Yeasts

PCR assays have not reached the same level of acceptance for the detection of human fungal pathogens as for other microorganisms , mainly because the low number of microorganisms challenges the detection limits of PCR. Therefore, whereas meta-analyses focusing on clinical validation suggest interest in adding PCR results to the diagnostic workup for invasive fungal disease (IFD) along with clinical evaluation, CT scans, classical mycology and antigen detection, no consensual PCR method has emerged. Compared with the end-point format of the 1990s, real-time quantitative PCR is a major breakthrough. This format prevents contamination with previously amplified products, provides the yield of amplification, allows for developing consensus procedures and should therefore be the only format used. An internal control is now mandatory to avoid false-negative results. Primer design strongly impacts on the objectives: pan-fungal primers can provide false-positive results due to environmental fungal DNA contamination; conversely, species-specific primers miss infections caused by untargeted fungi. Unresolved issues include the best specimens to be used; serum is currently preferred to blood because of the ease of the DNA extraction step. Work is in progress to establish standards at least for Aspergillus PCR, and the implementation of quality controls should help centres to improve assays. Eventually, the classical analysis of biomarker performance does not consider the evolving risk factors and changing treatments during IFD, which can lead to variable conclusions. New statistical methods such as event history analysis should be considered.

The aim of this work was to determine the effect of yeast extract and of its vitamin contents on autotrophic and heterotrophic growth and metabolism of four acetogenic bacteria from the human colon. Yeast extract exerted a stimulatory effect on autotrophic growth of the colonic acetogens, but concentration of this compound above 1-2 g. L Ϫ1 in the medium did not enhance utilization of H 2 /CO 2 . Vitamins provided by yeast extract were shown to be essential cofactors of the reductive pathway of acetate synthesis except for one Clostridium strain. Yeast extract was also necessary to maintain heterotrophic growth and acetate synthesis from glucose in acetogenic species, except in the Streptococcus strain. In the absence of yeast extract, vitamins could efficiently restore glucose fermentation via acetate. The reductive and oxidative pathways of acetate synthesis might, therefore, depend on vitamin cofactors supplied by yeast extract in most of the human acetogenic bacteria. Non-vitaminic factors appeared also to be involved in the metabolism of some of these acetogenic species.

The goal of the two experiments was to determine if Yeast had the two life-like characteristics, Metabolism and comprised of Cells, to determine whether Yeast is alive. In the two experiments, whether yeast excretes waste, more specifically CO2, and whether yeast reproduces over a period of time, were tested. The results of the two tests performed were ambiguous. The data for some of the experiments was inconsistent, and needed more testing. Also, some of the experiments were potentially flawed, and need to be performed again with corrections in order to produce credible data. Therefore, the group can conclude nothing regarding the two hypotheses.

A new-type in situ probe has been developed to acquire dark field images of yeast in bioreactors. It has been derived from an in situ bright field microscope that is able to measure cell density in bioreactors during fermentation processes. The illumination part of the probe has been replaced with a dark field device, in which an aspheric condenser is used, so that high contrast dark field images can be obtained. The technique of second imaging is implemented to improve the sharpness of the images by means of a relay lens. This new in situ probe is expected to enable the evaluation of the cell viability without staining owing to modern image processing.

Propolis or bee-glue is a sticky substance collected in the hives by the honeybee (Apis mellifica L., Apidae) from various tree sources. In the northern hemisphere (Europe, North and South America and western Asia), the tree sources are: Populus spp., Betula spp., Ul-423 Acta Pharm. 55 (2005) 423-430

Invertase and urease are enzyme entities highly associated with the cells of the astaxanthin-producer yeast Xanthophyllomyces dendrorhous (Phaffia rhodozyma) during any stage of its cell growth cycle. In this study cellobiose was a more efficient carbon source ...

For new species in Lipomyces are described. In terms of nuclear genome comparison the genus Lipomyces comprises three species-clusters. A key to the nine species now assigned to the genus, is given. The delimitation of species by (i) rRNA nucleotide sequence analyses of the 18S and 25S subunits and (II) nDNA homology determinations, do not invariably lead to the recognition of congruent taxa. The family of the Lipomycetaceae, nevertheless provides an illuminative model for phylogenetic studies in terms of more appropriate ribosomal nucleotide sequence analyses of both its teleomorphic and anamorphic members.

