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Sample records for viable cell yield

  1. Viable cell yield from active dry yeast products and effects of storage temperature and diluent on yeast cell viability.

    Science.gov (United States)

    Sullivan, M L; Bradford, B J

    2011-01-01

    Active dry yeast (ADY) products are commonly fed in the dairy industry, but research regarding quality control for such products is limited. The objectives of this study were to determine yeast viability in field samples relative to manufacturers' guarantees (experiment 1), measure the effects of high-temperature storage on yeast viability (experiment 1), and determine the effect of vitamin-trace mineral (VTM) premix on yeast viability (experiment 2). Commercially available ADY products were acquired in triplicate through normal distribution channels and stored at 4°C upon receipt. Initial samples were evaluated for colony-forming units and compared with product label guarantees. Only 1 of the 6 products sampled in experiment 1 met product guarantees for all 3 samples. To determine effects of storage temperature and duration on viability, ADY samples were stored in an incubator at 40°C with ambient humidity for 1, 2, and 3 mo. High-temperature storage significantly decreased viability over the 3-mo period; approximately 90% of viable cells were lost each month. Three of the 5 products sampled in experiment 2 met product guarantees. Fresh samples of 4 of these 5 ADY products were mixed in duplicate with ground corn (GC) or a VTM premix to achieve a target concentration of 2.2×10(8) cfu/g. For each product, GC and VTM samples were stored at ambient temperature (22°C) and at an elevated temperature (40°C) for 2 wk. No differences in viable yeast count were observed between GC and VTM samples immediately after mixing or after storage at ambient temperature. Yeast viability in GC and VTM samples decreased during storage at an elevated temperature. There also was a significant interaction of diluent and storage temperature; VTM samples had higher cell viability than GC samples when subjected to high-temperature storage. Results suggest that (1) ADY products failed to consistently meet product guarantees; (2) viability of ADY products was greatly diminished during

  2. Deletion of ultraconserved elements yields viable mice

    Energy Technology Data Exchange (ETDEWEB)

    Ahituv, Nadav; Zhu, Yiwen; Visel, Axel; Holt, Amy; Afzal, Veena; Pennacchio, Len A.; Rubin, Edward M.

    2007-07-15

    Ultraconserved elements have been suggested to retainextended perfect sequence identity between the human, mouse, and ratgenomes due to essential functional properties. To investigate thenecessities of these elements in vivo, we removed four non-codingultraconserved elements (ranging in length from 222 to 731 base pairs)from the mouse genome. To maximize the likelihood of observing aphenotype, we chose to delete elements that function as enhancers in amouse transgenic assay and that are near genes that exhibit markedphenotypes both when completely inactivated in the mouse as well as whentheir expression is altered due to other genomic modifications.Remarkably, all four resulting lines of mice lacking these ultraconservedelements were viable and fertile, and failed to reveal any criticalabnormalities when assayed for a variety of phenotypes including growth,longevity, pathology and metabolism. In addition more targeted screens,informed by the abnormalities observed in mice where genes in proximityto the investigated elements had been altered, also failed to revealnotable abnormalities. These results, while not inclusive of all thepossible phenotypic impact of the deleted sequences, indicate thatextreme sequence constraint does not necessarily reflect crucialfunctions required for viability.

  3. High speed flow cytometric separation of viable cells

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    Sasaki, Dennis T.; Van den Engh, Gerrit J.; Buckie, Anne-Marie

    1995-01-01

    Hematopoietic cell populations are separated to provide cell sets and subsets as viable cells with high purity and high yields, based on the number of original cells present in the mixture. High-speed flow cytometry is employed using light characteristics of the cells to separate the cells, where high flow speeds are used to reduce the sorting time.

  4. Viable Cell Culture Banking for Biodiversity Characterization and Conservation.

    Science.gov (United States)

    Ryder, Oliver A; Onuma, Manabu

    2018-02-15

    Because living cells can be saved for indefinite periods, unprecedented opportunities for characterizing, cataloging, and conserving biological diversity have emerged as advanced cellular and genetic technologies portend new options for preventing species extinction. Crucial to realizing the potential impacts of stem cells and assisted reproductive technologies on biodiversity conservation is the cryobanking of viable cell cultures from diverse species, especially those identified as vulnerable to extinction in the near future. The advent of in vitro cell culture and cryobanking is reviewed here in the context of biodiversity collections of viable cell cultures that represent the progress and limitations of current efforts. The prospects for incorporating collections of frozen viable cell cultures into efforts to characterize the genetic changes that have produced the diversity of species on Earth and contribute to new initiatives in conservation argue strongly for a global network of facilities for establishing and cryobanking collections of viable cells.

  5. Surface Charge Visualization at Viable Living Cells.

    Science.gov (United States)

    Perry, David; Paulose Nadappuram, Binoy; Momotenko, Dmitry; Voyias, Philip D; Page, Ashley; Tripathi, Gyanendra; Frenguelli, Bruno G; Unwin, Patrick R

    2016-03-09

    Scanning ion conductance microscopy (SICM) is demonstrated to be a powerful technique for quantitative nanoscale surface charge mapping of living cells. Utilizing a bias modulated (BM) scheme, in which the potential between a quasi-reference counter electrode (QRCE) in an electrolyte-filled nanopipette and a QRCE in bulk solution is modulated, it is shown that both the cell topography and the surface charge present at cellular interfaces can be measured simultaneously at high spatial resolution with dynamic potential measurements. Surface charge is elucidated by probing the properties of the diffuse double layer (DDL) at the cellular interface, and the technique is sensitive at both low-ionic strength and under typical physiological (high-ionic strength) conditions. The combination of experiments that incorporate pixel-level self-referencing (calibration) with a robust theoretical model allows for the analysis of local surface charge variations across cellular interfaces, as demonstrated on two important living systems. First, charge mapping at Zea mays root hairs shows that there is a high negative surface charge at the tip of the cell. Second, it is shown that there are distinct surface charge distributions across the surface of human adipocyte cells, whose role is the storage and regulation of lipids in mammalian systems. These are new features, not previously recognized, and their implications for the functioning of these cells are highlighted.

  6. Inkjet printing of viable human dental follicle stem cells

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    Mau Robert

    2015-09-01

    Full Text Available Inkjet printing technology has the potential to be used for seeding of viable cells for tissue engineering approaches. For this reason, a piezoelectrically actuated, drop-on-demand inkjet printing system was applied to deliver viable human dental follicle stem cells (hDFSC of sizes of about 15 μm up to 20 μm in diameter. The purpose of these investigations was to verify the stability of the printing process and to evaluate cell viability post printing. Using a Nanoplotter 2.1 (Gesim, Germany equipped with the piezoelectric printhead NanoTip HV (Gesim, Germany, a concentration of 6.6 ×106 cells ml−1 in DMEM with 10% fetal calf serum (FCS could be dispensed. The piezoelectric printhead has a nominal droplet volume of ~ 400 pl and was set to a voltage of 75 V and a pulse of 50 μs while dosing 50 000 droplets over a time of 100 seconds. The volume and trajectory of the droplet were checked by a stroboscope test right before and after the printing process. It was found that the droplet volume decreases significantly by 35% during printing process, while the trajectory of the droplets remains stable with only an insignificant number of degrees deviation from the vertical line. It is highly probable that some cell sedimentations or agglomerations affect the printing performance. The cell viability post printing was assessed by using the Trypan Blue dye exclusion test. The printing process was found to have no significant influence on cell survival. In conclusion, drop-on-demand inkjet printing can be a potent tool for the seeding of viable cells.

  7. The molecularly crowded cytoplasm of bacterialcCells : Dividing cells contrasted with viable but non-culturable (VBNC) bacterial cells

    NARCIS (Netherlands)

    Trevors, J. T.; van Elsas, J. D.; Bej, A. K.

    2013-01-01

    In this perspective, we discuss the cytoplasm in actively growing bacterial cells contrasted with viable but non-culturable (VBNC) cells. Actively growing bacterial cells contain a more molecularly crowded and organized cytoplasm, and are capable of completing their cell cycle resulting in cell

  8. Non-viable antagonist cells are associated with reduced biocontrol performance by viable cells of the yeast Papiliotrema flavescens against Fusarium head blight of wheat.

    Science.gov (United States)

    Microbially-based plant disease control products have achieved commercial market success, but the efficacy of such biocontrol products is sometimes deemed inconsistent. Improper processing of harvested microbial biomass or long-term storage can reduce the proportion of viable cells and necessitate t...

  9. Modelling the number of viable vegetative cells of Bacillus cereus passing through the stomach

    NARCIS (Netherlands)

    Wijnands, L.M.; Pielaat, A.; Dufrenne, J.B.; Zwietering, M.H.; Leusden, van F.M.

    2009-01-01

    Aims: Model the number of viable vegetative cells of B. cereus surviving the gastric passage after experiments in simulated gastric conditions. Materials and Methods: The inactivation of stationary and exponential phase vegetative cells of twelve different strains of Bacillus cereus, both mesophilic

  10. Combining ethidium monoazide treatment with real-time PCR selectively quantifies viable Batrachochytrium dendrobatidis cells.

    Science.gov (United States)

    Blooi, Mark; Martel, An; Vercammen, Francis; Pasmans, Frank

    2013-02-01

    Detection of the lethal amphibian fungus Batrachochytrium dendrobatidis relies on PCR-based techniques. Although highly accurate and sensitive, these methods fail to distinguish between viable and dead cells. In this study a novel approach combining the DNA intercalating dye ethidium monoazide (EMA) and real-time PCR is presented that allows quantification of viable B. dendrobatidis cells without the need for culturing. The developed method is able to suppress real-time PCR signals of heat-killed B. dendrobatidis zoospores by 99.9 % and is able to discriminate viable from heat-killed B. dendrobatidis zoospores in mixed samples. Furthermore, the novel approach was applied to assess the antifungal activity of the veterinary antiseptic F10(®) Antiseptic Solution. This disinfectant killed B. dendrobatidis zoospores effectively within 1 min at concentrations as low as 1:6400. Copyright © 2013 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  11. A simple way to identify non-viable cells within living plant tissue using confocal microscopy

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    Truernit Elisabeth

    2008-06-01

    Full Text Available Abstract Background Plant cell death is a normal process during plant development. Mutant plants may exhibit misregulation of this process, which can lead to severe growth defects. Simple ways of visualising cell death in living plant tissues can aid the study of plant development and physiology. Results Spectral variants of the fluorescent SYTOX dyes were tested for their usefulness for the detection of non-viable cells within plant embryos and roots using confocal laser-scanning microscopy. The dyes were selective for non-viable cells and showed very little background staining in living cells. Simultaneous detection of SYTOX dye and fluorescent protein (e.g. GFP fluorescence was possible. Conclusion The fluorescent SYTOX dyes are useful for an easy and quick first assay of plant cell viability in living plant samples using fluorescence and confocal laser-scanning microscopy.

  12. Separable Bilayer Microfiltration Device for Viable Label-free Enrichment of Circulating Tumour Cells

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    Zhou, Ming-Da; Hao, Sijie; Williams, Anthony J.; Harouaka, Ramdane A.; Schrand, Brett; Rawal, Siddarth; Ao, Zheng; Brennaman, Randall; Gilboa, Eli; Lu, Bo; Wang, Shuwen; Zhu, Jiyue; Datar, Ram; Cote, Richard; Tai, Yu-Chong; Zheng, Si-Yang

    2014-12-01

    The analysis of circulating tumour cells (CTCs) in cancer patients could provide important information for therapeutic management. Enrichment of viable CTCs could permit performance of functional analyses on CTCs to broaden understanding of metastatic disease. However, this has not been widely accomplished. Addressing this challenge, we present a separable bilayer (SB) microfilter for viable size-based CTC capture. Unlike other single-layer CTC microfilters, the precise gap between the two layers and the architecture of pore alignment result in drastic reduction in mechanical stress on CTCs, capturing them viably. Using multiple cancer cell lines spiked in healthy donor blood, the SB microfilter demonstrated high capture efficiency (78-83%), high retention of cell viability (71-74%), high tumour cell enrichment against leukocytes (1.7-2 × 103), and widespread ability to establish cultures post-capture (100% of cell lines tested). In a metastatic mouse model, SB microfilters successfully enriched viable mouse CTCs from 0.4-0.6 mL whole mouse blood samples and established in vitro cultures for further genetic and functional analysis. Our preliminary studies reflect the efficacy of the SB microfilter device to efficiently and reliably enrich viable CTCs in animal model studies, constituting an exciting technology for new insights in cancer research.

  13. Use of propidium monoazide for selective profiling of viable microbial cells during Gouda cheese ripening

    NARCIS (Netherlands)

    Erkus, O.; Jager, V.C. de; Geene, R.T.; Alen-Boerrigter, I.J. van; Hazelwood, L.; Hijum, S.A.F.T. van; Kleerebezem, M; Smid, E.J.

    2016-01-01

    DNA based microbial community profiling of food samples is confounded by the presence of DNA derived from membrane compromised (dead or injured) cells. Selective amplification of DNA from viable (intact) fraction of the community by propidium monoazide (PMA) treatment could circumvent this problem.

  14. In vitro and in vivo bioluminescent quantification of viable stem cells in engineered constructs.

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    Logeart-Avramoglou, Delphine; Oudina, Karim; Bourguignon, Marianne; Delpierre, Laetitia; Nicola, Marie-Anne; Bensidhoum, Morad; Arnaud, Eric; Petite, Herve

    2010-06-01

    Bioluminescent quantification of viable cells inside three-dimensional porous scaffolds was performed in vitro and in vivo. The assay quantified the bioluminescence of murine stem (C3H10T1/2) cells tagged with the luciferase gene reporter and distributed inside scaffolds of either soft, translucent, AN69 polymeric hydrogel or hard, opaque, coral ceramic materials. Quantitative evaluation of bioluminescence emitted from tagged cells adhering to these scaffolds was performed in situ using either cell lysates and a luminometer or intact cells and a bioluminescence imaging system. Despite attenuation of the signal when compared to cells alone, the bioluminescence correlated with the number of cells (up to 1.5 x 10(5)) present on each material scaffold tested, both in vitro and noninvasively in vivo (subcutaneous implants in the mouse model). The noninvasive bioluminescence measurement technique proved to be comparable to the cell-destructive bioluminescence measurement technique. Monitoring the kinetics of luciferase expression via bioluminescence enabled real-time assessment of cell survival and proliferation on the scaffolds tested over prolonged (up to 59 days) periods of time. This novel, sensitive, easy, fast-to-implement, quantitative bioluminescence assay has great, though untapped, potential for screening and determining noninvasively the presence of viable cells on biomaterial constructs in the tissue engineering and tissue regeneration fields.

  15. Mobilization of Viable Tumor Cells Into the Circulation During Radiation Therapy

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Olga A. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Molecular Radiation Biology Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); The Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC (Australia); Anderson, Robin L. [The Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC (Australia); Metastasis Research Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Russell, Prudence A. [Department of Anatomical Pathology, St. Vincent Hospital, Fitzroy, VIC (Australia); Ashley Cox, R. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Ivashkevich, Alesia [Molecular Radiation Biology Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Laboratory of DNA Repair and Genomics, Centre for Innate Immunity and Infectious Disease, Monash Institute for Medical Research, Monash University, Clayton, VIC (Australia); Swierczak, Agnieszka; Doherty, Judy P. [Metastasis Research Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Jacobs, Daphne H.M. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Smith, Jai [Molecular Radiation Biology Laboratory, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Siva, Shankar; Daly, Patricia E. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); Ball, David L. [Division of Radiation Oncology and Cancer Imaging, Peter MacCallum Cancer Centre, East Melbourne, VIC (Australia); The Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC (Australia); and others

    2014-02-01

    Purpose: To determine whether radiation therapy (RT) could mobilize viable tumor cells into the circulation of non-small cell lung cancer (NSCLC) patients. Methods and Materials: We enumerated circulating tumor cells (CTCs) by fluorescence microscopy of blood samples immunostained with conventional CTC markers. We measured their DNA damage levels using γ-H2AX, a biomarker for radiation-induced DNA double-strand breaks, either by fluorescence-activated cell sorting or by immunofluorescence microscopy. Results: Twenty-seven RT-treated NSCLC patients had blood samples analyzed by 1 or more methods. We identified increased CTC numbers after commencement of RT in 7 of 9 patients treated with palliative RT, and in 4 of 8 patients treated with curative-intent RT. Circulating tumor cells were also identified, singly and in clumps in large numbers, during RT by cytopathologic examination (in all 5 cases studied). Elevated γ-H2AX signal in post-RT blood samples signified the presence of CTCs derived from irradiated tumors. Blood taken after the commencement of RT contained tumor cells that proliferated extensively in vitro (in all 6 cases studied). Circulating tumor cells formed γ-H2AX foci in response to ex vivo irradiation, providing further evidence of their viability. Conclusions: Our findings provide a rationale for the development of strategies to reduce the concentration of viable CTCs by modulating RT fractionation or by coadministering systemic therapies.

  16. Desiccation induces viable but Non-Culturable cells in Sinorhizobium meliloti 1021.

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    Vriezen, Jan Ac; de Bruijn, Frans J; Nüsslein, Klaus R

    2012-01-20

    Sinorhizobium meliloti is a microorganism commercially used in the production of e.g. Medicago sativa seed inocula. Many inocula are powder-based and production includes a drying step. Although S. meliloti survives drying well, the quality of the inocula is reduced during this process. In this study we determined survival during desiccation of the commercial strains 102F84 and 102F85 as well as the model strain USDA1021.The survival of S. meliloti 1021 was estimated during nine weeks at 22% relative humidity. We found that after an initial rapid decline of colony forming units, the decline slowed to a steady 10-fold reduction in colony forming units every 22 days. In spite of the reduction in colony forming units, the fraction of the population identified as viable (42-54%) based on the Baclight live/dead stain did not change significantly over time. This change in the ability of viable cells to form colonies shows (i) an underestimation of the survival of rhizobial cells using plating methods, and that (ii) in a part of the population desiccation induces a Viable But Non Culturable (VBNC)-like state, which has not been reported before. Resuscitation attempts did not lead to a higher recovery of colony forming units indicating the VBNC state is stable under the conditions tested. This observation has important consequences for the use of rhizobia. Finding methods to resuscitate this fraction may increase the quality of powder-based seed inocula.

