Developing a novel selection method for alcoholic fermentation starters by exploring wine yeast microbiota from Greece

The selection of native yeast for alcoholic fermentation in wine focuses on ensuring the success of the process and promoting the quality of the final product. The purpose of this study was firstly to create a large collection of new yeast isolates and categorize them based on their oenological potential. Additionally, the geographical distribution of the most dominant species, Saccharomyces cerevisiae, was further explored. Towards this direction, fourteen spontaneously fermented wines from different regions of Greece were collected for yeast typing. The yeast isolates were subjected in molecular analyses and identification at species level. RAPD (Random Amplified Polymorphic DNA) genomic fingerprinting with the oligo-nucleotide primer M13 was used, combined with Matrix Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry (MALDI-TOF MS) technique. All yeast isolates were scrutinized for their sensitivity to killer toxin, production of non-desirable metabolites such as acetic acid and H2S, β-glucosidase production and resistance to the antimicrobial agent; SO2. In parallel, S. cerevisiae isolates were typed at strain level by interdelta – PCR genomic fingerprinting. S. cerevisiae strains were examined for their fermentative capacity in laboratory scale fermentation on pasteurized grape must. Glucose and fructose consumption was monitored daily and at the final point a free sorting task was conducted to categorize the samples according to their organoleptic profile. According to our results, among the 190 isolates, S. cerevisiae was the most dominant species while some less common non-Saccharomyces species such as Trigonopsis californica, Priceomyces carsonii, Zygosaccharomyces bailii, Brettanomyces bruxellensis and Pichia manshurica were identified in minor abundancies. According to phenotypic typing, most isolates were neutral to killer toxin test and exhibited low acetic acid production. Hierarchical Cluster Analysis revealed the presence of four yeast groups based on phenotypic fingerprinting. Strain level typing reported 20 different S. cerevisiae strains from which 65% indicated fermentative capacity and led to dry wines. Sensory evaluation results clearly discriminated the produced wines and consequently, the proposed yeast categorization was confirmed. A novel approach that employs biostatistical tools for a rapid screening and classification of indigenous wine yeasts with oenological potential, allowing a more efficient preliminary selection or rejection of isolates is proposed

TREM-1 expression on neutrophils and monocytes of septic patients: relation to the underlying infection and the implicated pathogen

<p>Abstract</p> <p>Background</p> <p>Current knowledge on the exact ligand causing expression of TREM-1 on neutrophils and monocytes is limited. The present study aimed at the role of underlying infection and of the causative pathogen in the expression of TREM-1 in sepsis.</p> <p>Methods</p> <p>Peripheral venous blood was sampled from 125 patients with sepsis and 88 with severe sepsis/septic shock. The causative pathogen was isolated in 91 patients. Patients were suffering from acute pyelonephritis, community-acquired pneumonia (CAP), intra-abdominal infections (IAIs), primary bacteremia and ventilator-associated pneumonia or hospital-acquired pneumonia (VAP/HAP). Blood monocytes and neutrophils were isolated. Flow cytometry was used to estimate the TREM-1 expression from septic patients.</p> <p>Results</p> <p>Within patients bearing intrabdominal infections, expression of TREM-1 was significantly lower on neutrophils and on monocytes at severe sepsis/shock than at sepsis. That was also the case for severe sepsis/shock developed in the field of VAP/HAP. Among patients who suffered infections by Gram-negative community-acquired pathogens or among patients who suffered polymicrobial infections, expression of TREM-1 on monocytes was significantly lower at the stage of severe sepsis/shock than at the stage of sepsis.</p> <p>Conclusions</p> <p>Decrease of the expression of TREM-1 on the membrane of monocytes and neutrophils upon transition from sepsis to severe sepsis/septic shock depends on the underlying type of infection and the causative pathogen.</p