Abbreviations: FUS/TLS: fusion involved in t(12:16) in malignant liposarcoma / translocation in liposarcoma; ; LC-MS/MS: liquid chromatography tandem mass spectrometry; OTUB1: otubain 1; OTUB2: otubain 2; Rack 1: receptor for activated kinase 1; Ub: ubiquitin Word counts: Total: 8121; Abstract: 191 1 Abstract

Beauveria bassiana colonizes insect hosts initially through a yeast phase, which is common in some artificial liquid cultures, but not reported on artificial solid media. We describe a yeast-like phase for B. bassiana isolate 447 (ATCC 20872) on MacConkey agar and its virulence toward Diatraea saccharalis and Tetranychus urticae. The yeast-like cells of B. bassiana developed by budding from germinating conidia after 24-h incubation. Cells were typically and fungal colonies were initially circular and mucoid, but later were covered with mycelia and conidia. Ability to produce yeast-like cells on MacConkey medium was relatively common among different B. bassiana isolates, but growth rate and timing of yeast-like cell production also varied. Metarhizium anisopliae and Paecilomyces spp. isolates did not grow as yeast-like cells on MacConkey medium. Yeast-like cells of B. bassiana 447 were more virulent against D. saccharalis than conidia when 107 cells/ml were used. At 108 cells/ml, the estimated mean survival time was 5.4 days for the yeast suspension and 7.7 days for the conidial suspension, perhaps due to faster germination. The LC50 was also lower for yeast than conidial suspensions. Yeast-like cells and conidia had similar virulence against T. urticae; the average mortalities with yeast-like cells and conidia were, respectively, 42.8 and 45.0%, with 107 cells/ml, and 77.8 and 74.4%, with 108 cells/ml. The estimated mean survival times were 3.6 and 3.9 for yeast and conidial suspensions, respectively. The bioassay results demonstrate the yeast-like structures produced on MacConkey agar are effective as inoculum for B. bassiana applications against arthropod pests, and possibly superior to conidia against some species. Obtaining well-defined yeast phase cultures of entomopathogenic hyphomycetes may be an important step in studies of the biology and nutrition, pathogenesis, and the genetic manipulation of these fungi.

Although most of what is known about the biology and function of arachidonic acid metabolites comes from the study of mammalian biology, these compounds can also be produced by lower eukaryotes, including yeasts and other fungi. It is also in this group of organisms that the least is known about the metabolic pathways leading to the production of these compounds as well as the functions of these compounds in the biology of fungi and yeasts. This review will deal with the discovery of oxylipins from polyunsaturated fatty acids, and more specifically the arachidonic acid derived eicosanoids, such as 3-hydroxy eicosatetraenoic acid, prostaglandin F 2α and prostaglandin E 2 , in yeasts starting in the early 1990s. This review will also focus on what is known about the metabolic pathways and/or proteins involved in the production of these compounds in pathogenic yeasts. The possible roles of these compounds in the biology, including the pathology, of these organisms will be discussed.

A total of 365 yeasts were isolated from the brines of soft, semihard and hard cheeses from different manufacturers. Identification was based on 131 characteristics, primarily employing a method with microtitration plates. Most brines exhibited a characteristic yeast flora. The predominant strains proved to be mainly Debaryomyces hansenii and Candida versatilis. In a few brines Trichosporon beigelii, C. rugosa, C. intermedia, Kluyveromyces marxianus, Saccharomyces sp. and C. tenuis/polymorpha were predominant. Also of importance were C. tropicalis, C. parapsilosis, C. zeylanoides, Issatchenkia orientalis and Geotrichum klebahnii. Not all strains could be clearly identified. Lists of characters are provided for subdividing D. hansenii and T. beigelii. The specificity of the yeast flora of brines is assumed to contribute to the sensory variety of cheeses.