  17. Mathematical modelling of the viable epidermis: impact of cell shape and vertical arrangement

    KAUST Repository

    Wittum, Rebecca

    2017-12-07

    In-silico methods are valuable tools for understanding the barrier function of the skin. The key benefit is that mathematical modelling allows the interplay between cell shape and function to be elucidated. This study focuses on the viable (living) epidermis. For this region, previous works suggested a diffusion model and an approximation of the cells by hexagonal prisms. The work at hand extends this in three ways. First, the extracellular space is treated with full spatial resolution. This induces a decrease of permeability by about 10%. Second, cells of tetrakaidecahedral shape are considered, in addition to the original hexagonal prisms. For both cell types, the resulting membrane permeabilities are compared. Third, for the first time, the influence of cell stacking in the vertical direction is considered. This is particularly important for the stratum granulosum, where tight junctions are present.

  18. Progress in biocatalysis with immobilized viable whole cells: systems development, reaction engineering and applications.

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    Polakovič, Milan; Švitel, Juraj; Bučko, Marek; Filip, Jaroslav; Neděla, Vilém; Ansorge-Schumacher, Marion B; Gemeiner, Peter

    2017-05-01

    Viable microbial cells are important biocatalysts in the production of fine chemicals and biofuels, in environmental applications and also in emerging applications such as biosensors or medicine. Their increasing significance is driven mainly by the intensive development of high performance recombinant strains supplying multienzyme cascade reaction pathways, and by advances in preservation of the native state and stability of whole-cell biocatalysts throughout their application. In many cases, the stability and performance of whole-cell biocatalysts can be highly improved by controlled immobilization techniques. This review summarizes the current progress in the development of immobilized whole-cell biocatalysts, the immobilization methods as well as in the bioreaction engineering aspects and economical aspects of their biocatalytic applications.

  19. Trypan blue dye enters viable cells incubated with the pore-forming toxin HlyII of Bacillus cereus.

    Directory of Open Access Journals (Sweden)

    Seav-Ly Tran

    Full Text Available Trypan blue is a dye that has been widely used for selective staining of dead tissues or cells. Here, we show that the pore-forming toxin HlyII of Bacillus cereus allows trypan blue staining of macrophage cells, despite the cells remaining viable and metabolically active. These findings suggest that the dye enters viable cells through the pores. To our knowledge, this is the first demonstration that trypan blue may enter viable cells. Consequently, the use of trypan blue staining as a marker of vital status should be interpreted with caution. The blue coloration does not necessarily indicate cell lysis, but may rather indicate pore formation in the cell membranes and more generally increased membrane permeability.

  20. IN-VITRO BIOREDUCTION OF HEXAVALENT CHROMIUM BY VIABLE WHOLE CELLS OF Arthrobacter sp. SUK 1201

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    Satarupa Dey

    2014-08-01

    Full Text Available A chromium resistant and reducing bacterium Arthrobacter sp. SUK 1201 was isolated from chromite mine overburden dumps of Orissa, India. Viable whole cells of this isolate was capable of completely reducing 100 µM Cr(VI in chemically defined MS medium within 28 h of incubation under batch cultivation. Reduction of chromate increased with increased cell density and was maximum at a density of 1010 cells/ml, but the reduction potential of the suspended cells decreased with increase in Cr(VI concentration in the medium. Chromate reducing efficiency was promoted when glycerol and glucose was used as electron donors, while the optimum pH and temperature of Cr(VI reduction was found to be 7.0 and 35°C respectively. The reduction process was inhibited by divalent cations Ni, Co and Cd, but not by Cu and Fe. Similarly, carbonyl cyanide m-chlorophenylhydrazone (CCCP, N,N,-Di cyclohexyl carboiimide (DCC, sodium azide and sodium fluoride were inhibitory to chromate reduction, while in presence of 2,4 dinitrophenol (2,4 DNP chromate reduction by SUK 1201 cells remained unaffected.

  1. Yielding elastic tethers stabilize robust cell adhesion.

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    Matt J Whitfield

    2014-12-01

    Full Text Available Many bacteria and eukaryotic cells express adhesive proteins at the end of tethers that elongate reversibly at constant or near constant force, which we refer to as yielding elasticity. Here we address the function of yielding elastic adhesive tethers with Escherichia coli bacteria as a model for cell adhesion, using a combination of experiments and simulations. The adhesive bond kinetics and tether elasticity was modeled in the simulations with realistic biophysical models that were fit to new and previously published single molecule force spectroscopy data. The simulations were validated by comparison to experiments measuring the adhesive behavior of E. coli in flowing fluid. Analysis of the simulations demonstrated that yielding elasticity is required for the bacteria to remain bound in high and variable flow conditions, because it allows the force to be distributed evenly between multiple bonds. In contrast, strain-hardening and linear elastic tethers concentrate force on the most vulnerable bonds, which leads to failure of the entire adhesive contact. Load distribution is especially important to noncovalent receptor-ligand bonds, because they become exponentially shorter lived at higher force above a critical force, even if they form catch bonds. The advantage of yielding is likely to extend to any blood cells or pathogens adhering in flow, or to any situation where bonds are stretched unequally due to surface roughness, unequal native bond lengths, or conditions that act to unzip the bonds.

  2. Yielding Elastic Tethers Stabilize Robust Cell Adhesion

    Science.gov (United States)

    Whitfield, Matt J.; Luo, Jonathon P.; Thomas, Wendy E.

    2014-01-01

    Many bacteria and eukaryotic cells express adhesive proteins at the end of tethers that elongate reversibly at constant or near constant force, which we refer to as yielding elasticity. Here we address the function of yielding elastic adhesive tethers with Escherichia coli bacteria as a model for cell adhesion, using a combination of experiments and simulations. The adhesive bond kinetics and tether elasticity was modeled in the simulations with realistic biophysical models that were fit to new and previously published single molecule force spectroscopy data. The simulations were validated by comparison to experiments measuring the adhesive behavior of E. coli in flowing fluid. Analysis of the simulations demonstrated that yielding elasticity is required for the bacteria to remain bound in high and variable flow conditions, because it allows the force to be distributed evenly between multiple bonds. In contrast, strain-hardening and linear elastic tethers concentrate force on the most vulnerable bonds, which leads to failure of the entire adhesive contact. Load distribution is especially important to noncovalent receptor-ligand bonds, because they become exponentially shorter lived at higher force above a critical force, even if they form catch bonds. The advantage of yielding is likely to extend to any blood cells or pathogens adhering in flow, or to any situation where bonds are stretched unequally due to surface roughness, unequal native bond lengths, or conditions that act to unzip the bonds. PMID:25473833

  3. Viable bacteria associated with red blood cells and plasma in freshly drawn blood donations.

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    Christian Damgaard

    Full Text Available Infection remains a leading cause of post-transfusion mortality and morbidity. Bacterial contamination is, however, detected in less than 0.1% of blood units tested. The aim of the study was to identify viable bacteria in standard blood-pack units, with particular focus on bacteria from the oral cavity, and to determine the distribution of bacteria revealed in plasma and in the red blood cell (RBC-fraction.Cross-sectional study. Blood were separated into plasma and RBC-suspensions, which were incubated anaerobically or aerobically for 7 days on trypticase soy blood agar (TSA or blue lactose plates. For identification colony PCR was performed using primers targeting 16S rDNA.Blood donors attending Capital Region Blood Bank, Copenhagen University Hospital, Rigshospitalet, Hvidovre, Denmark, October 29th to December 10th 2013.60 donors (≥50 years old, self-reported medically healthy.Bacterial growth was observed on plates inoculated with plasma or RBCs from 62% of the blood donations. Growth was evident in 21 (35% of 60 RBC-fractions and in 32 (53% of 60 plasma-fractions versus 8 of 60 negative controls (p = 0.005 and p = 2.6x10-6, respectively. Propionibacterium acnes was found in 23% of the donations, and Staphylococcus epidermidis in 38%. The majority of bacteria identified in the present study were either facultative anaerobic (59.5% or anaerobic (27.8% species, which are not likely to be detected during current routine screening.Viable bacteria are present in blood from donors self-reported as medically healthy, indicating that conventional test systems employed by blood banks insufficiently detect bacteria in plasma. Further investigation is needed to determine whether routine testing for anaerobic bacteria and testing of RBC-fractions for adherent bacteria should be recommended.

  4. Increasing Vero viable cell densities for yellow fever virus production in stirred-tank bioreactors using serum-free medium.

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    Mattos, Diogo A; Silva, Marlon V; Gaspar, Luciane P; Castilho, Leda R

    2015-08-20

    In this work, changes in Vero cell cultivation methods have been employed in order to improve cell growth conditions to obtain higher viable cell densities and to increase viral titers. The propagation of the 17DD yellow fever virus (YFV) in Vero cells grown on Cytodex I microcarriers was evaluated in 3-L bioreactor vessels. Prior to the current changes, Vero cells were repeatedly displaying insufficient microcarrier colonization. A modified cultivation process with four changes has resulted in higher cell densities and higher virus titers than previously observed for 17DD YFV. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Cellular bone matrices: viable stem cell-containing bone graft substitutes.

    Science.gov (United States)

    Skovrlj, Branko; Guzman, Javier Z; Al Maaieh, Motasem; Cho, Samuel K; Iatridis, James C; Qureshi, Sheeraz A

    2014-11-01

    Advances in the field of stem cell technology have stimulated the development and increased use of allogenic bone grafts containing live mesenchymal stem cells (MSCs), also known as cellular bone matrices (CBMs). It is estimated that CBMs comprise greater than 17% of all bone grafts and bone graft substitutes used. To critically evaluate CBMs, specifically their technical specifications, existing published data supporting their use, US Food and Drug Administration (FDA) regulation, cost, potential pitfalls, and other aspects pertaining to their use. Areview of literature. A series of Ovid, Medline, and Pubmed-National Library of Medicine/National Institutes of Health (www.ncbi.nlm.nih.gov) searches were performed. Only articles in English journals or published with English language translations were included. Level of evidence of the selected articles was assessed. Specific technical information on each CBM was obtained by direct communication from the companies marketing the individual products. Five different CBMs are currently available for use in spinal fusion surgery. There is a wide variation between the products with regard to the average donor age at harvest, total cellular concentration, percentage of MSCs, shelf life, and cell viability after defrosting. Three retrospective studies evaluating CBMs and fusion have shown fusion rates ranging from 90.2% to 92.3%, and multiple industry-sponsored trials are underway. No independent studies evaluating spinal fusion rates with the use of CBMs exist. All the commercially available CBMs claim to meet the FDA criteria under Section 361, 21 CFR Part 1271, and are not undergoing FDA premarket review. The CBMs claim to provide viable MSCs and are offered at a premium cost. Numerous challenges exist in regard to MSCs' survival, function, osteoblastic potential, and cytokine production once implanted into the intended host. Cellular bone matrices may be a promising bone augmentation technology in spinal fusion surgery

  6. Combined bromodeoxyuridine immunohistochemistry and Masson trichrome staining: facilitated detection of cell proliferation in viable vs. infarcted myocardium.

    Science.gov (United States)

    Lazarous, D F; Shou, M; Unger, E F

    1992-09-01

    Cells in the S-phase of the cell cycle can be identified in tissue sections by immunohistochemical localization of the thymidine analogue bromodeoxyuridine (BrdU). Generally, a single counterstain is used to visualize the underlying tissue; however, interpretation of morphologic detail is often difficult. We have utilized BrdU to localize proliferating cells in myocardium exposed to angiogenic mitogens. To facilitate identification of labelled nuclei in the context of infarcted vs. viable myocardium, BrdU immunohistochemistry was followed by a modified Masson trichrome stain. The time of exposure to the counterstains and the wash protocol were re-revised, permitting clear identification of the labelled brown nuclei against a background of red viable myocardium vs. blue infarct. The combined technique also provides color contrast suitable for computer-based image analysis.

  7. Quantitative assessment of viable cells of Lactobacillus plantarum strains in single, dual and multi-strain biofilms.

    Science.gov (United States)

    Fernández Ramírez, Mónica D; Kostopoulos, Ioannis; Smid, Eddy J; Nierop Groot, Masja N; Abee, Tjakko

    2017-03-06

    Biofilms of Lactobacillus plantarum are a potential source for contamination and recontamination of food products. Although biofilms have been mostly studied using single species or even single strains, it is conceivable that in a range of environmental settings including food processing areas, biofilms are composed of multiple species with each species represented by multiple strains. In this study six spoilage related L. plantarum strains FBR1-FBR6 and the model strain L. plantarum WCFS1 were characterised in single, dual and multiple strain competition models. A quantitative PCR approach was used with added propidium monoazide (PMA) enabling quantification of intact cells in the biofilm, representing the viable cell fraction that determines the food spoilage risk. Our results show that the performance of individual strains in multi-strain cultures generally correlates with their performance in pure culture, and relative strain abundance in multi-strain biofilms positively correlated with the relative strain abundance in suspended (planktonic) cultures. Performance of individual strains in dual-strain biofilms was highly influenced by the presence of the secondary strain, and in most cases no correlation between the relative contributions of viable planktonic cells and viable cells in the biofilm was noted. The total biofilm quantified by CV staining of the dual and multi-strain biofilms formed was mainly correlated to CV values of the dominant strain obtained in single strain studies. However, the combination of strain FBR5 and strain WCFS1 showed significantly higher CV values compared to the individual performances of both strains indicating that total biofilm formation was higher in this specific condition. Notably, L. plantarum FBR5 was able to outgrow all other strains and showed the highest relative abundance in dual and multi-strain biofilms. All the dual and multi-strain biofilms contained a considerable number of viable cells, representing a potential

  8. A multimodality imaging model to track viable breast cancer cells from single arrest to metastasis in the mouse brain

    Science.gov (United States)

    Parkins, Katie M.; Hamilton, Amanda M.; Makela, Ashley V.; Chen, Yuanxin; Foster, Paula J.; Ronald, John A.

    2016-10-01

    Cellular MRI involves sensitive visualization of iron-labeled cells in vivo but cannot differentiate between dead and viable cells. Bioluminescence imaging (BLI) measures cellular viability, and thus we explored combining these tools to provide a more holistic view of metastatic cancer cell fate in mice. Human breast carcinoma cells stably expressing Firefly luciferase were loaded with iron particles, injected into the left ventricle, and BLI and MRI were performed on days 0, 8, 21 and 28. The number of brain MR signal voids (i.e., iron-loaded cells) on day 0 significantly correlated with BLI signal. Both BLI and MRI signals decreased from day 0 to day 8, indicating a loss of viable cells rather than a loss of iron label. Total brain MR tumour volume on day 28 also correlated with BLI signal. Overall, BLI complemented our sensitive cellular MRI technologies well, allowing us for the first time to screen animals for successful injections, and, in addition to MR measures of cell arrest and tumor burden, provided longitudinal measures of cancer cell viability in individual animals. We predict this novel multimodality molecular imaging framework will be useful for evaluating the efficacy of emerging anti-cancer drugs at different stages of the metastatic cascade.

  9. Sensitive and Specific Biomimetic Lipid Coated Microfluidics to Isolate Viable Circulating Tumor Cells and Microemboli for Cancer Detection.

    Directory of Open Access Journals (Sweden)

    Jia-Yang Chen

    Full Text Available Here we presented a simple and effective membrane mimetic microfluidic device with antibody conjugated supported lipid bilayer (SLB "smart coating" to capture viable circulating tumor cells (CTCs and circulating tumor microemboli (CTM directly from whole blood of all stage clinical cancer patients. The non-covalently bound SLB was able to promote dynamic clustering of lipid-tethered antibodies to CTC antigens and minimized non-specific blood cells retention through its non-fouling nature. A gentle flow further flushed away loosely-bound blood cells to achieve high purity of CTCs, and a stream of air foam injected disintegrate the SLB assemblies to release intact and viable CTCs from the chip. Human blood spiked cancer cell line test showed the ~95% overall efficiency to recover both CTCs and CTMs. Live/dead assay showed that at least 86% of recovered cells maintain viability. By using 2 mL of peripheral blood, the CTCs and CTMs counts of 63 healthy and colorectal cancer donors were positively correlated with the cancer progression. In summary, a simple and effective strategy utilizing biomimetic principle was developed to retrieve viable CTCs for enumeration, molecular analysis, as well as ex vivo culture over weeks. Due to the high sensitivity and specificity, it is the first time to show the high detection rates and quantity of CTCs in non-metastatic cancer patients. This work offers the values in both early cancer detection and prognosis of CTC and provides an accurate non-invasive strategy for routine clinical investigation on CTCs.

  10. Viable cell sorting of dinoflagellates by multi-parametric flow cytometry.

    Science.gov (United States)

    Electronic cell sorting for isolation and culture of dinoflagellates and other marine eukaryotic phytoplankton was compared to the traditional method of manually picking of cells using a micropipette. Trauma to electronically sorted cells was not a limiting factor as fragile dinoflagellates, such a...

  11. Fuel cells are a commercially viable alternative for the production of "clean" energy.

    Science.gov (United States)

    Niakolas, Dimitris K; Daletou, Maria; Neophytides, Stylianos G; Vayenas, Constantinos G

    2016-01-01

    Fuel cells present a highly efficient and environmentally friendly alternative technology for decentralized energy production. The scope of the present study is to provide an overview of the technological and commercialization readiness level of fuel cells. Specifically, there is a brief description of their general advantages and weaknesses in correlation with various technological actions and political strategies, which are adopted towards their proper positioning in the global market. Some of the most important key performance indicators are also discussed, alongside with a few examples of broad commercialization. It is concluded that the increasing number of companies which utilize and invest on this technology, in combination with the supply chain improvements and the concomitant technological maturity and recognition, reinforce the fuel cell industry so as to become well-aligned for global success.