Aufklärung der genetischen Grundlagen der autosomal rezessiven Cutis laxa

Autosomal recessive cutis laxa (ARCL) is a highly heterogeneous group of disorders, which is mainly associated with a progeroid appearance, lax and wrinkled skin, mental retardation and osteopenia and whose pathogenesis is only partially unravelled. After excluding mutations in the human ARCL genes, GORAB and ATP6V0A2, in the patients of our ARCL cohort without lung involvement, we conducted further genetic analysis in consanguineous families. These families were formerly diagnosed as GO, ARCLII-WSS and de Barsy syndrome. Homozygocity mapping in ten kindreds revealed a candidate region on chromosome 17q25. Sequencing of several genes in the candidate region resulted in the identification of disease-causing mutations in the gene PYCR1, encoding for Δ1-pyrroline-5-carboxylate reductase, an enzyme which catalyzes the final step of the de novo synthesis of proline. In total, 22 families of our ARCL cohort were found to bear missense, frameshift and splice site but no nonsense mutations. Apart from the common features of all ARCL forms, intrauterine growth retardation, mental retardation and a typical facies were the constant findings in these patients. Brain and eye abnormalities such as agenesis of corpus callosum and cataract were also evident. Proline levels, measured in serum and cell lysates from patients with mutations in PYCR1 did not reveal significant changes when compared with control samples. Affected individuals showed a pronounced progeroid appearance, which was not progressive. The detection of mutations in patients diagnosed as de Barsy syndrome implies that PYCR1 is one of the genes responsible for this genetically heterogeneous phenotype. All mutations found were associated with a decrease of protein expression in patient fibroblasts. Furthermore, immunofluorescence studies revealed a mitochondrial residence of the protein, as PYCR1 was found to co- localize with specific mitochondrial marker proteins. Immunofluorescence analysis provided evidence for an altered mitochondrial morphology upon loss of PYCR1, which was further confirmed by electron microscopy. The alterations were found to become more severe after transient treatment with H2O2 resulting in a five-fold increase in the number of fragmented mitochondria in patient cell lines. Upon oxidative stress the fibroblasts with mutations in PYCR1 showed higher apoptosis rates compared to control cells. Our findings suggest that PYCR1 contributes to the maintenance of the normal mitochondrial morphology and protects against oxidative stress, most probably by inhibiting apoptosis. This study postulates a new association between mutations in PYCR1, altered mitochondrial function and progeroid changes in connective tissues.Die autosomal rezessive Cutis laxa (ARCL) ist eine sehr heterogene Gruppe von Erkrankungen, die hauptsächlich durch einen progeroiden Aspekt, laxe und faltige Haut, mentale Retardierung sowie Osteopenie gekennzeichnet sind und deren Pathogenese bisher erst partiell aufgeklärt werden konnte. In zehn konsanguinen Familien, bei denen im Vorfeld Gerodermia Osteodysplastika (GO), Wrinkly Skin Syndrome oder de Barsy Syndrom diagnostiziert, jedoch keine Mutationen nachgewiesen worden waren, führten wir eine Homozygotie-Kartierung durch. In der resultierenden Kandidatenregion auf Chromosom 17q25 wurden nach Sequenzierung mehrerer Gene pathogene Mutationen im PYCR1-Gen entdeckt. PYCR1 kodiert die Δ1-Pyrollin-5-Karboxylat Reduktase, ein Enzym welches für den letzten Schritt der de novo Synthese von Prolin verantwortlich ist. In insgesamt 22 Familien aus unserer Kohorte konnten wir Missense-, Frameshift- und Splicesite-Mutationen, allerdings keine Nonsense-Mutationen in PYCR1 identifizieren. Zusätzlich zu den typischen Merkmalen aller CL Subtypen, fanden sich bei diesen Patienten eine intrauterine Wachstumsverzögerung, eine schwerere mentale Retardierung und eine typische Facies, sowie eine Agenesie oder Hypoplasie des Corpus callosum und eine Katarakt. Die betroffenen Individuen wiesen einen deutlich progeroiden, aber nicht progressiven Phänotyp auf. Eine Hypoprolinämie konnte weder im Serum von Patienten mit PYCR1 Mutationen, noch in Lysaten von betroffenen Hautfibroblasten festgestellt werden. Auch ursprünglich als de Barsy Syndrom diagnostizierte Patienten wiesen Mutationen im PYCR1-Gen auf, was für einen hohe genetische Heterogenität dieses Syndroms spricht. In kultivierten Fibroblasten von Patienten mit einer PYCR1 Mutation fand sich eine verminderte Expression des PYCR1 Proteins. Immunofluoreszenz-Analysen zeigten, dass das PYCR1 Protein mitochondrial lokalisiert ist, da PYCR1 mit spezifischen mitochondrialen Markerproteinen kolokalisiert war. Außerdem wiesen Zellen mit einer PYCR1 Mutation eine veränderte mitochondriale Morphologie auf, was mittels elektronmikroskopischer Analyse bestätigt wurde. Nach Behandlung mit H2O2 waren die beobachteten Veränderungen deutlich ausgeprägter und es zeigte sich eine fünffach erhöhte Anzahl fragmentierter Mitochondrien in Patienten- Fibroblasten. Nach oxidativem Stress lag in Fibroblasten mit PYCR1 Mutationen eine höhere Apoptoserate im Vergleich zu Kontrollzellen vor. Unsere Ergebnisse legen nahe, dass PYCR1 zur Aufrechterhaltung der normalen mitochondrialen Morphologie beiträgt und gleichzeitig eine protektive Funktion gegen oxitativen Stress ausübt, möglicherweise durch Inhibition von Apoptose. Unsere Studie demonstriert einen neuartigen Zusammenhang zwischen Mutationen in PYCR1, gestörter mitochondrialer Funktion und progeroiden Veränderungen von Bindegewebsstrukturen