A total of forty-five wild yeast strains were isolated from five traditional Greek wheat sourdoughs. Strains were identified using the classical identification technique along with the sodium dodecyl sulfatepolyacrylamide gel electrophoresis of whole cell proteins (SDS-PAGE), Fourier transform-infrared spectroscopy (FT-IR) and the randomly amplified polymorphic DNA-polymerase chain reaction analysis (RAPD-PCR), The latter methods confirmed the classical identification. According to the results obtained, fourteen strains were identified as Saccharomyces cerevisiae strains, twenty-five as Pichia membranaefaciens strains and six as Yarrowia lipolytica.

Saccharomyces cerevisiae is one of the most important model organisms and has been a valuable asset to human civilization. However, despite its extensive use in the last 9,000 y, the existence of a seasonal cycle outside human-made environments has not yet been described. We demonstrate the role of social wasps as vector and natural reservoir of S. cerevisiae during all seasons. We provide experimental evidence that queens of social wasps overwintering as adults (Vespa crabro and Polistes spp.) can harbor yeast cells from autumn to spring and transmit them to their progeny. This result is mirrored by field surveys of the genetic variability of natural strains of yeast. Microsatellites and sequences of a selected set of loci able to recapitulate the yeast strain's evolutionary history were used to compare 17 environmental wasp isolates with a collection of strains from grapes from the same region and more than 230 strains representing worldwide yeast variation. The wasp isolates fall into subclusters representing the overall ecological and industrial yeast diversity of their geographic origin. Our findings indicate that wasps are a key environmental niche for the evolution of natural S. cerevisiae populations, the dispersion of yeast cells in the environment, and the maintenance of their diversity. The close relatedness of several wasp isolates with grape and wine isolates reflects the crucial role of human activities on yeast population structure, through clonal expansion and selection of specific strains during the biotransformation of fermented foods, followed by dispersal mediated by insects and other animals. evolutionary biology | genomics

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a rapid, highly accurate, and cost-effective method for routine identification of a wide range of microorganisms. We carried out a side by side comparative evaluation of the performance of Bruker Biotyper versus VITEK MS for identification of a large and diverse collection of microorganisms. Most difficult and/or unusual microorganisms, as well as commonly encountered microorganisms were selected, including Gram-positive and negative bacteria, mycobacteria, actinomycetes, yeasts and filamentous fungi. Six hundred forty two strains representing 159 genera and 441 species from clinical specimens previously identified at the Laboratoire de santé publique du Québec (LSPQ) by reference methods were retrospectively chosen for the study. They included 254 Gram-positive bacteria, 167 Gram-negative bacteria, 109 mycobacteria and aerobic actinomycetes and 112 yeasts and moulds. MALDI-TOF MS analyses were performed on both systems according to the manufacturer's instructions. Of the 642 strains tested, the name of the genus and / or species of 572 strains were referenced in the Bruker database while 406 were present in the VITEK MS IVD database. The Biotyper correctly identified 494 (86.4%) of the strains, while the VITEK MS correctly identified 362 (92.3%) of the strains (excluding 14 mycobacteria that were not tested). Of the 70 strains not present in the Bruker database at the species level, the Biotyper correctly identified 10 (14.3%) to the genus level and 2 (2.9%) to the complex/group level. For 52 (74.2%) strains, we obtained no identification, and an incorrect identification was given for 6 (8.6%) strains. Of the 178 strains not present in the VITEK MS IVD database at the species level (excluding 71 untested mycobacteria and actinomycetes), the VITEK MS correctly identified 12 (6.8%) of the strains each to the genus and to the complex/group level. For 97 (54.5%) strains, no identification was given and for 69 (38.7%) strains, an incorrect identification was obtained. Our study demonstrates that both systems gave a high level (above 85%) of correct identification for a wide range of PLOS ONE |

Since the primordial of humanity, pollen has been considered a good source of nutrients and energy. Its promising healing properties have also been referred to. The present study aimed to characterize, for the first time, eight commercial pollens from Portugal and Spain available on the market studying the legislation on labeling, pollinic origin, physicochemical and microbiological analyses and identification of yeasts. Eleven botanical families were found amongst the samples. The most abundant family and the most dominant pollen was Cistaceae. The moisture content, ash, a w , pH, reducing sugars, carbohydrates, proteins, lipids and energy were analyzed and the specific parameters were within the specifications required by some countries with legislation regarding these parameters. Microbiologically commercial pollen showed acceptable safety for the commercial quality and hygiene. All samples showed negative results for toxigenic species. The microorganisms studied were aerobic mesophiles, yeasts and moulds, coliforms, Escherichia coli, Staphylococcus aureus, Salmonella and sulfite-reducing Clostridium. During the work, six yeasts species were isolated from pollen, with Rhodotorula mucilaginosa being the most abundant, as it was present in four samples.