  12. A viable electrode material for use in microbial fuel cells for tropical regions

    DEFF Research Database (Denmark)

    Offei, Felix; Thygesen, Anders; Mensah, Moses

    2016-01-01

    Electrode materials are critical for microbial fuel cells (MFC) since they influence the construction and operational costs. This study introduces a simple and efficient electrode material in the form of palm kernel shell activated carbon (AC) obtained in tropical regions. The novel introduction...

  13. Progress in emerging techniques for characterization of immobilized viable whole-cell biocatalysts

    Czech Academy of Sciences Publication Activity Database

    Bučko, M.; Vikartovská, A.; Schenkmayerová, A.; Tkáč, J.; Filip, J.; Chorvát Jr., D.; Neděla, Vilém; Ansorge-Schumacher, M.B.; Gemeiner, P.

    2017-01-01

    Roč. 71, č. 11 (2017), s. 2309-2324 ISSN 0366-6352 Institutional support: RVO:68081731 Keywords : bioelectrocatalysis * imaging techniques * immobilized whole- cell biocatalyst * multienzyme cascade reactions * online kinetics Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 1.258, year: 2016

  14. Progress in biocatalysis with immobilized viable whole cells: systems development, reaction engineering and applications

    Czech Academy of Sciences Publication Activity Database

    Polakovič, M.; Švitel, J.; Bučko, M.; Filip, J.; Neděla, Vilém; Ansorge-Schumacher, M.B.; Gemeiner, P.

    2017-01-01

    Roč. 39, č. 5 (2017), s. 667-683 ISSN 0141-5492 Institutional support: RVO:68081731 Keywords : biocatalysis * immobilization methods * immobilized whole-cell biocatalyst * multienzyme cascade reactions * process economics * reaction engineering Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 1.730, year: 2016

  15. Identifying viable regulatory and innovation pathways for regenerative medicine: a case study of cultured red blood cells.

    Science.gov (United States)

    Mittra, J; Tait, J; Mastroeni, M; Turner, M L; Mountford, J C; Bruce, K

    2015-01-25

    The creation of red blood cells for the blood transfusion markets represents a highly innovative application of regenerative medicine with a medium term (5-10 year) prospect for first clinical studies. This article describes a case study analysis of a project to derive red blood cells from human embryonic stem cells, including the systemic challenges arising from (i) the selection of appropriate and viable regulatory protocols and (ii) technological constraints related to stem cell manufacture and scale up to clinical Good Manufacturing Practice (GMP) standard. The method used for case study analysis (Analysis of Life Science Innovation Systems (ALSIS)) is also innovative, demonstrating a new approach to social and natural science collaboration to foresight product development pathways. Issues arising along the development pathway include cell manufacture and scale-up challenges, affected by regulatory demands emerging from the innovation ecosystem (preclinical testing and clinical trials). Our discussion reflects on the efforts being made by regulators to adapt the current pharmaceuticals-based regulatory model to an allogeneic regenerative medicine product and the broader lessons from this case study for successful innovation and translation of regenerative medicine therapies, including the role of methodological and regulatory innovation in future development in the field. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Real-Time PCR Methodology for Selective Detection of Viable Escherichia coli O157:H7 Cells by Targeting Z3276 as a Genetic Marker

    Science.gov (United States)

    Chen, Jin-Qiang

    2012-01-01

    The goal of this study was to develop a sensitive, specific, and accurate method for the selective detection of viable Escherichia coli O157:H7 cells in foods. A unique open reading frame (ORF), Z3276, was identified as a specific genetic marker for the detection of E. coli O157:H7. We developed a real-time PCR assay with primers and probe targeting ORF Z3276 and confirmed that this assay was sensitive and specific for E. coli O157:H7 strains (n = 298). Using this assay, we can detect amounts of genomic DNA of E. coli O157:H7 as low as a few CFU equivalents. Moreover, we have developed a new propidium monoazide (PMA)–real-time PCR protocol that allows for the clear differentiation of viable from dead cells. In addition, the protocol was adapted to a 96-well plate format for easy and consistent handling of a large number of samples. Amplification of DNA from PMA-treated dead cells was almost completely inhibited, in contrast to the virtually unaffected amplification of DNA from PMA-treated viable cells. With beef spiked simultaneously with 8 × 107 dead cells/g and 80 CFU viable cells/g, we were able to selectively detect viable E. coli O157:H7 cells with an 8-h enrichment. In conclusion, this PMA–real-time PCR assay offers a sensitive and specific means to selectively detect viable E. coli O157:H7 cells in spiked beef. It also has the potential for high-throughput selective detection of viable E. coli O157:H7 cells in other food matrices and, thus, will have an impact on the accurate microbiological and epidemiological monitoring of food safety and environmental sources. PMID:22635992

  17. Viable Bacteria Associated with Red Blood Cells and Plasma in Freshly Drawn Blood Donations

    DEFF Research Database (Denmark)

    Damgaard, Christian; Magnussen, Karin; Enevold, Christian

    2015-01-01

    the oral cavity, and to determine the distribution of bacteria revealed in plasma and in the red blood cell (RBC)-fraction. DESIGN: Cross-sectional study. Blood were separated into plasma and RBC-suspensions, which were incubated anaerobically or aerobically for 7 days on trypticase soy blood agar (TSA......), self-reported medically healthy. RESULTS: Bacterial growth was observed on plates inoculated with plasma or RBCs from 62% of the blood donations. Growth was evident in 21 (35%) of 60 RBC-fractions and in 32 (53%) of 60 plasma-fractions versus 8 of 60 negative controls (p = 0.005 and p = 2.6x10...... of RBC-fractions for adherent bacteria should be recommended....

  18. A Viable Electrode Material for Use in Microbial Fuel Cells for Tropical Regions

    Directory of Open Access Journals (Sweden)

    Felix Offei

    2016-01-01

    Full Text Available Electrode materials are critical for microbial fuel cells (MFC since they influence the construction and operational costs. This study introduces a simple and efficient electrode material in the form of palm kernel shell activated carbon (AC obtained in tropical regions. The novel introduction of this material is also targeted at introducing an inexpensive and durable electrode material, which can be produced in rural communities to improve the viability of MFCs. The maximum voltage and power density obtained (under 1000 Ω load using an H-shaped MFC with AC as both anode and cathode electrode material was 0.66 V and 1.74 W/m3, respectively. The power generated by AC was as high as 86% of the value obtained with the extensively used carbon paper. Scanning electron microscopy and Denaturing Gradient Gel Electrophoresis (DGGE analysis of AC anode biofilms confirmed that electrogenic bacteria were present on the electrode surface for substrate oxidation and the formation of nanowires.

  19. Response of Listeria monocytogenes to disinfection stress at the single-cell and population levels as monitored by intracellular pH measurements and viable-cell counts

    DEFF Research Database (Denmark)

    Kastbjerg, Vicky Gaedt; Nielsen, Dennis S.; Arneborg, Nils

    2009-01-01

    .05). The protective effect of NaCl was reflected by viable-cell counts at a higher concentration of Incimaxx (0.0031%), where the salt-grown population survived better than the population grown without NaCl (P ... that a population of L. monocytogenes cells, whether planktonic or attached, is homogenous with respect to sensitivity to an acidic disinfectant studied on the single-cell level. Hence a major subpopulation more tolerant to disinfectants, and hence more persistent, does not appear to be present....

  20. A factor converting viable but nonculturable Vibrio cholerae to a culturable state in eukaryotic cells is a human catalase.

    Science.gov (United States)

    Senoh, Mitsutoshi; Hamabata, Takashi; Takeda, Yoshifumi

    2015-08-01

    In our previous work, we demonstrated that viable but nonculturable (VBNC) Vibrio cholerae O1 and O139 were converted to culturable by coculture with eukaryotic cells. Furthermore, we isolated a factor converting VBNC V. cholerae to culturable (FCVC) from a eukaryotic cell line, HT-29. In this study, we purified FCVC by successive column chromatographies comprising UNO Q-6 anion exchange, Bio-Scale CHT2-1 hydroxyapatite, and Superdex 200 10/300 GL. Homogeneity of the purified FCVC was demonstrated by SDS-PAGE. Nano-LC MS/MS analysis showed that the purified FCVC was a human catalase. An experiment of RNAi knockdown of catalase mRNA from HT-29 cells and treatment of the purified FCVC with a catalase inhibitor, 3-amino-1,2,4-triazole confirmed that the FCVC was a catalase. A possible role of the catalase in converting a VBNC V. cholerae to a culturable state in the human intestine is discussed. © 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  1. Drugs with anti-oxidant properties can interfere with cell viability measurements by assays that rely on the reducing property of viable cells.

    Science.gov (United States)

    Shenoy, Niraj; Stenson, Mary; Lawson, Joshua; Abeykoon, Jithma; Patnaik, Mrinal; Wu, Xiaosheng; Witzig, Thomas

    2017-02-27

    Cell viability assays such as Cell Titer Blue and Alamar Blue rely on the reducing property of viable cells to reduce the reagent dye to a product which gives a fluorescent signal. The current manufacture-recommended protocols do not take into account the possibility of the reagent substrate being reduced directly to the fluorescent product by drugs with an anti-oxidant property. After suspecting spurious results while determining the cytotoxic potential of a drug of interest (DOI) with known anti-oxidant property against a renal cell cancer (RCC) cell line, we aimed to establish that drugs with anti-oxidant property can indeed cause false-negative results with the current protocols of these assays by direct reduction of the reagent substrate. We also aimed to counter the same with a simple modification added to the protocol. Through our experiments, we conclusively demonstrate that drugs with anti-oxidant properties can indeed interfere with cell viability measurements by assays that rely on the reducing property of viable cells. A simple modification in the protocol, as elaborated in the manuscript, can prevent spurious results with these otherwise convenient assays.Laboratory Investigation advance online publication, 27 February 2017; doi:10.1038/labinvest.2017.18.

  2. A Novel Application for Low Frequency Electrochemical Impedance Spectroscopy as an Online Process Monitoring Tool for Viable Cell Concentrations

    Directory of Open Access Journals (Sweden)

    Christoph Slouka

    2016-11-01

    Full Text Available New approaches in process monitoring during industrial fermentations are not only limited to classical pH, dO2 and offgas analysis, but use different in situ and online sensors based on different physical principles to determine biomass, product quality, lysis and far more. One of the very important approaches is the in situ accessibility of viable cell concentration (VCC. This knowledge provides increased efficiency in monitoring and controlling strategies during cultivations. Electrochemical impedance spectroscopy—EIS—is used to monitor biomass in a fermentation of E. coli BL21(DE3, producing a recombinant protein using a fed batch-based approach. Increases in the double layer capacitance (Cdl, determined at frequencies below 1 kHz, are proportional to the increase of biomass in the batch and fed batch phase, monitored in offline and online modes for different cultivations. A good correlation of Cdl with cell density is found and in order to get an appropriate verification of this method, different state-of-the-art biomass measurements are performed and compared. Since measurements in this frequency range are largely determined by the double layer region between the electrode and media, rather minor interferences with process parameters (aeration, stirring are to be expected. It is shown that impedance spectroscopy at low frequencies is a powerful tool for cultivation monitoring.

  3. A viable foal obtained by equine somatic cell nuclear transfer using oocytes recovered from immature follicles of live mares.

    Science.gov (United States)

    Choi, Young-Ho; Norris, Jody D; Velez, Isabel C; Jacobson, Candace C; Hartman, David L; Hinrichs, Katrin

    2013-03-15

    The presence of heterogenous mitochondria from the host ooplast affects the acceptance of offspring obtained by somatic cell nuclear transfer. This might be avoided by obtaining oocytes from selected females, but is then complicated by low numbers of available oocytes. We examined the efficiency of equine somatic cell nuclear transfer using oocytes recovered by transvaginal aspiration of immature follicles from 11 mares. Use of metaphase I oocytes as cytoplasts and of scriptaid (a histone deacetylase inhibitor) treatment during oocyte activation were evaluated to determine if these approaches would increase blastocyst production. In experiment 1, blastocyst development was 0/14 for metaphase I oocytes and 4/103 (4%) for metaphase II oocytes. Three blastocysts were transferred to recipient mares, resulting in two pregnancies and one live foal, which died shortly after birth. In experiment 2, blastocyst development was 2/47 (4%) for control oocytes and 1/83 (1%) for scriptaid-treated oocytes. No foals were born from two blastocysts transferred in the control group. The blastocyst from the scriptaid treatment resulted in birth of a live foal. In conclusion, this is apparently the first report of production of a viable cloned foal from oocytes collected from immature follicles of live mares, supporting the possibility of cloning using oocytes from selected mares. Copyright © 2013 Elsevier Inc. All rights reserved.

  4. Clinical validation of an ultra high-throughput spiral microfluidics for the detection and enrichment of viable circulating tumor cells.

    Directory of Open Access Journals (Sweden)

    Bee Luan Khoo

    Full Text Available Circulating tumor cells (CTCs are cancer cells that can be isolated via liquid biopsy from blood and can be phenotypically and genetically characterized to provide critical information for guiding cancer treatment. Current analysis of CTCs is hindered by the throughput, selectivity and specificity of devices or assays used in CTC detection and isolation.Here, we enriched and characterized putative CTCs from blood samples of patients with both advanced stage metastatic breast and lung cancers using a novel multiplexed spiral microfluidic chip. This system detected putative CTCs under high sensitivity (100%, n = 56 (Breast cancer samples: 12-1275 CTCs/ml; Lung cancer samples: 10-1535 CTCs/ml rapidly from clinically relevant blood volumes (7.5 ml under 5 min. Blood samples were completely separated into plasma, CTCs and PBMCs components and each fraction were characterized with immunophenotyping (Pan-cytokeratin/CD45, CD44/CD24, EpCAM, fluorescence in-situ hybridization (FISH (EML4-ALK or targeted somatic mutation analysis. We used an ultra-sensitive mass spectrometry based system to highlight the presence of an EGFR-activating mutation in both isolated CTCs and plasma cell-free DNA (cf-DNA, and demonstrate concordance with the original tumor-biopsy samples.We have clinically validated our multiplexed microfluidic chip for the ultra high-throughput, low-cost and label-free enrichment of CTCs. Retrieved cells were unlabeled and viable, enabling potential propagation and real-time downstream analysis using next generation sequencing (NGS or proteomic analysis.

  5. Lymph Node Yield in Primary Retroperitoneal Lymph Node Dissection for Nonseminoma Germ Cell Tumors.

    Science.gov (United States)

    Nayan, Madhur; Jewett, Michael A S; Sweet, Joan; Anson-Cartwright, Lynn; Bedard, Philippe L; Moore, Malcolm; Chung, Peter; Warde, Padraig; Hamilton, Robert J

    2015-08-01

    The number of lymph nodes removed at surgery for various malignancies has diagnostic and prognostic value. However, there are limited data on the significance of the number of nodes removed at retroperitoneal lymph node dissection performed for testicular nonseminoma germ cell tumors. From 1979 to 2012 primary open retroperitoneal lymph node dissection was performed by a single experienced surgeon for clinical stage I/II testicular nonseminoma germ cell tumor in 157 patients. Node count was available in 111 cases (71%). Factors associated with total node count and nodes with viable cancer were assessed by linear regression. The association between node count and time to relapse was assessed by multivariate Cox proportional hazards models controlled for adjuvant chemotherapy. The median total lymph node count was 28 (IQR 19-38). Patient age, cancer laterality, body mass index, clinical stage, time from orchiectomy to retroperitoneal lymph node dissection, pathologist and lymph node dissection year were not associated with total lymph node count. A viable germ cell tumor was found in 70 patients (63%). Total node yield was not associated with nodal cancer metastasis. After lymph node dissection 17 patients (16%) received adjuvant chemotherapy. At a median 57-month followup 18 cases (17%) relapsed after primary retroperitoneal lymph node dissection. Increasing total node count was associated with a decreased risk of relapse on univariate and multivariate analysis (HR 0.96, 95% CI 0.92-0.99, p = 0.03 and HR 0.94, 95% CI 0.89-0.99, p = 0.017, respectively). No analyzed clinical or pathological variable was associated with the node yield of primary retroperitoneal lymph node dissection. However, there may be a relationship between the total node yield at retroperitoneal lymph node dissection and the risk of relapse. Copyright © 2015 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  6. Effects of blood transportation on human peripheral mononuclear cell yield, phenotype and function: implications for immune cell biobanking.

    Directory of Open Access Journals (Sweden)

    Anita Posevitz-Fejfár

    Full Text Available Human biospecimen collection, processing and preservation are rapidly emerging subjects providing essential support to clinical as well as basic researchers. Unlike collection of other biospecimens (e.g. DNA and serum, biobanking of viable immune cells, such as peripheral blood mononuclear cells (PBMC and/or isolated immune cell subsets is still in its infancy. While certain aspects of processing and freezing conditions have been studied in the past years, little is known about the effect of blood transportation on immune cell survival, phenotype and specific functions. However, especially for multicentric and cooperative projects it is vital to precisely know those effects. In this study we investigated the effect of blood shipping and pre-processing delay on immune cell phenotype and function both on cellular and subcellular levels. Peripheral blood was collected from healthy volunteers (n = 9: at a distal location (shipped overnight and in the central laboratory (processed immediately. PBMC were processed in the central laboratory and analyzed post-cryopreservation. We analyzed yield, major immune subset distribution, proliferative capacity of T cells, cytokine pattern and T-cell receptor signal transduction. Results show that overnight transportation of blood samples does not globally compromise T- cell subsets as they largely retain their phenotype and proliferative capacity. However, NK and B cell frequencies, the production of certain PBMC-derived cytokines and IL-6 mediated cytokine signaling pathway are altered due to transportation. Various control experiments have been carried out to compare issues related to shipping versus pre-processing delay on site. Our results suggest the implementation of appropriate controls when using multicenter logistics for blood transportation aiming at subsequent isolation of viable immune cells, e.g. in multicenter clinical trials or studies analyzing immune cells/subsets. One important conclusion might

  7. Quantitation of viable Coxiella burnetii in milk using an integrated cell culture-polymerase chain reaction (ICC-PCR) assay.

