectable with the other techniques.

Influence of spontaneous, "pied de cuve" and commercial dry yeast fermentation strategies on wine molecular composition and sensory properties.

While most producers in recent decades have relied on commercial yeasts (ADY) as their primary choice given their reliability and reproducibility, the fear of standardising the taste and properties of wine has led to the employment of alternative strategies that involve autochthonous yeasts such as pied de cuve (PdC) and spontaneous fermentation (SF). However, the impact of different fermentation strategies on wine has been a subject of debate and speculation. Consequently, this study describes, for the first time, the differences between the three kinds of fermentation at the metabolomic, chemical, and sensory levels in two wines: Chardonnay and Pinot Noir. The results showed how the yeast chosen significantly impacted the molecular composition of the wines, as revealed by metabolomic analysis that identified biomarkers with varying chemical compositions according to the fermentation modality. Notably, higher numbers of lipid markers were found for SF and PdC than ADY, which contained more peptides. Key molecules from the metabolic amino acid pathway, which are addressed in this article, showed evidence of such variations. In addition, the analysis of volatile aromatic compounds revealed an increase in groups of compounds specific to each fermentation. The sensorial analysis of Chardonnay wine showed a more qualitative sensory outcome (Higher fruit intensity) for ADY and SF compared to PdC. Our finding challenges the common speculation among wine producers that autochthonous yeast fermentations may offer greater complexity and uniqueness in comparison to commercial yeast fermentations.

Respective Impact of Day and Night Bed Baths on Critical Care Patients.

BACKGROUND: Despite data supporting the reduction of interactions with patients during nighttime, bed bathing is sometimes performed within this period in sedated patients unable to manage their own hygiene care. OBJECTIVES: To compare patient physiologic variables and adverse effect incidence between night and day bed baths. METHODS: This was a single-center prospective observational study in a 12-bed intensive care unit during 2 months. Night period was defined to run from 10 pm to 6 am. Night bed baths were provided to sedated ventilated patients whatever their sedation, if their Richmond Agitation Sedation Scale score was -2 or deeper. Bed bath-induced changes in physiological variables, treatments, and related unscheduled events were registered during both night and day bed baths. RESULTS: Twenty-one patients (aged 62.9 [52.5-73.2] years, 14 male patients) were included. We registered 97 night bed baths and 95 day bed baths. Heart rate increased only after day bed baths (85 beats/min [bpm] [69-97 bpm] vs 88 bpm [73-98 bpm], P = .02). Increase in Richmond Agitation Sedation Scale score occurred, respectively, during 13 (13.4%) and 8 (8.4%) night and day bed baths, without significant differences. Body temperature significantly decreased during both night and day bed baths (respectively, 37°C [36.6°C-37.4°C] vs 36.6°C [36.2°C-37.2°C], P < .0001; and 36.9°C [36.5°C-37.2°C] vs 36.7°C [36.2°C-37.2°C], P = .0006). Overall, unscheduled events, whether physiologic changes, pain, or calling a physician in rescue occurred in 97 procedures (50.5%), irrespective of their timing (night vs day, respectively 53% [54.6%] vs 44% [46.3%], P = .31). DISCUSSION: Although unscheduled events occurred in half of bed baths, differences evidenced between nighttime and daytime bed baths were scarce. The appropriateness of nighttime bed bathing remains questionable.

Metschnikowia pulcherrima Influences the Expression of Genes Involved in PDH Bypass and Glyceropyruvic Fermentation in Saccharomyces cerevisiae.

Previous studies reported that the use of Metschnikowia pulcherrima in sequential culture fermentation with Saccharomyces cerevisiae mainly induced a reduction of volatile acidity in wine. The impact of the presence of this yeast on the metabolic pathway involved in pyruvate dehydrogenase (PDH) bypass and glycerol production in S. cerevisiae has never been investigated. In this work, we compared acetic acid and glycerol production kinetics between pure S. cerevisiae culture and its sequential culture with M. pulcherrima during alcoholic fermentation. In parallel, the expression levels of the principal genes involved in PDH bypass and glyceropyruvic fermentation in S. cerevisiae were investigated. A sequential culture of M. pulcherrima/S. cerevisiae at an inoculation ratio of 10:1 produced 40% less acetic acid than pure S. cerevisiae culture and led to the enhancement of glycerol content (12% higher). High expression levels of pyruvate decarboxylase PDC1 and PDC5, acetaldehyde dehydrogenase ALD6, alcohol dehydrogenase ADH1 and glycerol-3-phosphate dehydrogenase PDC1 genes during the first 3 days of fermentation in sequential culture conditions are highlighted. Despite the complexity of correlating gene expression levels to acetic acid formation kinetics, we demonstrate that the acetic acid production pathway is altered by sequential culture conditions. Moreover, we show for the first time that the entire acetic acid and glycerol metabolic pathway can be modulated in S. cerevisiae by the presence of M. pulcherrima at the beginning of fermentation.

Molecular cloning, heterologous expression, and characterization of Ornithine decarboxylase from Oenococcus oeni.

Ornithine decarboxylase (ODC) is responsible for the production of putrescine, the major biogenic amine found in wine. Oenococcus oeni is the most important lactic acid bacterium in the winemaking process and is involved in malolactic fermentation. We report here the characterization of ODC from an O. oeni strain isolated from wine. Screening of 263 strains isolated from wine and cider from all over the world revealed that the presence of the odc gene appears to be strain specific in O. oeni. After cloning, heterologous expression in Escherichia coli, and characterization, the enzyme was found to have a molecular mass of 85 kDa and a pI of 6.2 and revealed maximal activity at pH 5.5 and an optimum temperature of 35°C. Kinetic studies showed that O. oeni ODC is specific for L-ornithine with a K(m) value of 1 mM and a V(max) of 0.57 U·mg(-1). The hypothesis that cadaverine, which results from lysine decarboxylation, may be linked to putrescine production is not valid since O. oeni ODC cannot decarboxylate L-lysine. As no lysine decarboxylase was detected in any of the O. oeni genomes sequenced, cadaverine synthesis may result from another metabolic pathway. This work is the first characterization of an ODC from a lactic acid bacterium isolated from a fermented product.

Anti-ulcer activity of Synadenium grantii latex

Synadenium grantii Hook f., Euphorbiaceae, is popularly known as leitosinha or janaúba. The diluted latex (18 drops/L of water) is commonly used in the south of Brazil to treat gastric disturbances. This study evaluated phytochemical screening and toxicity using Artemia salina Leach of crude bark extract and also latex. The toxicity and the anti-ulcer activity of S. grantii latex were also tested in rats. Phytochemical results showed presence of tannins, terpenes, unsaponificable substances, coumarins and anthraquinones in the crude bark extract and terpenes in the latex. Gas chromatography-mass spectrometry (GC-MS) analysis demonstrated the presence of diterpene tigliane esters in the latex, identified as 12-deoxyphorbol-13-(2-metilpropionate) and phorbol 12,13,20-triacetate. The toxicity results using A. salina presented CL50 26.58μg/mL and CL50 778.66μg/mL, for the latex and the crude bark extract respectively. The toxicological hepatic parameters of the diluted latex were not different to the control group (p<0.05). The eosinophils cells showed an increase in both the diluted and pure latex groups. The pure latex showed gastric protection of 90% (p<0.05) and the diluted latex showed 6% compared to the negative control. Therefore, our data indicate that S. grantii latex, under research conditions presented gastric protection. Pure latex showed more toxicity than the diluted latex.