We have investigated the bacteria and yeast ecology of the typical Italian Ciauscolo salami that is produced in Central Italy using a polyphasic approach based on culture-dependent and -independent methods. The physico-chemical analyses showed a progressive drop in pH and water activity (a w ) during ripening. The viable counts revealed a dominance of lactic acid bacteria (LAB) over coagulase negative cocci (CNC) and yeasts. From the molecular identification of the isolates, the prevalence of Lactobacillus curvatus, Lb. plantarum and Staphylococcus xylosus was shown among the bacteria, while Debaryomyces hansenii was the prevalent species among the yeasts, and it was isolated throughout the whole ripening process. Minority species, namely Rhodotorula mucillaginosa and Trichosporon brassicae, were also recovered from the meat batter.

Recombinant DNA (rDNA) technologies allow the production of a wide range of peptides, proteins and metabolites from naturally non-producing cells. Since human insulin was the first heterologous compound produced in a laboratory in 1977, rDNA technology has become one of the most important technologies developed in the 20th century. Recombinant protein and metabolites production is a multi-billion dollar market. The development of a new product begins with the choice of the cell factory. The final application of the compound dictates the main criteria that should be taken into consideration: (1) quality, (2) quantity, (3) yield and (4) space time yield of the desired product. Quantity and quality are the most predominant requirements that must be considered for the commercial production of a protein. Quantity and yield are the requirements for the production of a metabolite. Finally, space time yield is crucial for any production process. It therefore becomes clear why the perfect host does not exist yet, and why-despite important advances in rDNA applications in higher eukaryotic cells-microbial biodiversity continues to represent a potential source of attractive cell factories. In this review, we compare the advantages and limitations of the principal yeast and bacterial workhorse systems.

The microbial diversity of water kefir, made from a mixture of water, dried figs, a slice of lemon and sucrose was studied. The microbial consortia residing in the granules of three water kefirs of different origins were analyzed. A collection of 453 bacterial isolates was obtained on different selective/differential media. Bacterial isolates were grouped with randomly amplified polymorphic DNA (RAPD)-PCR analyses. One representative of each RAPD genotype was identified by comparative 16S rDNA gene sequencing. The predominant genus in water kefirs I and II was Lactobacillus, which accounted for 82.1% in water kefir I and 72.1% in water kefir II of the bacterial isolates. The most abundant species in water kefirs I and II were Lactobacillus hordei and Lb. nagelii followed by considerably lower numbers of Lb. casei. Other lactic acid bacteria (LAB) were identified as Leuconostoc mesenteroides and Lc. citreum in all three water kefirs. The most abundant species in water kefir III was Lc. mesenteroides (28%) and Lc. citreum (24.3%). A total of 57 LAB belonging to the species of Lb. casei, Lb. hordei, Lb. nagelii, Lb. hilgardii and Lc. mesenteroides were able to produce exopolysacchrides from sucrose. Non LABs were identified as Acetobacter fabarum and Ac. orientalis. The Acetobacter species were more prevalent in consortium III. Cluster analyses of RAPD-PCR patterns revealed an interspecies diversity among the Lactobacillus and Acetobacter strains. Aditionally, Saccharomyces cerevisiae, Lachancea fermentati, Hanseniaospora valbyensis and Zygotorulaspora florentina were isolated and identified by comparison of partial 26S rDNA sequences and FTIR spectroscopy.