    Science.gov (United States)

    Stewart, Diana; Shieh, Y-Carol; Tortorello, Mary; Kukreja, Ankush; Shazer, Arlette; Schlesser, Joseph

    2015-11-01

    The obligate intracellular pathogen Coxiella burnetii has long been considered the most heat resistant pathogen in raw milk, making it the reference pathogen for determining pasteurisation conditions for milk products. New milk formulations and novel non-thermal processes require validation of effectiveness which requires a more practical method for analysis than using the currently used animal model for assessing Coxiella survival. Also, there is an interest in better characterising thermal inactivation of Coxiella in various milk formulations. To avoid the use of the guinea pig model for evaluating Coxiella survival, an Integrated Cell Culture-PCR (ICC-PCR) method was developed for determining Coxiella viability in milk. Vero cell cultures were directly infected from Coxiella-contaminated milk in duplicate 24-well plates. Viability of the Coxiella in milk was shown by a ≥ 0.5 log genome equivalent (ge)/ml increase in the quantity of IS111a gene from the baseline post-infection (day 0) level after 9-11 d propagation. Coxiella in skim, 2%, and whole milk, and half and half successfully infected Vero cells and increased in number by at least 2 logs using a 48-h infection period followed by 9-d propagation time. As few as 125 Coxiella ge/ml in whole milk was shown to infect and propagate at least 2 logs in the optimised ICC-PCR assay, though variable confirmation of propagation was shown for as low as 25 Coxiella ge/ml. Applicability of the ICC-PCR method was further proven in an MPN format to quantitate the number of viable Coxiella remaining in whole milk after 60 °C thermal treatment at 0, 20, 40, 60 and 90 min.

  8. Polyelectrolyte Complex Beads by Novel Two-Step Process for Improved Performance of Viable Whole-Cell Baeyer-Villiger Monoxygenase by Immobilization

    Directory of Open Access Journals (Sweden)

    Tomáš Krajčovič

    2017-11-01

    Full Text Available A novel immobilization matrix for the entrapment of viable whole-cell Baeyer–Villiger monooxygenase was developed. Viable recombinant Escherichia coli cells overexpressing cyclohexanone monooxygenase were entrapped in polyelectrolyte complex beads prepared by a two-step reaction of oppositely-charged polymers including highly defined cellulose sulphate. Immobilized cells exhibited higher operational stability than free cells during 10 repeated cycles of Baeyer–Villiger biooxidations of rac-bicyclo[3.2.0]hept-2-en-6-one to the corresponding lactones (1R,5S-3-oxabicyclo-[3.3.0]oct-6-en-3-one and (1S,5R-2-oxabicyclo-[3.3.0]oct-6-en-3-one. The morphology of polyelectrolyte complex beads was characterised by environmental scanning electron microscopy; the spatial distribution of polymers in the beads and cell viability were examined using confocal laser scanning microscopy, and the texture was characterised by the mechanical resistance measurements.

  9. Enhanced inhibition of murine prostatic carcinoma growth by immunization with or administration of viable human umbilical vein endothelial cells and CRM197

    Directory of Open Access Journals (Sweden)

    Zhang Huiyong

    2011-02-01

    Full Text Available Vaccination with xenogeneic and syngeneic endothelial cells is effective for inhibiting tumor growth. Nontoxic diphtheria toxin (CRM197, as an immunogen or as a specific inhibitor of heparin-binding EGF-like growth factor, has shown promising antitumor activity. Therefore, immunization with or administration of viable human umbilical vein endothelial cells (HUVECs combined with CRM197 could have an enhanced antitumor effect. Six-week-old C57BL/6J male mice were vaccinated with viable HUVECs, 1 x 10(6 viable HUVECs combined with 100 μg CRM197, or 100 μg CRM197 alone by ip injections once a week for 4 consecutive weeks. RM-1 cells (5 x 10(5 were inoculated by sc injection as a preventive procedure. During the therapeutic procedure, 6-week-old male C57BL/6J mice were challenged with 1 x 10(5 RM-1 cells, then injected sc with 1 x 10(6 viable HUVECs, 1 x 10(6 viable HUVECs + 100 μg CRM197, and 100 μg CRM197 alone twice a week for 4 consecutive weeks. Tumor volume and life span were monitored. We also investigated the effects of immunization with HUVECs on the aortic arch wall and on wound healing. Vaccination with or administration of viable HUVECs+CRM197 enhanced the inhibition of RM-1 prostatic carcinoma by 24 and 29%, respectively, and prolonged the life span for 3 and 4 days, respectively, compared with those of only vaccination or administration with viable HUVECs of tumor-bearing C57BL/6J mice. Furthermore, HUVEC immunization caused some damage to the aortic arch wall but did not have remarkable effects on the rate of wound healing; the wounds healed in approximately 13 days. Treatment with CRM197 in combination with viable HUVECs resulted in a marked enhancement of the antitumor effect in the preventive or therapeutic treatment for prostatic carcinoma in vivo, suggesting a novel combination for anti-cancer therapy.

  10. Controlling apoptosis to optimize yields of proteins from mammalian cells.

    Science.gov (United States)

    Zustiak, Matthew P; Dorai, Haimanti; Betenbaugh, Michael J; Sauerwald, Tina M

    2012-01-01

    Apoptosis is the foremost method of cell death in bioreactors and can be caused by nutrient limitation, toxin accumulation, and growth factor withdrawal. By delaying the onset of this form of programmed cell death, one can achieve longer sustained viabilities in culture, thereby increasing product yield. Described here is a genetic-based, step-by-step method to generate an apoptosis-resistant cell line. This cell line, then, can be used as a platform for biotherapeutic protein production. The key steps include antiapoptotic transgene selection and transfection followed by clonal isolation and screening. With the proper screening methods, one can obtain a robust cell line that resists the harsh conditions of late-stage and/or high-density culture.

  11. Assessing prognosis and optimizing treatment in patients with postchemotherapy viable nonseminomatous germ-cell tumors (NSGCT): results of the sCR2 international study

    DEFF Research Database (Denmark)

    Fizazi, K.; Oldenburg, J.; Dunant, A.

    2008-01-01

    malignant cells, and a good International Germ Cell Consensus Classification group at presentation. Patients were assigned to one of three risk groups defined in sCR1: no risk factor (good risk), one risk factor (intermediate risk) and two to three risk factors (poor risk group). The 5-year PFS rate was 92...... with surveillance and treatment only at relapse. CONCLUSION: In patients with postchemotherapy viable NSGCT, a complete resection of residual masses should be rigorously pursued. These data validate the sCR1 prognostic index. Given their excellent outcome, patients in the favorable group may not require......BACKGROUND: The purpose of this study was to validate a prognostic index [surgical complete response 1 (sCR1)] in patients with postchemotherapy viable nonseminomatous germ-cell tumors (NSGCT). PATIENTS AND METHODS: Data and specimens from 61 patients with normalized tumor markers...

  12. Viable Cancer Cells in the Remnant Stomach are a Potential Source of Peritoneal Metastasis after Curative Distal Gastrectomy for Gastric Cancer.

    Science.gov (United States)

    Murata, Satoshi; Yamamoto, Hiroshi; Yamaguchi, Tsuyoshi; Kaida, Sachiko; Ishida, Mitsuaki; Kodama, Hirokazu; Takebayashi, Katsushi; Shimizu, Tomoharu; Miyake, Toru; Tani, Tohru; Kushima, Ryoji; Tani, Masaji

    2016-09-01

    The mechanisms underlying peritoneal metastasis (PM) after curative gastrectomy for gastric cancer (GC) are not well elucidated. This study assessed whether viable cancer cells, including cancer stemlike cells (CSCs), were present in the remnant stomach immediately before gastrointestinal (GI) tract reconstruction because these could be a source of PM after gastrectomy. Saline fluid used for remnant stomach lumen irrigation before GI reconstruction was prospectively collected from 142 consecutive patients undergoing distal gastrectomy for GC and cytologically examined. Proliferative activity (Ki67 staining) and stemness (expression of the CSC surface markers CD44s or CD44v6) were evaluated in detected cancer cells. Viable cancer cells were detected in 33 (23.2 %) of the 142 remnant stomachs. These cells formed clusters and stained positively for Ki67, indicating proliferation. Cancer cells in remnant stomachs and surface cancer cells in primary GCs from 10 (30.3 %) of these 33 cases also stained positively for CD44s or CD44v6. In a multiple logistic regression analysis, advanced cancer (odds ratio [OR], 4.65; 95 % confidence interval [CI], 1.32-16.4; P = 0.017), tumor size of 40 mm or larger (OR, 3.78; 95 % CI, 1.12-12.8; P = 0.033), and histologic differentiation (OR, 3.10; 95 % CI, 1.30-7.40; P = 0.011) were associated independently with the presence of cancer cells in the remnant stomach. Viable, proliferative, and clustered cancer cells, including CSCs, were found in remnant gastric lumens immediately before GI reconstruction, indicating a possible cellular source of PM after curative gastrectomy for GC. Dissemination of gastric contents into the peritoneal cavity should be avoided during GI reconstruction.

  13. Designing primers and evaluation of the efficiency of propidium monoazide – Quantitative polymerase chain reaction for counting the viable cells of Lactobacillus gasseri and Lactobacillus salivarius

    Directory of Open Access Journals (Sweden)

    Chieh-Hsien Lai

    2017-07-01

    Full Text Available The purpose of this study is to evaluate the efficiency of using propidium monoazide (PMA real-time quantitative polymerase chain reaction (qPCR to count the viable cells of Lactobacillus gasseri and Lactobacillus salivarius in probiotic products. Based on the internal transcription spacer and 23S rRNA genes, two primer sets specific for these two Lactobacillus species were designed. For a probiotic product, the total deMan Rogosa Sharpe plate count was 8.65±0.69 log CFU/g, while for qPCR, the cell counts of L. gasseri and L. salivarius were 8.39±0.14 log CFU/g and 8.57±0.24 log CFU/g, respectively. Under the same conditions, for its heat-killed product, qPCR counts for L. gasseri and L. salivarius were 6.70±0.16 log cells/g and 7.67±0.20 log cells/g, while PMA-qPCR counts were 5.33±0.18 log cells/g and 5.05±0.23 log cells/g, respectively. For cell dilutions with a viable cell count of 8.5 log CFU/mL for L. gasseri and L. salivarius, after heat killing, the PMA-qPCR count for both Lactobacillus species was near 5.5 log cells/mL. When the PMA-qPCR counts of these cell dilutions were compared before and after heat killing, although some DNA might be lost during the heat killing, significant qPCR signals from dead cells, i.e., about 4–5 log cells/mL, could not be reduced by PMA treatment. Increasing PMA concentrations from 100 μM to 200 μM or light exposure time from 5 minutes to 15 minutes had no or, if any, only minor effect on the reduction of qPCR signals from their dead cells. Thus, to differentiate viable lactic acid bacterial cells from dead cells using the PMA-qPCR method, the efficiency of PMA to reduce the qPCR signals from dead cells should be notable.

  14. Designing primers and evaluation of the efficiency of propidium monoazide - Quantitative polymerase chain reaction for counting the viable cells of Lactobacillus gasseri and Lactobacillus salivarius.

    Science.gov (United States)

    Lai, Chieh-Hsien; Wu, Sih-Rong; Pang, Jen-Chieh; Ramireddy, Latha; Chiang, Yu-Cheng; Lin, Chien-Ku; Tsen, Hau-Yang

    2017-07-01

    The purpose of this study is to evaluate the efficiency of using propidium monoazide (PMA) real-time quantitative polymerase chain reaction (qPCR) to count the viable cells of Lactobacillus gasseri and Lactobacillus salivarius in probiotic products. Based on the internal transcription spacer and 23S rRNA genes, two primer sets specific for these two Lactobacillus species were designed. For a probiotic product, the total deMan Rogosa Sharpe plate count was 8.65±0.69 log CFU/g, while for qPCR, the cell counts of L. gasseri and L. salivarius were 8.39±0.14 log CFU/g and 8.57±0.24 log CFU/g, respectively. Under the same conditions, for its heat-killed product, qPCR counts for L. gasseri and L. salivarius were 6.70±0.16 log cells/g and 7.67±0.20 log cells/g, while PMA-qPCR counts were 5.33±0.18 log cells/g and 5.05±0.23 log cells/g, respectively. For cell dilutions with a viable cell count of 8.5 log CFU/mL for L. gasseri and L. salivarius, after heat killing, the PMA-qPCR count for both Lactobacillus species was near 5.5 log cells/mL. When the PMA-qPCR counts of these cell dilutions were compared before and after heat killing, although some DNA might be lost during the heat killing, significant qPCR signals from dead cells, i.e., about 4-5 log cells/mL, could not be reduced by PMA treatment. Increasing PMA concentrations from 100 μM to 200 μM or light exposure time from 5 minutes to 15 minutes had no or, if any, only minor effect on the reduction of qPCR signals from their dead cells. Thus, to differentiate viable lactic acid bacterial cells from dead cells using the PMA-qPCR method, the efficiency of PMA to reduce the qPCR signals from dead cells should be notable. Copyright © 2016. Published by Elsevier B.V.

  15. Not single but periodic injections of synovial mesenchymal stem cells maintain viable cells in knees and inhibit osteoarthritis progression in rats.

    Science.gov (United States)

    Ozeki, N; Muneta, T; Koga, H; Nakagawa, Y; Mizuno, M; Tsuji, K; Mabuchi, Y; Akazawa, C; Kobayashi, E; Matsumoto, K; Futamura, K; Saito, T; Sekiya, I

    2016-06-01

    We investigated the effects of single or repetitive intra-articular injections of synovial mesenchymal stem cells (MSCs) on a rat osteoarthritis (OA) model, and elucidated the behaviors and underlying mechanisms of the stem cells after the injection. One week after the transection of the anterior cruciate ligament (ACL) of wild type Lewis rats, one million synovial MSCs were injected into the knee joint every week. Cartilage degeneration was evaluated with safranin-o staining after the first injection. To analyze cell kinetics or MSC properties, luciferase, LacZ, and GFP expressing synovial MSCs were used. To confirm the role of MSCs, species-specific microarray and PCR analyses were performed using human synovial MSCs. Histological analysis for femoral and tibial cartilage showed that a single injection was ineffective but weekly injections had significant chondroprotective effects for 12 weeks. Histological and flow-cytometric analyses of LacZ and GFP expressing synovial MSCs revealed that injected MSCs migrated mainly into the synovium and most of them retained their undifferentiated MSC properties though the migrated cells rapidly decreased. In vivo imaging analysis revealed that MSCs maintained in knees while weekly injection. Species-specific microarray and PCR analyses showed that the human mRNAs on day 1 for 21 genes increased over 50-fold, and increased the expressions of PRG-4, BMP-2, and BMP-6 genes encoding chondroprotective proteins, and TSG-6 encoding an anti-inflammatory one. Not single but periodic injections of synovial MSCs maintained viable cells without losing their MSC properties in knees and inhibited osteoarthritis (OA) progression by secretion of trophic factors. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  16. Macrophage conditioned medium induced cellular network formation in MCF-7 cells through enhanced tunneling nanotube formation and tunneling nanotube mediated release of viable cytoplasmic fragments

    Energy Technology Data Exchange (ETDEWEB)

    Patheja, Pooja, E-mail: pooja.patheja8@gmail.com [Laser Biomedical Applications Section, Raja Ramanna Centre for Advanced Technology, Indore 452013, Madhya Pradesh (India); Homi Bhabha National Institute, Training School Complex, Anushaktinagar, Mumbai 400094, Maharashtra (India); Sahu, Khageswar [Laser Biomedical Applications Section, Raja Ramanna Centre for Advanced Technology, Indore 452013, Madhya Pradesh (India)

    2017-06-15

    Infiltrating macrophages in tumor microenvironment, through their secreted cytokines and growth factors, regulate several processes of cancer progression such as cancer cell survival, proliferation, invasion, metastasis and angiogenesis. Recently, intercellular cytoplasmic bridges between cancer cells referred as tunneling nanotubes (TNTs) have been recognized as novel mode of intercellular communication between cancer cells. In this study, we investigated the effect of inflammatory mediators present in conditioned medium derived from macrophages on the formation of TNTs in breast adenocarcinoma cells MCF-7. Results show that treatment with macrophage conditioned medium (MφCM) not only enhanced TNT formation between cells but also stimulated the release of independently migrating viable cytoplasmic fragments, referred to as microplasts, from MCF-7 cells. Time lapse microscopy revealed that microplasts were released from parent cancer cells in extracellular space through formation of TNT-like structures. Mitochondria, vesicles and cytoplasm could be transferred from parent cell body to microplasts through connecting TNTs. The microplasts could also be resorbed into the parent cell body by retraction of the connecting TNTs. Microplast formation inhibited in presence cell migration inhibitor, cytochalasin-B. Notably by utilizing migratory machinery within microplasts, distantly located MCF-7 cells formed several TNT based intercellular connections, leading to formation of physically connected network of cells. Together, these results demonstrate novel role of TNTs in microplast formation, novel modes of TNT formation mediated by microplasts and stimulatory effect of MφCM on cellular network formation in MCF-7 cells mediated through enhanced TNT and microplast formation.

  17. Bacillus subtilis mutants with knockouts of the genes encoding ribonucleases RNase Y and RNase J1 are viable, with major defects in cell morphology, sporulation, and competence.