Malolactic Fermentation&mdash;Theoretical Advances and Practical Considerations

There are two main fermentations associated with the vinification process. Alcoholic fermentation (AF), which is conducted by yeasts and malolactic fermentation (MLF), which takes place as a result of the metabolic activity of lactic acid bacteria (LAB) of the genera Oenococcus, former-Lactobacillus, Pediococcus and Leuconostoc. MLF is defined as the biotransformation of L-malic acid to L-lactic acid and carbon dioxide and in addition to deacidification, contributes significantly to microbial stability and often to the improvement of the sensory profile of wines. Therefore, the abiotic and biotic factors that affect MLF, along with its correlation with quality characteristics, has been in the epicenter of intensive research. In addition, practical issues that accompany MLF have also been considered and adequately assessed. The aim of the present review was to explore and critically discuss MLF from both theoretical and practical perspectives

Malolactic Fermentation—Theoretical Advances and Practical Considerations

There are two main fermentations associated with the vinification process. Alcoholic fermentation (AF), which is conducted by yeasts and malolactic fermentation (MLF), which takes place as a result of the metabolic activity of lactic acid bacteria (LAB) of the genera Oenococcus, former-Lactobacillus, Pediococcus and Leuconostoc. MLF is defined as the biotransformation of L-malic acid to L-lactic acid and carbon dioxide and in addition to deacidification, contributes significantly to microbial stability and often to the improvement of the sensory profile of wines. Therefore, the abiotic and biotic factors that affect MLF, along with its correlation with quality characteristics, has been in the epicenter of intensive research. In addition, practical issues that accompany MLF have also been considered and adequately assessed. The aim of the present review was to explore and critically discuss MLF from both theoretical and practical perspectives

Removal of rare earth elements from solutions using metakaolin alkali activated materials

Η παρούσα διπλωματική εργασία εκπονήθηκε στην Ερευνητική μονάδα «Τεχνολογίες Διαχείρισης Μεταλλευτικών & Μεταλλουργικών Αποβλήτων & Αποκατάστασης Εδαφών» της Σχολής Μηχανικών Ορυκτών Πόρων του Πολυτεχνείου Κρήτης.Περίληψη: Σκοπός αυτής της εργασίας ήταν η αξιολόγηση της ικανότητας γεωπολυμερών μετακαολίνη για την απομάκρυνση σπάνιων γαιών (REEs) από υδατικά διαλύματα και πιο συγκεκριμένα σκανδίου (Sc) και νεοδυμίου (Nd). Οι συγκεντρώσεις των σπανίων γαιών στα διαλύματα μετρήθηκαν με συσκευή ICP-MS. Τα γεωπολυμερή μετακαολίνη χαρακτηρίστηκαν με τη χρήση αναλυτικών τεχνικών όπως η τεχνική περίθλασης ακτίνων Χ (XRD), η φασματοφωτομετρία υπέρυθρου (FTIR), και η ηλεκτρονική μικροσκοπία σάρωσης (SEM/EDX). Πραγματοποιήθηκαν κινητικά πειράματα και πειράματα ισορροπίας και τα αποτελέσματα σχολιάστηκαν και αξιολογήθηκαν. Για τα κινητικά δεδομένα χρησιμοποιήθηκαν τα μοντέλα ψευδο-πρώτης και ψευδο-δεύτερης τάξης Lagergren και στην ισορροπία, μελετήθηκαν τα μοντέλα ισόθερμων Langmuir και Freundlich. Σε κάθε περίπτωση αξιολογήθηκε ο τύπος του μοντέλου που ταιριάζει καλύτερα στα πειραματικά αποτελέσματα.Summarization: The aim of this study was to evaluate the removal capacity of metakaolin based inorganic polymers (IPs) for the removal of Rare Earth Elements (REEs) from aqueous solutions, namely scandium (Sc) and neodymium (Nd). Analytical techniques involving XRD, FTIR and SEM/EDS were used for the characterization and morphology analysis of the produced metakaolin IPs. Kinetic and equilibrium experiments were performed and the results were evaluated. Kinetic models of pseudo-first and pseudo-second order Lagergren were applied, while at equilibrium the isotherm models of Langmuir and Freundlich were studied