Soybeans soaked in tap water for 24 to 36 h at 20, 30 or 37 °C underwent a natural fermentation that was characterized by the growth of microorganisms to 10 s 10 l° cfu/ml (depending on temperature) and a reduction of pH from 6.5 to 4.5. Lactobacillus casei. Streptococcus faecium, Staphylococcus epidermidis and Streptococcus dysgalactiae dominated the fermentation but, significant contributions were also made by Klebsiella pneumoniae, Klebsiella ozaenae, Enterobacter cloacae. Enterobacter agglomerans, Citrobacter diversus and Bacillus breuis, and the yeasts Pichia burtonii, Candida diddensiae and Rhodotorula rubra. Fermentation of surface-decontaminated beans in sterile water with pure cultures of these isolates showed L. casei, Strep. faecium and Staph. epidermidis to be the main species responsible for the pH reduction. Soybeans were the main source of microorganisms for the fermentation. Boiled beans did not undergo an acid fermentation.

To obtain insight into the genome-wide transcriptional response of heterologous carotenoid production in Saccharomyces cerevisiae, the transcriptome of two different S. cerevisiae strains overexpressing carotenogenic genes from the yeast Xanthophyllomyces dendrorhous grown in carbon-limited chemostat cultures was analysed. The strains exhibited different absolute carotenoid levels as well as different intermediate profiles. These discrepancies were further sustained by the difference of the transcriptional response exhibited by the two strains. Transcriptome analysis of the strain producing high carotenoid levels resulted in specific induction of genes involved in pleiotropic drug resistance (PDR). These genes encode ABC-type and major facilitator transporters which are reported to be involved in secretion of toxic compounds out of cells. β-Carotene was found to be secreted when sunflower oil was added to the medium of S. cerevisiae cells producing high levels of carotenoids, which was not observed when added to X. dendrorhous cells. Deletion of pdr10, one of the induced ABC transporters, decreased the transformation efficiency of a plasmid containing carotenogenic genes. The few transformants that were obtained had decreased growth rates and lower carotenoid production levels compared to a pdr5 deletion and a reference strain transformed with the same genes. Our results suggest that production of high amounts of carotenoids in S. cerevisiae leads to membrane stress, in which Pdr10 might play an important role, and a cellular response to secrete carotenoids out of the cell.

Bacteria, yeasts and filamentous fungi were isolated during natural coffee processing. Bacteria were isolated in greater numbers at the beginning of the fermentation, when the moisture of the coffee beans was around 68%. Gram-positive bacteria represented 85.5% of all bacteria isolated, and Bacillus was the predominant genus (51%). Gram-negative species of the genera Serratia, Enterobacter and Acinetobacter were also found. Approximately 22% of 940 randomly chosen isolates of microorganisms were yeasts. Debaryomyces (27%), Pichia (18.9%) and Candida (8.0%) were the most commonly found genera, and these three genera tended to appear more often as the fruit was fermented and dried. Aspergillus was the most abundant genus besides Penicillium, Fusarium and Cladosporium, with 42.6% of the total fungi isolates. The genera and species identified included members known to have pectinase and cellulase activities. Of the 10 organic acids analyzed and quantified in coffee beans, acetic and lactic acids may have been generated by microbial activity. Butyric acid was not detected in any sample. (R.F. Schwan).

In this study we investigated yeast biodiversity and dynamics during the production of a sweet wine obtained from dried grapes. Two wineries were selected in the Collio region and grapes, grape juices and wines during fermentations were analyzed by culture-dependent methods (plating on WLN medium) and cultureindependent methods (PCR-DGGE). Moreover, the capability of the Saccharomyces cerevisiae starter cultures to take over the fermentation was assessed by RAPD-PCR. On WLN agar several species of non-Saccharomyces yeasts (Hanseniaspora, Metschnikowia, Pichia, Candida, Torulaspora and Debaryomyces), but also strains of S. cerevisiae, were isolated. After inoculation of the starter cultures, only colonies typical of S. cerevisiae were observed. Using PCR-DGGE, the great biodiversity of moulds on the grapes was underlined, both at the DNA and RNA level, while the yeast contribution started to become important only in the musts. Here, bands belonging to species of Candida zemplinina and Hanseniaspora uvarum were visible. Lastly, when the S. cerevisiae isolates were compared by RAPD-PCR, it was determined that only in one of the fermentations followed, the inoculated strain conducted the alcoholic fermentation. In the second fermentation, the starter culture was not able to promptly implant and other populations of S. cerevisiae could be isolated, most likely contributing to the final characteristics of the sweet wine produced.