    Science.gov (United States)

    Figaro, Sabine; Durand, Sylvain; Gilet, Laetitia; Cayet, Nadège; Sachse, Martin; Condon, Ciarán

    2013-05-01

    The genes encoding the ribonucleases RNase J1 and RNase Y have long been considered essential for Bacillus subtilis cell viability, even before there was concrete knowledge of their function as two of the most important enzymes for RNA turnover in this organism. Here we show that this characterization is incorrect and that ΔrnjA and Δrny mutants are both viable. As expected, both strains grow relatively slowly, with doubling times in the hour range in rich medium. Knockout mutants have major defects in their sporulation and competence development programs. Both mutants are hypersensitive to a wide range of antibiotics and have dramatic alterations to their cell morphologies, suggestive of cell envelope defects. Indeed, RNase Y mutants are significantly smaller in diameter than wild-type strains and have a very disordered peptidoglycan layer. Strains lacking RNase J1 form long filaments in tight spirals, reminiscent of mutants of the actin-like proteins (Mre) involved in cell shape determination. Finally, we combined the rnjA and rny mutations with mutations in other components of the degradation machinery and show that many of these strains are also viable. The implications for the two known RNA degradation pathways of B. subtilis are discussed.

  18. Effects of the oral administration of viable and heat-killed Streptococcus bovis HC5 cells to pre-sensitized BALB/c mice.

    Directory of Open Access Journals (Sweden)

    Aline D Paiva

    Full Text Available Antimicrobial peptides have been suggested as an alternative to classical antibiotics in livestock production and bacteriocin-producing bacteria could be added to animal feeds to deliver bacteriocins in the gastrointestinal (GI tract of ruminant and monogastric animals. In this study, viable (V and heat-killed (HK Streptococcus bovis HC5 cells were orally administered to pre-sensitized mice in order to assess the effects of a bacteriocin-producing bacteria on histological parameters and the immune response of the GI tract of monogastric animals. The administration of V and HK S. bovis HC5 cells during 58 days to BALB/c mice did not affect weight gain, but an increase in gut permeability was detected in animals receiving the HK cells. Viable and heat killed cells caused similar morphological alterations in the GI tract of the animals, but the most prominent effects were detected in the small intestine. The oral administration of S. bovis HC5 also influenced cytokine production in the small intestine, and the immune-mediated activity differed between V and HK cells. The relative expression of IL-12 and INF-γ was significantly higher in the small intestine of mice treated with V cells, while an increase in IL-5, IL-13 and TNF-α expression was only detected in mice treated with HK cells. Considering that even under a condition of severe challenge (pre-sensitization followed by daily exposure to the same bacterial immunogen the general health of the animals was maintained, it appears that oral administration of S. bovis HC5 cells could be a useful route to deliver bacteriocin in the GI tract of livestock animals.

  19. A non-aggressive, highly efficient, enzymatic method for dissociation of human brain-tumors and brain-tissues to viable single-cells.

    Science.gov (United States)

    Volovitz, Ilan; Shapira, Netanel; Ezer, Haim; Gafni, Aviv; Lustgarten, Merav; Alter, Tal; Ben-Horin, Idan; Barzilai, Ori; Shahar, Tal; Kanner, Andrew; Fried, Itzhak; Veshchev, Igor; Grossman, Rachel; Ram, Zvi

    2016-06-01

    Conducting research on the molecular biology, immunology, and physiology of brain tumors (BTs) and primary brain tissues requires the use of viably dissociated single cells. Inadequate methods for tissue dissociation generate considerable loss in the quantity of single cells produced and in the produced cells' viability. Improper dissociation may also demote the quality of data attained in functional and molecular assays due to the presence of large quantities cellular debris containing immune-activatory danger associated molecular patterns, and due to the increased quantities of degraded proteins and RNA. Over 40 resected BTs and non-tumorous brain tissue samples were dissociated into single cells by mechanical dissociation or by mechanical and enzymatic dissociation. The quality of dissociation was compared for all frequently used dissociation enzymes (collagenase, DNase, hyaluronidase, papain, dispase) and for neutral protease (NP) from Clostridium histolyticum. Single-cell-dissociated cell mixtures were evaluated for cellular viability and for the cell-mixture dissociation quality. Dissociation quality was graded by the quantity of subcellular debris, non-dissociated cell clumps, and DNA released from dead cells. Of all enzymes or enzyme combinations examined, NP (an enzyme previously not evaluated on brain tissues) produced dissociated cell mixtures with the highest mean cellular viability: 93 % in gliomas, 85 % in brain metastases, and 89 % in non-tumorous brain tissue. NP also produced cell mixtures with significantly less cellular debris than other enzymes tested. Dissociation using NP was non-aggressive over time-no changes in cell viability or dissociation quality were found when comparing 2-h dissociation at 37 °C to overnight dissociation at ambient temperature. The use of NP allows for the most effective dissociation of viable single cells from human BTs or brain tissue. Its non-aggressive dissociative capacity may enable ambient

  20. Prognostic impact of the number of viable circulating cells with high telomerase activity in gastric cancer patients: a prospective study.

    Science.gov (United States)

    Ito, Hiroaki; Inoue, Haruhiro; Kimura, Satoshi; Ohmori, Tohru; Ishikawa, Fumihiro; Gohda, Keigo; Sato, Jun

    2014-07-01

    The identification of circulating tumor cells (CTCs) in peripheral blood is a useful approach to estimate prognosis, monitor disease progression and measure treatment effects in several types of malignancies. We have previously used OBP-401, a telomerase-specific, replication-selective, oncolytic adenoviral agent carrying the green fluorescent protein (GFP) gene. GFP-positive cells (GFP+ cells) were counted under a fluorescence microscope. Our results showed that the number of at least 7.735 µm in diameter GFP+ cells (L-GFP+ cells) in the peripheral blood was a significant marker of prognosis in gastric cancer patients. However, tumor cells undergoing epithelial-mesenchymal transition (EMT) have been reported to be smaller in size than cells without EMT features; thus, CTCs undergoing EMT may escape detection with this technique. Therefore, in this study, we analyzed the relationship between patient outcome and the number of GFP+ cells of any size. We obtained peripheral blood samples from 65 patients with gastric cancer. After infection of OBP-401, GFP+ cells were counted and measured. The relationship between the number of GFP+ cells and surgical outcome was analyzed. The median follow-up period of the surviving patients was 36 months. A significant difference in overall survival was found between patients with 0-5 and patients with ≥6 L-GFP+ cells. No clear relationship was established between the number of small-sized GFP+ cells and patient prognosis. The number of L-GFP+ cells was significantly related to overall survival in patients with gastric cancer. The detection of L-GFP+ cells using OBP-401 may be a useful prognostic marker in gastric cancer.

  1. Binding of CLL subset 4 B-cell receptor immunoglobulins to viable human memory B lymphocytes requires a distinctive IGKV somatic mutation.

    Science.gov (United States)

    Catera, Rosa; Liu, Yun; Gao, Chao; Yan, Xiao-Jie; Magli, Amanda; Allen, Steven L; Kolitz, Jonathan E; Rai, Kanti R; Chu, Charles C; Feizi, Ten; Stamatopoulos, Kostas; Chiorazzi, Nicholas

    2017-01-12

    Amino acid replacement mutations in certain CLL stereotyped B-cell receptor (BCR) immunoglobulins (IGs) at defined positions within antigen-binding sites strongly imply antigen selection. Prime examples of this are CLL subset 4 BCR IGs using IGHV4-34/IGHD5-18/IGHJ6 and IGKV2-30/IGKJ2 rearrangements. Conspicuously and unlike most CLL IGs, subset 4 IGs do not bind apoptotic cells. By testing the (auto)antigenic reactivities of subset 4 IGs toward viable lymphoid-lineage cells and specific autoantigens typically bound by IGHV4-34+ IGs, we found IGs from both subset 4 and non-subset 4 IGHV4-34-expressing CLL cases bind naïve B cells. However, only subset 4 IGs react with memory B cells. Furthermore, subset 4 IGs do not bind DNA nor i or I carbohydrate antigens, common targets of IGHV4-34-utilizing antibodies in systemic lupus erythematosus and cold agglutinin disease, respectively. Notably, we found that subset 4 IG binding to memory B lymphocytes depends on an aspartic acid at position 66 of FR3 in the rearranged IGKV2-30 gene; this amino acid residue is acquired by somatic mutation. Our findings illustrate the importance of positive and negative selection criteria for structural elements in CLL IGs and suggest that autoantigens driving normal B cells to become subset 4 CLL cells differ from those driving IGHV4-34+ B cells in other diseases.

  2. Fluorescence Quenching Property of C-Phycocyanin from Spirulina platensis and its Binding Efficacy with Viable Cell Components.

    Science.gov (United States)

    Paswan, Meenakshi B; Chudasama, Meghna M; Mitra, Madhusree; Bhayani, Khushbu; George, Basil; Chatterjee, Shruti; Mishra, Sandhya

    2016-03-01

    Phycocyanin is a natural brilliant blue colored, fluorescent protein, which is commonly present in cyanobacteria. In this study, C-phycocyanin was extracted and purified from Spirulina platensis, which are multicellular and filamentous cyanobacteria of greater importance because of its various biological and pharmacological potential. It was analyzed for its binding affinity towards blood cells, algal cells, genomic DNA of microalgae, and bacteria at different temperature and incubation time. It showed good binding affinity with these components even at low concentration of 2.5 μM. The purpose of this study was to evaluate the applicability of C-phycocyanin as a green fluorescent dye substituting carcinogenic chemical dyes.

  3. Managing Viable Knowledge

    NARCIS (Netherlands)

    Achterbergh, J.M.I.M.; Vriens, D.J.

    2002-01-01

    In this paper, Beer's Viable System Model (VSM) is applied to knowledge management. Based on the VSM, domains of knowledge are identified that an organization should possess to maintain its viability. The logic of the VSM is also used to support the diagnosis, design and implementation of the

  4. Determination of Viable Salmonella Typhimurium Cells in Heat Treated Milk By PMA/Real-Time PCR Method

    Directory of Open Access Journals (Sweden)

    Zülal Kesmen

    2017-06-01

    Full Text Available Applying different technological processes during the production of food has a lethal effect on the bacteria but DNA of these bacterial strains may cause false positive results when detected by real time PCR technique because they preserve their existence for a certain period of time. To overcome this shortcoming of the real time PCR technique, a new method has been developed in recent years, based on the removal of dead cell DNA from the medium by treatment with Propodium Monoazide (PMA before DNA extraction. In this study, real-time PCR method was combined with PMA application for the detection of live cells of Salmonella Typhimurium in heat treated milk samples. For this purpose, milk samples inoculated with S. Tyhimurium were heat treated at different temperatures (60, 65, 70 and 75°C and times (15, 60, 300, 900 sec and number of live bacteria was determined comparatively by direct real-time PCR, PMA/real-time PCR and conventional cultural method. As a result, unlike the direct real time PCR technique, PMA/real-time PCR method prevents to a certain extent of false positive results from dead cells at all tested temperatures and times but higher results were obtained from PMA/real-time PCR method when compared to conventional cultural results. Therefore, further studies should be carried out to optimize the conditions of the PMA application in order to eliminate the high positive results detected by the PMA / real-time PCR method

  5. Advances toward regenerative medicine in the central nervous system: challenges in making stem cell therapy a viable clinical strategy.

    Science.gov (United States)

    Stoll, Elizabeth A

    2014-01-01

    Over recent years, there has been a great deal of interest in the prospects of stem cell-based therapies for the treatment of nervous system disorders. The eagerness of scientists, clinicians, and spin-out companies to develop new therapies led to premature clinical trials in human patients, and now the initial excitement has largely turned to skepticism. Rather than embracing a defeatist attitude or pressing blindly ahead, I argue it is time to evaluate the challenges encountered by regenerative medicine in the central nervous system and the progress that is being made to solve these problems. In the twenty years since the adult brain was discovered to have an endogenous regenerative capacity, much basic research has been done to elucidate mechanisms controlling proliferation and cellular identity; how stem cells may be directed into neuronal lineages; genetic, pharmacological, and behavioral interventions that modulate neurogenic activity; and the exact nature of limitations to regeneration in the adult, aged, diseased and injured CNS. These findings should prove valuable in designing realistic clinical strategies to improve the prospects of stem cell-based therapies. In this review, I discuss how basic research continues to play a critical role in identifying both barriers and potential routes to regenerative therapy in the CNS.

  6. Rotary orbital suspension culture of embryonic stem cell-derived neural stem/progenitor cells: impact of hydrodynamic culture on aggregate yield, morphology and cell phenotype.

    Science.gov (United States)

    Laundos, Tiago L; Silva, Joana; Assunção, Marisa; Quelhas, Pedro; Monteiro, Cátia; Oliveira, Carla; Oliveira, Maria J; Pêgo, Ana P; Amaral, Isabel F

    2017-08-01

    Embryonic stem (ES)-derived neural stem/progenitor cells (ES-NSPCs) constitute a promising cell source for application in cell therapies for the treatment of central nervous system disorders. In this study, a rotary orbital hydrodynamic culture system was applied to single-cell suspensions of ES-NSPCs, to obtain homogeneously-sized ES-NSPC cellular aggregates (neurospheres). Hydrodynamic culture allowed the formation of ES-NSPC neurospheres with a narrower size distribution than statically cultured neurospheres, increasing orbital speeds leading to smaller-sized neurospheres and higher neurosphere yield. Neurospheres formed under hydrodynamic conditions (72 h at 55 rpm) showed higher cell compaction and comparable percentages of viable, dead, apoptotic and proliferative cells. Further characterization of cellular aggregates provided new insights into the effect of hydrodynamic shear on ES-NSPC behaviour. Rotary neurospheres exhibited reduced protein levels of N-cadherin and β-catenin, and higher deposition of laminin (without impacting fibronectin deposition), matrix metalloproteinase-2 (MMP-2) activity and percentage of neuronal cells. In line with the increased MMP-2 activity levels found, hydrodynamically-cultured neurospheres showed higher outward migration on laminin. Moreover, when cultured in a 3D fibrin hydrogel, rotary neurospheres generated an increased percentage of neuronal cells. In conclusion, the application of a constant orbital speed to single-cell suspensions of ES-NSPCs, besides allowing the formation of homogeneously-sized neurospheres, promoted ES-NSPC differentiation and outward migration, possibly by influencing the expression of cell-cell adhesion molecules and the secretion of proteases/extracellular matrix proteins. These findings are important when establishing the culture conditions needed to obtain uniformly-sized ES-NSPC aggregates, either for use in regenerative therapies or in in vitro platforms for biomaterial development or

  7. Induction of Viable but Nonculturable Salmonella in Exponentially Grown Cells by Exposure to a Low-Humidity Environment and Their Resuscitation by Catalase.

    Science.gov (United States)

    Morishige, Yuta; Koike, Atsushi; Tamura-Ueyama, Ai; Amano, Fumio

    2017-02-01

    Salmonella is a major cause of foodborne disease that sometimes occurs in massive outbreaks around the world. This pathogen is tolerant of low-humidity conditions. We previously described a method for induction of viable but nonculturable (VBNC) Salmonella enterica serovar Enteritidis by treatment with hydrogen peroxide (H2O2) and subsequent resuscitation with 0.3 mM sodium pyruvate. Here, we report a new method for the induction of the VBNC state in Salmonella Enteritidis cells, one involving dehydration. Exposure of Salmonella Enteritidis cells to dehydration stress under poor nutritional conditions (0.9% [wt/vol] NaCl) and 10 to 20% relative humidity at room temperature decreased the presence of culturable population to 0.0067%, but respiratory and glucose uptake active populations were maintained at 0.46 and 1.12%, respectively, meaning that approximately 1% may have entered the VBNC state. Furthermore, these VBNC cells could be resuscitated to acquire culturability by incubation with catalase in M9 minimal medium without glucose in a manner dependent on the dose of catalase but not sodium pyruvate. These results suggest that a low-humidity environment could cause Salmonella Enteritidis cells to enter the VBNC state and the cells could then be resuscitated for growth by treatment with catalase, suggesting a potential risk of Salmonella Enteritidis to survive in low water activity foods in the VBNC state and to start regrowth for foodborne illness.