Effect of Non-<i>Saccharomyces</i> Species Monocultures on Alcoholic Fermentation Behavior and Aromatic Profile of Assyrtiko Wine

Six wild-type non-Saccharomyces strains, belonging to the species Zygosaccharomyces bailii, Priceomyces carsonii, Trigonopsis californica, and Pichia manshurica, were evaluated for white wine production using Assyrtiko grapes from Santorini in Greece. Fermentation kinetics, in terms of glucose and fructose consumption and sensory analysis, was first employed to test the enological potential of the yeast strains. Based on their performance, two strains of Z. bailii (Zb-A19Y5 and Zb-K29Y2) and one strain of T. californica (Tc-A9Y1) selected for further analysis. The selected strains were tested in larger fermentation volumes for sugar consumption, while the produced wines were assessed for classical enological parameters, volatile compounds (GC/MS), and sensory analysis. Tc-A9Y1 could lead to dry wine (1 g/L of residual sugars) with 1.6 vol (12%) less alcohol than the other experimental wines. The wines that were fermented with the strain Zb-K29Y2 exhibited very high concentrations of isoamyl alcohol (604.33 ± 76.8 mg/L), but at the same time, they were characterized by low fruity notes. None of the produced wines presented any off-flavor aromas. Exploiting non-Saccharomyces strains with great fermentation capacity, which are able to produce high-quality wines and adapted to global warming conditions, is a new challenge for the wine industry

Effect of Myclobutanil Pesticide on the Physiological Behavior of Two Newly Isolated Saccharomyces cerevisiae Strains during Very-High-Gravity Alcoholic Fermentation

Yeasts are able to act as biosorbents, as their cell wall includes several components capable of binding organic xenobiotic compounds that can potentially be removed during various fermentation processes. In the present investigation, two novel Saccharomyces cerevisiae strains (LMBF-Y 16 and LMBF-Y-18), previously isolated from grapes, were studied regarding their physiological behavior (dry cell weight&mdash;DCW production, substrate uptake, and ethanol and glycerol biosynthesis) during fermentations of grape must, in some cases enriched with commercial glucose and fructose (initial total sugar concentration approximately 150 and 250 g/L, respectively). Myclobutanil (a chiral triazole fungicide broadly used as a protective agent of vine) was also added to the culture media at various concentrations in order to assess the ability of the yeasts to simultaneously perform alcoholic fermentations and detoxify the medium (i.e., to remove the fungicide). In the first set of experiments and for both tested strains, trials were carried out in either 250 mL or 2.0 L agitated shake flasks in either synthetic glucose-based experiments or grape musts. Since the results obtained in the trials where the cultures were placed in 2.0 L flasks with grape musts as substrates were superior in terms of both DCW and ethanol production, these experimental conditions were selected for the subsequent studies. Both strains showed high fermentative efficiency, producing high amounts of DCW (9.5&ndash;10.5 g/L) in parallel with high ethanol production, which in some cases achieved values very close to the maximum theoretical ethanol production yield (&asymp;0.49 g of ethanol per g of sugar). When using grape must with initial total sugars at approximately 250 g/L (very high gravity fermentation media, close to winemaking conditions), significantly high ethanol quantities (i.e., ranging between 105 and 123 g/L) were produced. Myclobutanil addition slightly negatively affected sugar conversion into ethanol; however, in all cases, ethanol production was very satisfactory. A non-negligible myclobutanil removal during fermentation, which ranged between 5%&ndash;27%, as a result of the adsorptive or degradative capacity of the yeast was also reported. The presence of myclobutanil had no effect on DCW production and resulted in no significant differences in the biosynthesis of glycerol. Therefore, these newly isolated yeast strains could be excellent candidates for simultaneous high ethanol production and parallel pesticide removal in a general biorefinery concept demonstrating many environmental benefits