Some oenological practices, such as the massive utilisation of commercial yeast and the consequent colonisation of wineries, can contribute to reducing the native yeast biodiversity. In this context, the vineyard could be a reservoir of autochthonous yeasts of oenological interest. Thus, the evaluation of the influence of different agricultural parameters on the biodiversity of yeast population in the vineyard is necessary. This work shows the results of the influence of some floor management strategies of the vineyard in the natural yeast population associated with the grape-berries. With this objective, a three year sampling plan was designed in the Shiraz vineyards of the Madrid region using three floor management strategies: bare soil by tillage, bare soil maintained with herbicides and soil maintained with cover crop. The results of this study have shown that bare soil by tillage could be a sustainable alternative for managing the soil, due to the reduced use of agrochemicals and the resulting high yeasts biodiversity. Nevertheless, the presence of herbicides in the vineyard has a minor impact on the diversity of grape associated yeast communities, and this could have increased the yeast populations. Hence, from the fermentative yeasts' (like Saccharomyces) point of view, in hot and arid environments where soils may be affected by the tillage management, the best option could be the maintenance of the bare soil with the use of herbicides.

The microbial populations associated with the different sections of broccoli harvested from three locations in Australia were studied during storage at 5, 15 and 208C. Bacterial and yeast populations associated with the outer florets and cut surfaces of the stem were generally 10-fold or more higher than those associated with inner florets or non-cut stems, respectively. The predominating bacterial species varied with the origin of the broccoli. Pseudomonas fluorescens, Ps. 5 7 corrugata and Ps. viridiflava predominated at populations of 10 -10 cfu / g on broccoli harvested from Victoria, Ps. 3 6 fluorescens, Ps. mendocina and Ps. fragii and Arthrobacter spp. (10 -10 cfu / g) were prevalent on broccoli harvested from Queensland. Broccoli harvested from New South Wales exhibited a predominance of Ps. fluorescens, Arthrobacter spp. and 3 5

The aim of the present study was the microbiological and technological characterization of laboratorymade sourdoughs for use in barley-flour-based bread-making. A defined multi-strain starter culture consisting of selected lactic acid bacteria (LAB) and yeasts from wheat sourdoughs was inoculated into three flour-water mixtures, composed of: (i) 100% wheat flour (ii) 50% wheat flour and 50% hull-less barley flour (composite flour); (iii) 100% hull-less barley flour. After two months of continuous propagation, the chemical characteristics of the three sourdoughs were investigated by measuring: pH, total titratable acidity and concentrations of various microbial metabolites by HPLC (i.e. lactic, acetic, phenyllactic and butyric acids and diacetyl). The microbial traits were studied through viable counts, isolation and typing of LAB and yeasts and PCR-DGGE analyses. Only Saccharomyces cerevisiae and Lactobacillus plantarum were detectable in the sourdoughs together with other lactobacilli species which were different depending on the type of flour blend used. The molecular typing of the isolates highlighted that only a few strains among those initially inoculated prevailed. The volume increases of the three types of sourdough were also investigated and a correlation was seen between an increase in the barley flour content and a reduction in the dough volume.

In winemaking, nutrient supplementation is a common practice for optimising fermentation and producing quality
wine. Nutritionally suboptimal grape juices are often enriched with nutrients in order to manipulate the production
of yeast aroma compounds. Nutrients are also added to active dry yeast (ADY) rehydration media to enhance
subsequent fermentation performance. In this study we demonstrate that nutrient supplementation at rehydration
also has a significant effect on the formation of volatile sulfur compounds during wine fermentations. The
concentration of the ‘fruity’ aroma compounds, the polyfunctional thiols 3-mercaptohexan-1-ol (3MH) and 3-
mercaptohexyl acetate (3MHA), was increased while the concentration of the ‘rotten egg’ aroma compound,
hydrogen sulfide (H2S), was decreased. Nutrient supplementation of the rehydration media also changed the
kinetics of H2S production during fermentation by advancing onset of H2S production. Microarray analysis revealed
that this was not due to expression changes within the sulfate assimilation pathway, which is known to be a major
contributor to H2S production. To gain insight into possible mechanisms responsible for this effect, a component
of the rehydration nutrient mix, the tri-peptide glutathione (GSH) was added at rehydration and studied for its
subsequent effects on H2S formation. GSH was found to be taken up during rehydration and to act as a source for
H2S during the following fermentation. These findings represent a potential approach for managing sulfur aroma
production through the use of rehydration nutrients.