  8. Freezing Nitrogen Ethanol Composite May be a Viable Approach for Cryotherapy of Human Giant Cell Tumor of Bone.

    Science.gov (United States)

    Wu, Po-Kuei; Chen, Cheng-Fong; Wang, Jir-You; Chen, Paul Chih-Hsueh; Chang, Ming-Chau; Hung, Shih-Chieh; Chen, Wei-Ming

    2017-06-01

    Liquid nitrogen has been used as adjuvant cryotherapy for treating giant cell tumor (GCT) of bone. However, the liquid phase and ultrafreezing (-196° C) properties increase the risk of damage to the adjacent tissues and may lead to perioperative complications. A novel semisolid cryogen, freezing nitrogen ethanol composite, might mitigate these shortcomings because of less-extreme freezing. We therefore wished to evaluate freezing nitrogen ethanol composite as a coolant to determine its properties in tumor cryoablation. (1) Is freezing nitrogen ethanol composite-mediated freezing effective for tumor cryoablation in an ex vivo model, and if yes, is apoptosis involved in the tumor-killing mechanism? (2) Does freezing nitrogen ethanol composite treatment block neovascularization and neoplastic progression of the grafted GCTs and is it comparable to that of liquid nitrogen in an in vivo chicken model? (3) Can use of freezing nitrogen ethanol composite as an adjuvant to curettage result in successful short-term treatment, defined as absence of GCT recurrence at a minimum of 1 year in a small proof-of-concept clinical series? The cryogenic effect on bone tissue mediated by freezing nitrogen ethanol composite and liquid nitrogen was verified by thermal measurement in a time-course manner. Cryoablation on human GCT tissue was examined ex vivo for effect on morphologic features (cell shrinkage) and DNA fragmentation (apoptosis). The presumed mechanism was investigated by molecular analysis of apoptosis regulatory proteins including caspases 3, 8, and 9 and Bax/Bcl-2. Chicken chorioallantoic membrane was used as an in vivo model to evaluate the effects of freezing nitrogen ethanol composite and liquid nitrogen treatment on GCT-derived neovascularization and tumor neoplasm. A small group of patients with GCT of bone was treated by curettage and adjuvant freezing nitrogen ethanol composite cryotherapy in a proof-of-concept study. Tumor recurrence and perioperative

  9. Quality of raw cow milk in Republic of Macedonia determined through the testing of somatic cell count and total viable count

    Directory of Open Access Journals (Sweden)

    Angelovski Ljupco

    2008-11-01

    Full Text Available Somatic cells count and total viable count are criteria used to estimate the compliance of raw cow milk with the Book of rules for demands for safety and hygiene and procedures for official controls of milk and milk products, Official Gazette of RM 157/2007. According to the given demands, raw milk operators are obliged to conduct all procedures and to guarantee that milk is in compliance with the criteria laid down in Book of rules. At the same time, Republic of Macedonia have to fulfill EU criteria laid down in Directive 92/46 (Council directive 92/46/EEC laying down the health rules for the production and placing on the market of raw milk, heat-treated milk and milkbased products for quality of raw milk as part of implementation of community legislation and milk production. The independent laboratory for milk quality control at FVM-Skopje, in frame of its activities in the period February- August 2008 has conducted a study for obtaining preliminary results for the situation with raw milk quality produced in R. of Macedonia for somatic cells counts and total viable count. In the study we analyzed 2065 samples for TVC and 1625 samples for SCC of raw milk samples produced in different parts of the country. From the tested samples only 41,8% fulfill criteria for SCC and 41,45% criteria for TVC lay down in Book of rules for 2008. Assessment of the results in light of Council Directive it is obvious that only 42,7% of the samples for SCC and 10,7% for TVC fulfill the criteria of Council Directive having in mind different requirements vs. Book of rules.

  10. Biologic properties of viable deletion mutants of simian virus 40 (SV40) rescued from the cells of an SV40-induced hamster lymphocytic leukemia.

    Science.gov (United States)

    Diamandopoulos, G T; Carmichael, G

    1983-12-01

    A lymphocytic leukemia induced by the oncogenic DNA simian virus 40 (SV40) in an inbred LSH/SsLak Syrian golden hamster was evoked to produce infectious SV40 by fusion of the leukemia cells with grivet monkey kidney (GMK) cells and by exposure of the leukemia cells to the chemical inducers mitomycin C and cycloheximide. Plaque-purified viable substrains of the rescued SV40 when studied by restriction endonuclease digestion of viral DNA were found to contain small deletions within the Hind III restriction fragment C. These deletions lay near the viral origin of DNA replication. Ten plaque-purified substrains of the rescued virus identified by immunofluorescence as being SV40 were found, when compared to the wild-type SV40, to replicate slowly and to form small plaques. Although these substrains transformed NIH/3T3 cells as efficiently as the wild-type SV40 in tissue culture, they were generally less oncogenic in vivo--7 of the 10 failed to induce tumors. The 3 oncogenic SV40-rescued substrains were not found to exhibit "lymphocytotropism," i.e., the capacity to infect and neoplastically transform preferentially hamster lymphocytes. Thus the hamster lymphocytic leukemia originally induced by the wild-type SV40 was most likely a chance-stochastic event rather than the result of tropism-determinism mediated by the virus, as is usually the case with leukemogenic RNA viruses.

  11. [Factors impacting yield of CD34(+) cells from healthy donors mobilized with rhG-CSF].

    Science.gov (United States)

    Zhu, Ling; Zhou, Li-Kun; Xue, Mei; Yan, Hong-Min; Liu, Jing; Wang, Zhi-Dong; Ding, Li; Wang, Heng-Xiang

    2009-12-01

    This study was aimed to explore the effect of donor characteristics (age, sex and so on.) on CD34(+) cell yields in apheresis from healthy donors mobilized by recombinant granulocyte colony-stimulating factor(rhG-CSF). In 61 healthy donors, the characteristics associated with CD34(+) cell yield were analysed. The relationship between the CD34(+) cell yields and donor characteristics was statistically assessed with multivariate forward, backward and stepwise regression methods. A variety of parameters were analyzed which included donor age, sex, weight, height, body mass index (BMI) and time for collection of peripheral blood apheresis, while the mean number of peripheral blood mononuclear cells (MNCs), CD34(+) cell count, CD34(+) cell proportion based on MNC and CD34(+) cell count per kg of donor weight were used as the variables. The results showed that age of donors was the main factor impacting CD34(+) cell yields (-0.60 donors could reach the peak CD34(+) cell yields. It is concluded that the age of the donors is the first factor determining the choice of donors for allogeneic hematopoietic stem cell transplantation, the sex, height, weight and BMI are secondary factors impacting yield of CD34(+) cells from donors mobilized with rhG-CSF.

  12. Primordial germ cell-mediated chimera technology produces viable pure-line Houbara bustard offspring: potential for repopulating an endangered species.

    Directory of Open Access Journals (Sweden)

    Ulrich Wernery

    2010-12-01

    Full Text Available The Houbara bustard (Chlamydotis undulata is a wild seasonal breeding bird populating arid sandy semi-desert habitats in North Africa and the Middle East. Its population has declined drastically during the last two decades and it is classified as vulnerable. Captive breeding programmes have, hitherto, been unsuccessful in reviving population numbers and thus radical technological solutions are essential for the long term survival of this species. The purpose of this study was to investigate the use of primordial germ cell-mediated chimera technology to produce viable Houbara bustard offspring.Embryonic gonadal tissue was dissected from Houbara bustard embryos at eight days post-incubation. Subsequently, Houbara tissue containing gonadal primordial germ cells (gPGCs was injected into White Leghorn chicken (Gallus gallus domesticus embryos, producing 83/138 surviving male chimeric embryos, of which 35 chimeric roosters reached sexual maturity after 5 months. The incorporation and differentiation of Houbara gPGCs in chimeric chicken testis were assessed by PCR with Houbara-specific primers and 31.3% (5/16 gonads collected from the injected chicken embryos showed the presence of donor Houbara cells. A total of 302 semen samples from 34 chimeric roosters were analyzed and eight were confirmed as germline chimeras. Semen samples from these eight roosters were used to artificially inseminate three female Houbara bustards. Subsequently, 45 Houbara eggs were obtained and incubated, two of which were fertile. One egg hatched as a male live born Houbara; the other was female but died before hatching. Genotyping confirmed that the male chick was a pure-line Houbara derived from a chimeric rooster.This study demonstrates for the first time that Houbara gPGCs can migrate, differentiate and eventually give rise to functional sperm in the chimeric chicken testis. This approach may provide a promising tool for propagation and conservation of endangered avian

  13. Effects of water-filtered infrared-A and of heat on cell death, inflammation, antioxidative potential and of free radical formation in viable skin--first results.

    Science.gov (United States)

    Piazena, Helmut; Pittermann, Wolfgang; Müller, Werner; Jung, Katinka; Kelleher, Debra K; Herrling, Thomas; Meffert, Peter; Uebelhack, Ralf; Kietzmann, Manfred

    2014-09-05

    The effects of water-filtered infrared-A (wIRA) and of convective heat on viability, inflammation, inducible free radicals and antioxidative power were investigated in natural and viable skin using the ex vivo Bovine Udder System (BUS) model. Therefore, skin samples from differently treated parts of the udder of a healthy cow were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test, by prostaglandin E2 (PGE2) measurement and by electron spin resonance (ESR) spectroscopy. Neither cell viability, the inflammation status, the radical status or the antioxidative defence systems of the skin were significantly affected by wIRA applied within 30 min by using an irradiance of 1900 W m(-2) which is of relevance for clinical use, but which exceeded the maximum solar IR-A irradiance at the Earth's surface more than 5 times and which resulted in a skin surface temperature of about 45 °C without cooling and of about 37 °C with convective cooling by air ventilation. No significant effects on viability and on inflammation were detected when convective heat was applied alone under equivalent conditions in terms of the resulting skin surface temperatures and exposure time. As compared with untreated skin, free radical formation was almost doubled, whereas the antioxidative power was reduced to about 50% after convective heating to about 45 °C. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Of energy and survival incognito: a relationship between viable but non-culturable cells formation and inorganic polyphosphate and formate metabolism in Campylobacter jejuni.

    Science.gov (United States)

    Kassem, Issmat I; Chandrashekhar, Kshipra; Rajashekara, Gireesh

    2013-01-01

    Campylobacter jejuni is a Gram-negative food-borne bacterium that can cause mild to serious diseases in humans. A variety of stress conditions including exposure to formic acid, a weak organic acid, can cause C. jejuni to form viable but non-culturable cells (VBNC), which was proposed as a potential survival mechanism. The inability to detect C. jejuni VBNC using standard culturing techniques may increase the risk of exposure to foods contaminated with this pathogen. However, little is known about the cellular mechanisms and triggers governing VBNC formation. Here, we discuss novel mechanisms that potentially affect VBNC formation in C. jejuni and emphasize the impact of formic acid on this process. Specifically, we highlight findings that show that impairing inorganic polyphosphate (poly-P) metabolism reduces the ability of C. jejuni to form VBNC in a medium containing formic acid. We also discuss the potential effect of poly-P and formate metabolism on energy homeostasis and cognate VBNC formation. The relationship between poly-P metabolism and VBNC formation under acid stress has only recently been identified and may represent a breakthrough in understanding this phenomenon and its impact on food safety.

  15. EMERGING ENGINEERING PRINCIPLES FOR YIELD IMPROVEMENT IN MICROBIAL CELL DESIGN

    Directory of Open Access Journals (Sweden)

    Santiago Comba

    2012-10-01

    Full Text Available Metabolic Engineering has undertaken a rapid transformation in the last ten years making real progress towards the production of a wide range of molecules and fine chemicals using a designed cellular host. However, the maximization of product yields through pathway optimization is a constant and central challenge of this field. Traditional methods used to improve the production of target compounds from engineered biosynthetic pathways in non-native hosts include: codon usage optimization, elimination of the accumulation of toxic intermediates or byproducts, enhanced production of rate-limiting enzymes, selection of appropriate promoter and ribosome binding sites, application of directed evolution of enzymes, and chassis re-circuit. Overall, these approaches tend to be specific for each engineering project rather than a systematic practice based on a more generalizable strategy. In this mini-review, we highlight some novel and extensive approaches and tools intended to address the improvement of a target product formation, founded in sophisticated principles such as dynamic control, pathway genes modularization, and flux modeling.

  16. CD14+ cells from peripheral blood positively regulate hematopoietic stem and progenitor cell survival resulting in increased erythroid yield

    OpenAIRE

    Heideveld, Esther; Masiello, Francesca; Marra, Manuela; Esteghamat, Fatemehsadat; Yağcı, Nurcan; von Lindern, Marieke; Migliaccio, Anna Rita F.; van den Akker, Emile

    2015-01-01

    Expansion of erythroblasts from human peripheral blood mononuclear cells is 4- to 15-fold more efficient than that of CD34+ cells purified from peripheral blood mononuclear cells. In addition, purified CD34+ and CD34− populations from blood do not reconstitute this erythroid yield, suggesting a role for feeder cells present in blood mononuclear cells that increase hematopoietic output. Immunodepleting peripheral blood mononuclear cells for CD14+ cells reduced hematopoietic stem and progenitor...

  17. Seed priming and sulfur effects on soybean cell membrane stability and yield in saline soil

    National Research Council Canada - National Science Library

    Bejandi, Teymur Khandan; Sedghi, Mohammad; Sharifi, Raouf Seyed; Namvar, Ali; Molaei, Peyman

    2009-01-01

    The objective of this work was to determine the effects of seed priming and sulfur application on cell membrane characteristics, seedling emergence, chlorophyll content and grain yield of soybean (Glycine max) in saline soil...

  18. CD14+ cells from peripheral blood positively regulate hematopoietic stem and progenitor cell survival resulting in increased erythroid yield

    NARCIS (Netherlands)

    Heideveld, Esther; Masiello, Francesca; Marra, Manuela; Esteghamat, Fatemehsadat; Yağcı, Nurcan; von Lindern, Marieke; Migliaccio, Anna Rita F.; van den Akker, Emile

    2015-01-01

    Expansion of erythroblasts from human peripheral blood mononuclear cells is 4- to 15-fold more efficient than that of CD34(+) cells purified from peripheral blood mononuclear cells. In addition, purified CD34(+) and CD34(-) populations from blood do not reconstitute this erythroid yield, suggesting

  19. CD14+ cells from peripheral blood positively regulate hematopoietic stem and progenitor cell survival resulting in increased erythroid yield

    Science.gov (United States)

    Heideveld, Esther; Masiello, Francesca; Marra, Manuela; Esteghamat, Fatemehsadat; Yağcı, Nurcan; von Lindern, Marieke; Migliaccio, Anna Rita F.; van den Akker, Emile

    2015-01-01

    Expansion of erythroblasts from human peripheral blood mononuclear cells is 4- to 15-fold more efficient than that of CD34+ cells purified from peripheral blood mononuclear cells. In addition, purified CD34+ and CD34− populations from blood do not reconstitute this erythroid yield, suggesting a role for feeder cells present in blood mononuclear cells that increase hematopoietic output. Immunodepleting peripheral blood mononuclear cells for CD14+ cells reduced hematopoietic stem and progenitor cell expansion. Conversely, the yield was increased upon co-culture of CD34+ cells with CD14+ cells (full contact or transwell assays) or CD34+ cells re-constituted in conditioned medium from CD14+ cells. In particular, CD14++CD16+ intermediate monocytes/macrophages enhanced erythroblast outgrowth from CD34+ cells. No effect of CD14+ cells on erythroblasts themselves was observed. However, 2 days of co-culturing CD34+ and CD14+ cells increased CD34+ cell numbers and colony-forming units 5-fold. Proliferation assays suggested that CD14+ cells sustain CD34+ cell survival but not proliferation. These data identify previously unrecognized erythroid and non-erythroid CD34− and CD34+ populations in blood that contribute to the erythroid yield. A flow cytometry panel containing CD34/CD36 can be used to follow specific stages during CD34+ differentiation to erythroblasts. We have shown modulation of hematopoietic stem and progenitor cell survival by CD14+ cells present in peripheral blood mononuclear cells which can also be found near specific hematopoietic niches in the bone marrow. PMID:26294724

  20. High-Yield Method for Dispersing Simian Kidneys for Cell Cultures

    Science.gov (United States)

    de Oca, H. Montes; Probst, P.; Grubbs, R.

    1971-01-01

    A technique for dispersion of animal tissue cells is described. The proposed technique is based on the concomitant use of trypsin and disodium ethylenediamine tetraacetate (EDTA). The use of the two dispersing agents (trypsin and disodium EDTA) markedly enhances cell yield as compared with the standard cell dispersion methods. Moreover, significant reduction in the amount of time required for complete tissue dispersal, presence of a very low number of nonviable cells, less cell clumping, and more uniform monolayer formation upon cultivation compare favorably with the results usually obtained with the standard trypsinization technique. Images PMID:4993235

  1. Where Do All the Phytoplankton Go? Challenges in Keeping Track of Viable Cells in Phytoplankton Communities Using Flow Cytometry and Cell Staining

    Science.gov (United States)

    Simmons, L. J.; Fobbe, D. J.; Berges, J. A.

    2016-02-01

    Understanding the dynamics of phytoplankton communities has traditionally focused on differences in growth and related processes among taxa. It is now appreciated that differences in mortality could be equally important in contributing to these dynamics. Studying mortality in communities is difficult, especially on relevant time scales, which could be as short as hours to days. Flow cytometry can potentially provide solutions, because it can allow discrimination of different taxa, and when combined with staining, distinguish live and dead cells. We applied flow cytometry and staining to phytoplankton communities in a model system: a small, well-studied, urban pond in southeastern Wisconsin. Using flow cytometry, it was possible to resolve up to six dominant taxa (most stain also affected other fluorescence channels, requiring compensation. Correlations of numbers of dead cells with environmental factors (e.g. temperature, nutrient concentrations, irradiance) were generally poor, suggesting the greater importance of biotic versus abiotic variables in community mortality dynamics. Ongoing work is focusing on the effects of viral pathogens, grazing and allelopathic interactions using experimental manipulations and individual-based modeling.

  2. Co-infusion of autologous adipose tissue derived neuronal differentiated mesenchymal stem cells and bone marrow derived hematopoietic stem cells, a viable therapy for post-traumatic brachial plexus injury: A case report

    Directory of Open Access Journals (Sweden)

    Umang G Thakkar

    2014-08-01

    Full Text Available Stem cell therapy is emerging as a viable approach in regenerative medicine. A 31-year-old male with brachial plexus injury had complete sensory-motor loss since 16 years with right pseudo-meningocele at C5-D1 levels and extra-spinal extension up to C7-D1, with avulsion on magnetic resonance imaging and irreversible damage. We generated adipose tissue derived neuronal differentiated mesenchymal stem cells (N-AD-MSC and bone marrow derived hematopoietic stem cells (HSC-BM. Neuronal stem cells expressed β-3 tubulin and glial fibrillary acid protein which was confirmed on immunofluorescence. On day 14, 2.8 ml stem cell inoculum was infused under local anesthesia in right brachial plexus sheath by brachial block technique under ultrasonography guidance with a 1.5-inch-long 23 gauge needle. Nucleated cell count was 2 × 10 4 /μl, CD34+ was 0.06%, and CD45-/90+ and CD45-/73+ were 41.63% and 20.36%, respectively. No untoward effects were noted. He has sustained recovery with re-innervation over a follow-up of 4 years documented on electromyography-nerve conduction velocity study.