NAD + -dependent formate dehydrogenase (FDH, EC 1.2.1.2) is one of the best enzymes for the purpose of NADH regeneration in dehydrogenase-based synthesis of optically active compounds. Low operational stability and high production cost of native FDHs limit their application in commercial production of chiral compounds. The review summarizes the results on engineering of bacterial and yeast FDHs aimed at improving their chemical and thermal stability, catalytic activity, switch in coenzyme specificity from NAD + to NADP + and overexpression in Escherichia coli cells.

mRNA synthesis in all organisms is performed by RNA polymerases, which work as nanomachines on DNA templates. The rate at which their product is made is an important parameter in gene expression. Transcription rate encompasses two related, yet different, concepts: the nascent transcription rate, which measures the in situ mRNA production by RNA polymerase, and the rate of synthesis of mature mRNA, which measures the contribution of transcription to the mRNA concentration. Both parameters are useful for molecular biologists, but they are not interchangeable and they are expressed in different units. It is important to distinguish when and where each one should be used. We propose that for functional genomics the use of nascent transcription rates should be restricted to the evaluation of the transcriptional process itself, whereas mature mRNA synthesis rates should be employed to address the transcriptional input to mRNA concentration balance leading to variation of gene expression.

Eight non-Saccharomyces wine strains, previously selected for their ability to modulate the final concentrations of various volatile compounds and to persist with Saccharomyces cerevisiae in mixed inocula fermentations of grape juice, have been analyzed in the present work to test their ability to release mannoproteins. The eight strains were members of different genera originally isolated from grape: Hansensiaspora osmophila, Lachancea thermotolerans, Metschnikowia pulcherrima, Pichia fermentans, Saccharomycodes ludwigii, Starmerella bacillaris, Torulaspora delbrueckii and Zygosaccharomyces florentinus. A synthetic polysaccharide-free grape juice, was used to characterize the mannoproteins released during the alcoholic fermentation. Mannoproteins profiles were characterized by gel electrophoresis and carbohydrate composition was analyzed both by HPLC and by mass spectrometry. The eight non-Saccharomyces yeasts demonstrated a higher capacity to release polysaccharides compared to S. cerevisiae. The proteins released by the eight yeast strains showed a wide variety of protein sizes, ranging from 25 kDa to greater than 250 kDa. The mass spectrometric profile of the N-glycans ranged from 1600 to 4000 Da and was characteristic for each strain. Detailed investigation of the degree of polymerization of released N-glycans revealed variable composition from 8 to 15 units of monosaccharides. Food Microbiology 43 (2014) 5e15 P. Domizio et al. / Food Microbiology 43 (2014) 5e15

Twenty-one yeast-like microorganisms were isolated from tannery effluents and from a nickel-copper mine in Argentina. They were tested for their Cu(II), Ni(II), Cd(II) and Cr(VI) tolerance in qualitative assays on solid medium. Three isolates were selected for their multiple tolerance to the different heavy metals and highest tolerance to Cr(VI). According to morphological and physiological analysis and 26S rDNA D1/D2 domain sequences the isolates were characterized as: Lecythophora sp. NGV-1, Candida sp. NGV-9 and Aureobasidium pullulans VR-8. Resistance of the three strains to high Cr(VI) concentrations and their ability to remove Cr(VI) were assessed using YNB-glucose medium supplemented with 0.5 and 1 mM Cr(VI). Chromate removal activity was estimated by measuring remaining Cr(VI) concentration in the supernatant using the colorimetric 1,5-diphenylcarbazide method and total chromium was determined by flame atomic absorption spectroscopy. The results indicate that the initial Cr(VI) concentration negatively influenced growth and the specific growth rate but stimulated the metabolic activity of the three strains; resistance to Cr(VI) by these strains was mainly due to reduction of Cr(VI) rather than chromium bioaccumulation. This study showed the potential ability of these strains as tools for bioremediation of Cr(VI) from contaminated sites.