  3. Potential of Escherichia coli 0157:H7 to persist and form viable but non-culturable cells on a food-contact surface subjected to cycles of soiling and chemical treatment

    DEFF Research Database (Denmark)

    Marouani-Gadri, Nesrine; Firmesse, Olivier; Chassaing, Danielle

    2010-01-01

    only, a further increase in this proportion occurred 24 h after the CT, suggesting that some of the surviving viable but non-culturable cells finally died. This study shows that conditions leading to E. coli O157:H7 persistence are not likely to arise when good refrigeration and hygiene practices...... no longer detectable after the first week. However, on 66-hour biofilms with 6.7 log CFU/cm², after initially decreasing, E. coli numbers reached 6.6 log CFU/cm² and 8.3 log viable cells/cm² on the 11th day. When E. coli was cultured with a Comamonas testosteroni previously shown to increase E. coli biofilm...

  4. The role of stem cell mobilization regimen on lymphocyte collection yield in patients with multiple myeloma.

    Science.gov (United States)

    Hiwase, D K; Hiwase, S; Bailey, M; Bollard, G; Schwarer, A P

    2008-01-01

    The lymphocyte dose (LY-DO) infused during an autograft influences absolute lymphocyte (ALC) recovery and survival following autologous stem cell transplantation (ASCT) in multiple myeloma (MM) patients. Factors influencing lymphocyte yield (LY-C) during leukapheresis have been poorly studied. Factors that could influence survival, LY-C and CD34(+) cell yield were analyzed in 122 MM patients. Three mobilization regimens were used, granulocyte-colony-stimulating factor (G-CSF) alone (n=13), cyclophosphamide 1-2 g/m(2) plus G-CSF (LD-CY, n=62) and cyclophosphamide 3-4 g/m(2) and G-CSF (ID-CY, n=47). Using multivariate analysis, age, LY-C, ALC on day 30 (ALC-30) and International Staging System stage significantly influenced overall (OS) and progression-free survival (PFS) following ASCT. PFS (56 versus 29 months, P=0.05) and OS (72 versus 49 months; P=0.07) were longer in the LY-C>or=0.12x10(9)/kg group than the LY-Cradiotherapy and number of leukaphereses significantly influenced LY-C. Significantly higher LY-C was obtained with G-CSF alone compared with the LD-CY and ID-CY groups. CD34(+) count on the day of leukapheresis, prior chemotherapy with prednisone, cyclophosphamide, adriamycin and BCNU or melphalan, and stem cell mobilization regimen significantly influenced CD34(+) cell yield. LY-C influenced ALC-15 and survival following ASCT. Factors that influenced CD34(+) cell yield and LY-C during leukapheresis were different. Mobilization should be tailored to maximize the LY-C and CD34(+) cell yield.

  5. Microbial Electrolysis Cells for High Yield Hydrogen Gas Production from Organic Matter

    KAUST Repository

    Logan, Bruce E.

    2008-12-01

    The use of electrochemically active bacteria to break down organic matter, combined with the addition of a small voltage (>0.2 V in practice) in specially designed microbial electrolysis cells (MECs), can result in a high yield of hydrogen gas. While microbial electrolysis was invented only a few years ago, rapid developments have led to hydrogen yields approaching 100%, energy yields based on electrical energy input many times greater than that possible by water electrolysis, and increased gas production rates. MECs used to make hydrogen gas are similar in design to microbial fuel cells (MFCs) that produce electricity, but there are important differences in architecture and analytical methods used to evaluate performance. We review here the materials, architectures, performance, and energy efficiencies of these MEC systems that show promise as a method for renewable and sustainable energy production, and wastewater treatment. © 2008 American Chemical Society.

  6. Microbial electrolysis cells for high yield hydrogen gas production from organic matter.

    Science.gov (United States)

    Logan, Bruce E; Call, Douglas; Cheng, Shaoan; Hamelers, Hubertus V M; Sleutels, Tom H J A; Jeremiasse, Adriaan W; Rozendal, René A

    2008-12-01

    The use of electrochemically active bacteria to break down organic matter, combined with the addition of a small voltage (> 0.2 V in practice) in specially designed microbial electrolysis cells (MECs), can result in a high yield of hydrogen gas. While microbial electrolysis was invented only a few years ago, rapid developments have led to hydrogen yields approaching 100%, energy yields based on electrical energy input many times greater than that possible by water electrolysis, and increased gas production rates. MECs used to make hydrogen gas are similar in design to microbial fuel cells (MFCs) that produce electricity, but there are important differences in architecture and analytical methods used to evaluate performance. We review here the materials, architectures, performance, and energy efficiencies of these MEC systems that show promise as a method for renewable and sustainable energy production, and wastewater treatment.

  7. Cell type-specific and common characteristics of exosomes derived from mouse cell lines: Yield, physicochemical properties, and pharmacokinetics.

    Science.gov (United States)

    Charoenviriyakul, Chonlada; Takahashi, Yuki; Morishita, Masaki; Matsumoto, Akihiro; Nishikawa, Makiya; Takakura, Yoshinobu

    2017-01-01

    Exosomes are small membrane vesicles secreted from cells and are expected to be used as drug delivery systems. Important characteristics of exosomes, such as yield, physicochemical properties, and pharmacokinetics, may be different among different cell types. However, there is limited information about the effect of cell type on these characteristics. In the present study, we evaluated these characteristics of exosomes derived from five different types of mouse cell lines: B16BL6 murine melanoma cells, C2C12 murine myoblast cells, NIH3T3 murine fibroblasts cells, MAEC murine aortic endothelial cells, and RAW264.7 murine macrophage-like cells. Exosomes were collected using a differential ultracentrifugation method. The exosomes collected from all the cell types were negatively charged globular vesicles with a diameter of approximately 100nm. C2C12 and RAW264.7 cells produced more exosomes than the other types of cells. The exosomes were labeled with a fusion protein of Gaussia luciferase and lactadherin to evaluate their pharmacokinetics. After intravenous injection into mice, all the exosomes rapidly disappeared from the systemic circulation and mainly distributed to the liver. In conclusion, the exosome yield was significantly different among the cell types, and all the exosomes evaluated in this study showed comparable physicochemical and pharmacokinetic properties. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Constitutive expression of cell wall invertase genes increases grain yield and starch content in maize.

    Science.gov (United States)

    Li, Bei; Liu, Hua; Zhang, Yue; Kang, Tao; Zhang, Li; Tong, Jianhua; Xiao, Langtao; Zhang, Hongxia

    2013-12-01

    Grain size, number and starch content are important determinants of grain yield and quality. One of the most important biological processes that determine these components is the carbon partitioning during the early grain filling, which requires the function of cell wall invertase. Here, we showed the constitutive expression of cell wall invertase-encoding gene from Arabidopsis, rice (Oryza sativa) or maize (Zea mays), driven by the cauliflower mosaic virus (CaMV) 35S promoter, all increased cell wall invertase activities in different tissues and organs, including leaves and developing seeds, and substantially improved grain yield up to 145.3% in transgenic maize plants as compared to the wild-type plants, an effect that was reproduced in our 2-year field trials at different locations. The dramatically increased grain yield is due to the enlarged ears with both enhanced grain size and grain number. Constitutive expression of the invertase-encoding gene also increased total starch content up to 20% in the transgenic kernels. Our results suggest that cell wall invertase gene can be genetically engineered to improve both grain yield and grain quality in crop plants. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  9. Microbial Electrolysis Cells for High Yield Hydrogen Gas Production from Organic Matter

    NARCIS (Netherlands)

    Logan, B.E.; Call, D.; Cheng, S.; Hamelers, H.V.M.; Sleutels, T.H.J.A.; Jeremiasse, A.W.; Rozendal, R.A.

    2008-01-01

    The use of electrochemically active bacteria to break down organic matter, combined with the addition of a small voltage (>0.2 V in practice) in specially designed microbial electrolysis cells (MECs), can result in a high yield of hydrogen gas. While microbial electrolysis was invented only a few

  10. High Diagnostic Yield of Dedicated Pulmonary Screening before Hematopoietic Cell Transplantation in Children

    NARCIS (Netherlands)

    Versluijs, Anne Birgitta; van der Ent, Korstiaan; Boelens, Jaap J.; Wolfs, Tom; de Jong, Pim; Bierings, Marc B.

    2015-01-01

    Pulmonary complications are an important cause for treatment-related morbidity and mortality in hematopoietic cell transplantation (HCT) in children. The aim of this study was to investigate the yield of our pre-HCT pulmonary screening program. We also describe our management guidelines based on

  11. Temperature-induced labelling of Fluo-3 AM selectively yields brighter nucleus in adherent cells.

    Science.gov (United States)

    Meng, Guixian; Pan, Leiting; Li, Cunbo; Hu, Fen; Shi, Xuechen; Lee, Imshik; Drevenšek-Olenik, Irena; Zhang, Xinzheng; Xu, Jingjun

    2014-01-17

    Fluo-3 is widely used to study cell calcium. Two traditional approaches: (1) direct injection and (2) Fluo-3 acetoxymethyl ester (AM) loading, often bring conflicting results in cytoplasmic calcium ([Ca(2+)]c) and nuclear calcium ([Ca(2+)]n) imaging. AM loading usually yields a darker nucleus than in cytoplasm, while direct injection always induces a brighter nucleus which is more responsive to [Ca(2+)]n detection. In this work, we detailedly investigated the effects of loading and de-esterification temperatures on the fluorescence intensity of Fluo-3 in response to [Ca(2+)]n and [Ca(2+)]c in adherent cells, including osteoblast, HeLa and BV2 cells. Interestingly, it showed that fluorescence intensity of nucleus in osteoblast cells was about two times larger than that of cytoplasm when cells were loaded with Fluo-3 AM at 4 °C and allowed a subsequent step for de-esterification at 20 °C. Brighter nuclei were also acquired in HeLa and BV2 cells using the same experimental condition. Furthermore, loading time and adhesion quality of cells had effect on fluorescence intensity. Taken together, cold loading and room temperature de-esterification treatment of Fluo-3 AM selectively yielded brighter nucleus in adherent cells. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. A novel bioreactor and culture method drives high yields of platelets from stem cells.

    Science.gov (United States)

    Avanzi, Mauro P; Oluwadara, Oluwasijibomi E; Cushing, Melissa M; Mitchell, Maxwell L; Fischer, Stephen; Mitchell, W Beau

    2016-01-01

    Platelet (PLT) transfusion is the primary treatment for thrombocytopenia. PLTs are obtained exclusively from volunteer donors, and the PLT product has only a 5-day shelf life, which can limit supply and result in PLT shortages. PLTs derived from stem cells could help to fill this clinical need. However, current culture methods yield far too few PLTs for clinical application. To address this need, a defined, serum-free culture method was designed using a novel bioreactor to increase the yield of PLTs from stem cell-derived megakaryocytes. CD34 cells isolated from umbilical cord blood were expanded with a variety of reagents and on a nanofiber membrane using serum-free medium. These cells were then differentiated into megakaryocytic lineage by culturing with thrombopoietin and stem cell factor in serum-free conditions. Polyploidy was induced by addition of Rho kinase inhibitor or actin polymerization inhibitor to the CD41 cells. A novel bioreactor was developed that recapitulated aspects of the marrow vascular niche. Polyploid megakaryocytes that were subjected to flow in the bioreactor extended proPLTs and shed PLTs, as confirmed by light microscopy, fluorescence imaging, and flow cytometry. CD34 cells were expanded 100-fold. CD41 cells were expanded 100-fold. Up to 100 PLTs per input megakaryocyte were produced from the bioreactor, for an overall yield of 10(6) PLTs per input CD34 cell. The PLTs externalized P-selectin after activation. Functional PLTs can be produced ex vivo on a clinically relevant scale using serum-free culture conditions with a novel stepwise approach and an innovative bioreactor. © 2015 AABB.

  13. Construction high-yield candidate influenza vaccine viruses in Vero cells by reassortment.

    Science.gov (United States)

    Yu, Wei; Yang, Fan; Yang, Jinghui; Ma, Lei; Cun, Yina; Song, Shaohui; Liao, Guoyang

    2016-11-01

    Usage of influenza vaccine is the best choice measure for preventing and conclusion of influenza virus infection. Although it has been used of chicken embryo to produce influenza vaccine, following with WHO recommended vaccine strain, there were uncontrollable factors and its deficiencies, specially, during an influenza pandemic in the world. The Vero cells are used for vaccine production of a few strains including influenza virus, because of its homology with human, recommended by WHO. However, as known most of the influenza viruses strains could not culture by Vero cells. It was used two high-yield influenza viruses adapted in Vero cells as donor viruses, such as A/Yunnan/1/2005Va (H3N2) and B/Yunnan/2/2005Va (B), to construct high-yield wild influenza virus in Vero cells under antibody selection pressure. After reassortment and passages, it obtained the new Vaccine strains with A/Tianjin/15/2009Va (H1N1), A/Fujian/196/2009Va (H3N2) and B/Chongqing/1384/2010Va (B), which was not only completely keeping their original antigenic (HA and NA), but also grown well in Vero cells with high-yield. All results of gene analysis and HA, HI shown that this reassortment method could be used to find new direction to product the influenza vaccine. J. Med. Virol. 88:1914-1921, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  14. Increased bacterial cell density and recombinant protein yield using a commercial microbial cultivation system.

    Science.gov (United States)

    Peck, Grantley R; Bowden, Timothy R; Shiell, Brian J; Michalski, Wojtek P

    2014-01-01

    EnBase (BioSilta, Finland) is a microbial cultivation system that replicates fed-batch systems through sustained release of glucose by enzymatic degradation of a polymeric substrate. Achievable bacterial cell densities and recombinant capripoxvirus protein expression levels, solubility, and antigenicity using the EnBase system were assessed. BL21-AI Escherichia coli expressing capripoxvirus proteins achieved up to eightfold higher cell densities when grown in EnBase media compared with standard media. Greater yields of capripoxvirus proteins were attained using EnBase media, either through increases in the amount of expressed protein per cell in conjunction with higher cell density or through the increase in cell density alone. Addition of EnBase booster enhanced protein yield for one of the proteins tested but reduced yield for the other. However, the amount of soluble forms of the capripoxvirus proteins tested was not different from that observed from cultures grown under standard conditions. Purified capripoxvirus proteins expressed using EnBase or standard media were assessed for their performance by enzyme-linked immunosorbent assay (ELISA) and were shown to be equally capable of specifically binding capripoxvirus antibodies.

  15. A new somatic cell count index to more accurately predict milk yield losses

    Directory of Open Access Journals (Sweden)

    J. Jeretina

    2017-10-01

    Full Text Available Intramammary infection and clinical mastitis in dairy cows leads to considerable economic losses for farmers. The somatic cell concentration in cow's milk has been shown to be an excellent indicator for the prevalence of subclinical mastitis. In this study, a new somatic cell count index (SCCI was proposed for the accurate prediction of milk yield losses caused by elevated somatic cell count (SCC. In all, 97 238 lactations (55 207 Holstein cows from 2328 herds were recorded between 2010 and 2014 under different scenarios (high and low levels of SCC, four lactation stages, different milk yield intensities, and parities (1, 2, and  ≥  3. The standard shape of the curve for SCC was determined using completed standard lactations of healthy cows. The SCCI was defined as the sum of the differences between the measured interpolated values of the natural logarithm of SCC (ln(SCC and the values for the standard shape of the curve for SCC for a particular period, divided by the total area enclosed by the standard curve and upper limit of ln(SCC  =  10 for SCC. The phenotypic potential of milk yield (305-day milk yield – MY305 was calculated using regression coefficients estimated from the linear regression model for parity and breeding values of cows for milk yield. The extent of daily milk yield loss caused by increased SCC was found to be mainly related to the early stage of lactation. Depending on the possible scenarios, the estimated milk yield loss from MY305 for primiparous cows was at least 0.8 to 0.9 kg day−1 and for multiparous cows it ranged from 1.3 to 4.3 kg day−1. Thus, the SCCI was a suitable indicator for estimating daily milk yield losses associated with increased SCC and might provide farmers reliable information to take appropriate measures for ensuring good health of cows and reducing milk yield losses at the herd level.

  16. High Yield of Adult Oligodendrocyte Lineage Cells Obtained from Meningeal Biopsy

    Directory of Open Access Journals (Sweden)

    Sissi Dolci

    2017-10-01

    Full Text Available Oligodendrocyte loss can lead to cognitive and motor deficits. Current remyelinating therapeutic strategies imply either modulation of endogenous oligodendrocyte precursors or transplantation of in vitro expanded oligodendrocytes. Cell therapy, however, still lacks identification of an adequate source of oligodendrocyte present in adulthood and able to efficiently produce transplantable cells. Recently, a neural stem cell-like population has been identified in meninges. We developed a protocol to obtain high yield of oligodendrocyte lineage cells from one single biopsy of adult rat meningeal tissue. From 1 cm2 of adult rat spinal cord meninges, we efficiently expanded a homogenous culture of 10 millions of meningeal-derived oligodendrocyte lineage cells in a short period of time (approximately 4 weeks. Meningeal-derived oligodendrocyte lineage cells show typical mature oligodendrocyte morphology and express specific oligodendrocyte markers, such as galactosylceramidase and myelin basic protein. Moreover, when transplanted in a chemically demyelinated spinal cord model, meningeal-derived oligodendrocyte lineage cells display in vivo-remyelinating potential. This oligodendrocyte lineage cell population derives from an accessible and adult source, being therefore a promising candidate for autologous cell therapy of demyelinating diseases. In addition, the described method to differentiate meningeal-derived neural stem cells into oligodendrocyte lineage cells may represent a valid in vitro model to dissect oligodendrocyte differentiation and to screen for drugs capable to promote oligodendrocyte regeneration.