The present work was designed to assess the antimicrobial activity of the methanolic extracts of the bark of Ficus racemosa L. and root of Cisampelos pareira L. var. hirsuta (DC) Forman. The antimicrobial activity of the extracts was assessed by disc diffusion method using four microbial strains. Many plants of the family Moraceae are used in the treatment of infectious diseases. Ficus racemosa, belongs to this family, is used traditionally against dyspepsia, dermal infections, abdominal pains and diarrhoea. Similarly, Cissampelos pareira, a plant belongs to family Minispermaceae found to have analgesic, antiinflammatory and febrifuge properties. In the present study, both the plant extracts exhibited the antimicrobial effect against four strains of Escherichia coli, Pseudomonas aeroginosa, Staphylococcus aureus and Bacillus subtilis. The effects were compared with that of the standard antibiotic loaded discs, Ofloxacin 5 µg/disc. Finally, the petridishes were incubated at 37 0 C for 24 hours. After the incubation period the zone of inhibition produced by samples and standard were measured. Comparatively, the activity of Cissampelos pareira was found better than that of Ficus racemosa. But, in case of P. aeuroginosa, it showed less activity than Ficus racemosa. Cissampelos pareira showed better activity against B. subtilis which is comparable with that of the standard drug Ofloxacin. It was clearly observed that methanolic extracts of both the herbals exhibit impact antimicrobial activity against B. subtilis.

Aims: To establish the site of microbial growth on naturally black fermented table olives, and to monitor the population dynamics of yeasts and selected micro-organisms together with the changes in organic acid profile and pH in the cover brine during fermentation. Methods and Results: During fermentation, the numbers of Enterobacteriaceae and Pseudomonas spp. in the brine decreased whilst lactic acid bacteria and yeast populations increased. Scanning electron microscopy showed that a yeast-rich biofilm developed on the epicuticular wax of the olive skin during fermentation. Yeasts also predominated in the stomatal openings, but bacteria were more numerous in intercellular spaces in the sub-stomatal flesh. Citric, malic and tartaric acids were the major organic acids accumulating in the brine during fermentation. Conclusions: Micro-organisms associated with the skin, stomata and flesh in fermenting black olives may experience different local conditions to those prevailing in the cover brine. Significant and Impact of the Study: These are the first observations of the microorganisms associated with the fruit of naturally fermented black olives and of the accumulation of specific organic acids during fermentation.

In this study, four different laboratory scale gluten-free (GF) sourdoughs were developed from buckwheat or teff flours. The fermentations were initiated by the spontaneous biota of the flours and developed under two technological conditions (A and B). Sourdoughs were propagated by continuous back-slopping until the stability was reached. The composition of the stable biota occurring in each sourdough was assessed using both culture-dependent and-independent techniques. Overall, a broad spectrum of lactic acid bacteria (LAB) and yeasts species, belonging mainly to the genera Lactobacillus, Pediococcus, Leuconostoc, Kazachstania and Candida, were identified in the stable sourdoughs. Buckwheat and teff sourdoughs were dominated mainly by obligate or facultative heterofermentative LAB, which are commonly associated with traditional wheat or rye sourdoughs. However, the spontaneous fermentation of the GF flours resulted also in the selection of species which are not consider endemic to traditional sourdoughs, i.e. Pediococcus pentosaceus, Leuconostoc holzapfelii, Lactobacillus gallinarum, Lactobacillus vaginalis, Lactobacillus sakei, Lactobacillus graminis and Weissella cibaria. In general, the composition of the stable biota was strongly affected by the fermentation conditions, whilst Lactobacillus plantarum dominated in all buckwheat sourdoughs. Lactobacillus pontis is described for the first time as dominant species in teff sourdough. Among yeasts, Saccharomyces cerevisiae and Candida glabrata dominated teff sourdoughs, whereas the solely Kazachstania barnetti was isolated in buckwheat sourdough developed under condition A. This study allowed the identification and isolation of LAB and yeasts species which are highly competitive during fermentation of buckwheat or teff flours. Representatives of these species can be selected as starters for the production of sourdough destined to GF bread production.