  17. Engineering high yields of secondary metabolites in Rubia cell cultures through transformation with rol genes.

    Science.gov (United States)

    Bulgakov, Victor P; Shkryl, Yuri N; Veremeichik, Galina N

    2010-01-01

    Among the different methods currently used to improve yields of secondary metabolites in cultured plant cells, the method involving transformation by rol genes represents an example of relatively new technology. These genes, isolated from plasmids of the plant pathogen Agrobacterium rhizogenes, are potential activators of secondary metabolism in transformed cells from the Solanaceae, Araliaceae, Rubiaceae, Vitaceae, and Rosaceae families. In some cases, the activator effect of individual rol genes was sufficient to overcome the inability of cultured plant cells to produce large amounts of secondary metabolites. Stimulation of production characteristics of cultured plant cells mediated by the rol genes was shown to be remarkably stable over long-term cultivation. In this chapter, we describe transformation of Rubia cordifolia L. cells with the rol genes as an example of metabolic engineering of secondary metabolites.

  18. Improved poliovirus D-antigen yields by application of different Vero cell cultivation methods.

    Science.gov (United States)

    Thomassen, Yvonne E; Rubingh, Olaf; Wijffels, René H; van der Pol, Leo A; Bakker, Wilfried A M

    2014-05-19

    Vero cells were grown adherent to microcarriers (Cytodex 1; 3 g L(-1)) using animal component free media in stirred-tank type bioreactors. Different strategies for media refreshment, daily media replacement (semi-batch), continuous media replacement (perfusion) and recirculation of media, were compared with batch cultivation. Cell densities increased using a feed strategy from 1×10(6) cells mL(-1) during batch cultivation to 1.8, 2.7 and 5.0×10(6) cells mL(-1) during semi-batch, perfusion and recirculation, respectively. The effects of these different cell culture strategies on subsequent poliovirus production were investigated. Increased cell densities allowed up to 3 times higher D-antigen levels when compared with that obtained from batch-wise Vero cell culture. However, the cell specific D-antigen production was lower when cells were infected at higher cell densities. This cell density effect is in good agreement with observations for different cell lines and virus types. From the evaluated alternative culture methods, application of a semi-batch mode of operations allowed the highest cell specific D-antigen production. The increased product yields that can easily be reached using these higher cell density cultivation methods, showed the possibility for better use of bioreactor capacity for the manufacturing of polio vaccines to ultimately reduce vaccine cost per dose. Further, the use of animal-component-free cell- and virus culture media shows opportunities for modernization of human viral vaccine manufacturing. Copyright © 2014. Published by Elsevier Ltd.

  19. Accelerated high-yield generation of limb-innervating motor neurons from human stem cells

    Science.gov (United States)

    Amoroso, Mackenzie W.; Croft, Gist F.; Williams, Damian J.; O’Keeffe, Sean; Carrasco, Monica A.; Davis, Anne R.; Roybon, Laurent; Oakley, Derek H.; Maniatis, Tom; Henderson, Christopher E.; Wichterle, Hynek

    2013-01-01

    Human pluripotent stem cells are a promising source of differentiated cells for developmental studies, cell transplantation, disease modeling, and drug testing. However, their widespread use even for intensely studied cell types like spinal motor neurons is hindered by the long duration and low yields of existing protocols for in vitro differentiation and by the molecular heterogeneity of the populations generated. We report a combination of small molecules that within 3 weeks induce motor neurons at up to 50% abundance and with defined subtype identities of relevance to neurodegenerative disease. Despite their accelerated differentiation, motor neurons expressed combinations of HB9, ISL1 and column-specific markers that mirror those observed in vivo in human fetal spinal cord. They also exhibited spontaneous and induced activity, and projected axons towards muscles when grafted into developing chick spinal cord. Strikingly, this novel protocol preferentially generates motor neurons expressing markers of limb-innervating lateral motor column motor neurons (FOXP1+/LHX3−). Access to high-yield cultures of human limb-innervating motor neuron subtypes will facilitate in-depth study of motor neuron subtype-specific properties, disease modeling, and development of large-scale cell-based screening assays. PMID:23303937

  20. Three-dimensional spheroid culture of human umbilical cord mesenchymal stem cells promotes cell yield and stemness maintenance.

    Science.gov (United States)

    Li, Yi; Guo, Gang; Li, Li; Chen, Fei; Bao, Ji; Shi, Yu-Jun; Bu, Hong

    2015-05-01

    Mesenchymal stem cell (MSC) transplantation is a promising treatment of many diseases. However, conventional techniques with cells being cultured as a monolayer result in slow cell proliferation and insufficient yield to meet clinical demands. Three-dimensional (3D) culture systems are gaining attention with regard to recreating a complex microenvironment and to understanding the conditions experienced by cells. Our aim is to establish a novel 3D system for the culture of human umbilical cord MSCs (hUC-MSCs) within a real 3D microenvironment but with no digestion or passaging. Primary hUC-MSCs were isolated and grown in serum-free medium (SFM) on a suspension Rocker system. Cell characteristics including proliferation, phenotype and multipotency were recorded. The therapeutic effects of 3D-cultured hUC-MSCs on carbon tetrachloride (CCl4)-induced acute liver failure in mouse models were examined. In the 3D Rocker system, hUC-MSCs formed spheroids in SFM and maintained high viability and active proliferation. Compared with monolayer culture, the 3D-culture system yielded more hUC-MSCs cells within the same volume. The spheroids expressed higher levels of stem cell markers and displayed stronger multipotency. After transplantation into mouse, 3D hUC-MSCs significantly promoted the secretion of interferon-γ and interleukin-6 but inhibited that of tumor necrosis factor-α, thereby alleviating liver necrosis and promoting regeneration following CCl4 injury. The 3D culture of hUC-MSCs thus promotes cell yield and stemness maintenance and represents a promising strategy for hUC-MSCs expansion on an industrial scale with great potential for cell therapy and biotechnology.

  1. Transient treatment with epigenetic modifiers yields stable neuroblastoma stem cells resembling aggressive large-cell neuroblastomas.

    Science.gov (United States)

    Ikegaki, Naohiko; Shimada, Hiroyuki; Fox, Autumn M; Regan, Paul L; Jacobs, Joshua R; Hicks, Sakeenah L; Rappaport, Eric F; Tang, Xao X

    2013-04-09

    Cancer stem cells (CSCs) are plastic in nature, a characteristic that hampers cancer therapeutics. Neuroblastoma (NB) is a pediatric tumor of neural crest origin, and half of the cases are highly aggressive. By treating NB cell lines [SKNAS, SKNBE(2)C, CHP134, and SY5Y] with epigenetic modifiers for a short time, followed by sphere-forming culture conditions, we have established stem cell-like NB cells that are phenotypically stable for more than a year. These cells are characterized by their high expression of stemness factors, stem cell markers, and open chromatin structure. We referred to these cells as induced CSCs (iCSCs). SKNAS iCSC and SKNBE(2)C iCSC clones (as few as 100 cells) injected s.c. into SCID/Beige mice formed tumors, and in one case, SKNBE(2)C iCSCs metastasized to the adrenal gland, suggesting their increased metastatic potential. SKNAS iCSC xenografts showed the histologic appearance of totally undifferentiated large-cell NBs (LCNs), the most aggressive and deadly form of NB in humans. Immunohistochemical analyses showed that SKNAS iCSC xenografts expressed high levels of the stem cell marker CXCR4, whereas the SKNAS monolayer cell xenografts did not. The patterns of CXCR4 and MYC expression in SKNAS iCSC xenografts resembled those in the LCNs. The xenografts established from the NB iCSCs shared two common features: the LCN phenotype and high-level MYC/MYCN expression. These observations suggest both that NB cells with large and vesicular nuclei, representing their open chromatin structure, are indicative of stem cell-like tumor cells and that epigenetic changes may have contributed to the development of these most malignant NB cells.

  2. A High Yield and Cost-efficient Expression System of Human Granzymes in Mammalian Cells.

    Science.gov (United States)

    Dotiwala, Farokh; Fellay, Isabelle; Filgueira, Luis; Martinvalet, Denis; Lieberman, Judy; Walch, Michael

    2015-06-10

    When cytotoxic T lymphocytes (CTL) or natural killer (NK) cells recognize tumor cells or cells infected with intracellular pathogens, they release their cytotoxic granule content to eliminate the target cells and the intracellular pathogen. Death of the host cells and intracellular pathogens is triggered by the granule serine proteases, granzymes (Gzms), delivered into the host cell cytosol by the pore forming protein perforin (PFN) and into bacterial pathogens by the prokaryotic membrane disrupting protein granulysin (GNLY). To investigate the molecular mechanisms of target cell death mediated by the Gzms in experimental in-vitro settings, protein expression and purification systems that produce high amounts of active enzymes are necessary. Mammalian secreted protein expression systems imply the potential to produce correctly folded, fully functional protein that bears posttranslational modification, such as glycosylation. Therefore, we used a cost-efficient calcium precipitation method for transient transfection of HEK293T cells with human Gzms cloned into the expression plasmid pHLsec. Gzm purification from the culture supernatant was achieved by immobilized nickel affinity chromatography using the C-terminal polyhistidine tag provided by the vector. The insertion of an enterokinase site at the N-terminus of the protein allowed the generation of active protease that was finally purified by cation exchange chromatography. The system was tested by producing high levels of cytotoxic human Gzm A, B and M and should be capable to produce virtually every enzyme in the human body in high yields.

  3. Tumor-selective replication herpes simplex virus-based technology significantly improves clinical detection and prognostication of viable circulating tumor cells

    DEFF Research Database (Denmark)

    Zhang, Wen; Bao, Li; Yang, Shaoxing

    2016-01-01

    Detection of circulating tumor cells remains a significant challenge due to their vast physical and biological heterogeneity. We developed a cell-surface-marker-independent technology based on telomerase-specific, replication-selective oncolytic herpes-simplex-virus-1 that targets telomerase...

  4. Enzymatic isolation of viable human odontoblasts.

    Science.gov (United States)

    Cuffaro, H M; Pääkkönen, V; Tjäderhane, L

    2016-05-01

    To improve an enzymatic method previously used for isolation of rat odontoblasts to isolate viable mature human odontoblasts. Collagenase I, collagenase I/hyaluronidase mixture and hyaluronidase were used to extract mature human odontoblasts from the pulp chamber. Detachment of odontoblasts from dentine was determined with field emission scanning electron microscopy (FESEM) and to analyse the significance of differences in tubular diameter, and the t-test was used. MTT-reaction was used to analyse cell viability, and nonparametric Kruskal-Wallis and Mann-Whitney post hoc tests were used to analyse the data. Immunofluorescent staining of dentine sialoprotein (DSP), aquaporin-4 (AQP4) and matrix metalloproteinase-20 (MMP-20) and quantitative PCR (qPCR) of dentine sialophosphoprotein (DSPP) were used to confirm the odontoblastic nature of the cells. MTT-reaction and FESEM demonstrated collagenase I/hyaluronidase resulted in more effective detachment and higher viability than collagenase I alone. Hyaluronidase alone was not able to detach odontoblasts. Immunofluorescence revealed the typical odontoblastic-morphology with one process, and DSP, AQP4 and MMP-20 were detected. Quantitative PCR of DSPP confirmed that the isolated cells expressed this odontoblast-specific gene. The isolation of viable human odontoblasts was successful. The cells demonstrated morphology typical for odontoblasts and expressed characteristic odontoblast-type genes and proteins. This method will enable new approaches, such as apoptosis analysis, for studies using fully differentiated odontoblasts. © 2015 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  5. The impact of silicon solar cell architecture and cell interconnection on energy yield in hot & sunny climates

    KAUST Repository

    Haschke, Jan

    2017-03-23

    Extensive knowledge of the dependence of solar cell and module performance on temperature and irradiance is essential for their optimal application in the field. Here we study such dependencies in the most common high-efficiency silicon solar cell architectures, including so-called Aluminum back-surface-field (BSF), passivated emitter and rear cell (PERC), passivated emitter rear totally diffused (PERT), and silicon heterojunction (SHJ) solar cells. We compare measured temperature coefficients (TC) of the different electrical parameters with values collected from commercial module data sheets. While similar TC values of the open-circuit voltage and the short circuit current density are obtained for cells and modules of a given technology, we systematically find that the TC under maximum power-point (MPP) conditions is lower in the modules. We attribute this discrepancy to additional series resistance in the modules from solar cell interconnections. This detrimental effect can be reduced by using a cell design that exhibits a high characteristic load resistance (defined by its voltage-over-current ratio at MPP), such as the SHJ architecture. We calculate the energy yield for moderate and hot climate conditions for each cell architecture, taking into account ohmic cell-to-module losses caused by cell interconnections. Our calculations allow us to conclude that maximizing energy production in hot and sunny environments requires not only a high open-circuit voltage, but also a minimal series-to-load-resistance ratio.

  6. A high-yielding, generic fed-batch process for recombinant antibody production of GS-engineered cell lines

    DEFF Research Database (Denmark)

    Fan, Li; Zhao, Liang; Sun, Yating

    2009-01-01

    . Compared to batch cultures, the fed-batch technology generated the magnitude of the increase in cell yields (5 fold) and final antibody concentrations (4-8 fold). The majority of the increase in final antibody concentration was functions of the increased cell density and the prolonged culture time....... This generic and high-yielding fed-batch process would shorten development time, and ensure process stability, thereby facilitating the manufacture of therapeutic antibodies by GS-engineered cell lines....

  7. High-yield Escherichia coli-based cell-free expression of human proteins

    Energy Technology Data Exchange (ETDEWEB)

    Michel, Erich; Wuethrich, Kurt, E-mail: wuthrich@mol.biol.ethz.ch [ETH Zurich, Institute of Molecular Biology and Biophysics (Switzerland)

    2012-05-15

    Production of sufficient amounts of human proteins is a frequent bottleneck in structural biology. Here we describe an Escherichia coli-based cell-free system which yields mg-quantities of human proteins in N-terminal fusion constructs with the GB1 domain, which show significantly increased translation efficiency. A newly generated E. coli BL21 (DE3) RIPL-Star strain was used, which contains a variant RNase E with reduced activity and an excess of rare-codon tRNAs, and is devoid of lon and ompT protease activity. In the implementation of the expression system we used freshly in-house prepared cell extract. Batch-mode cell-free expression with this setup was up to twofold more economical than continuous-exchange expression, with yields of 0.2-0.9 mg of purified protein per mL of reaction mixture. Native folding of the proteins thus obtained is documented with 2D [{sup 15}N,{sup 1}H]-HSQC NMR.

  8. The binding of rhBMP-2 to the receptors of viable MC3T3-E1 cells and the question of cooperativity

    Energy Technology Data Exchange (ETDEWEB)

    Wiemann, M.; Rumpf, H.M.; Bingmann, D.; Jennissen, H.P. [Universitaetsklinikum Essen (Germany). Inst. fuer Physiologie; Universitaetsklinikum Essen (Germany). Inst. fuer Physiologiesche Chemie

    2001-12-01

    The binding of rhBMP-2 to its receptors, the signal transduction cascade and the final responses of bone cells, osteoprogenitor cells and derived cell lines is of high fundamental and clinical interest. In this report concentration-response curves of the osteoblast cell line MC3T3-E1 under influence of rhBMP-2 was investigated. The biological response of the cells (corresponding to a down-stream effect of the receptor state-function) was monitored in pilot experiments by the MC3T3-cell alkaline phosphatase-induction test (MC3T3-cell ALP-induction test). It is shown that the MC3T3-cell ALP-induction test is a good tool for measuring biologically active recombinant human BMP-2 (rhBMP-2) in crude extracts of E. coli as well as in highly purified form. In addition this test is very sensitive to chemically induced structural changes of rhBMP-2 such as those resulting from a radiolabeling of rhBMP-2 by the Bolton-Hunter procedure. The latter procedure reduces the biological activity of rhBMP-2 by a factor of 3-4. The measured concentration-response curves could all be non-linearly fitted to a rectangular hyperbola. The half-maximal saturation, K{sub 0.5}, is found between 30-100 nM rhBMP-2 (= 0.8-2.5 {mu}g/ml). The effect of rhBMP-2 shows a plateau i.e. maximal response at ca. 300-1000 nM rhBMP-2 (= 8-25 {mu}g/ml). The data thus indicate a non-cooperative binding-response behavior. This was unexpected since BMP-2 binds simultaneously to two cooperating receptors of type 1 and type 2. However in the very low concentration range of employed rhBMP-2 a variable response of the cells was measured so that a full exclusion of cooperativity cannot be concluded at the present time. This will be clarified by future experiments. (orig.)

  9. An invention of thermo-responsive polymer surface, yielding cell sheet based regenerative therapies in cardiology and ophthalmology

    Directory of Open Access Journals (Sweden)

    Sawa Y

    2015-12-01

    Full Text Available The Invention: In vitro cell culture methodologies provide a conducive environment for the cells taken out of their native environment to grow and proliferate in a non-physiological environment, the culture dish. Research experiments have been focusing on various criteria for assessing how far it is possible to recapitulate the native extra-cellular environment in vitro. Scaffolds, culture media, growth factors and cell surface modified culture dishes are some of the components that provide a conducive environment for in vitro cell culture. Cells that are grown in culture dishes using conventional methodologies are usually detached using enzymatic treatment with Trypsin, Collagenase etc [1], to be transplanted when it comes to a clinical or experimental application. Such enzymes used in separating the cells may have some damaging effects to the cell membranes which might impair the cell function [1]. However, cells if can be grown as a monolayer and be harvested as a contiguous cell sheet, it is considered suitable for transplantation in certain specific applications. In addition to that, if enzymatic digestion which has some detrimental effects on the detached cells could be avoided, that is an added advantage. The work by Prof. Okano and team from the Tokyo Women's Medical University, Japan, on thermo-responsive polymer surfaces has yielded a solution which has both the advantages viz., detachability of cells grown as a monolayer in the form of a cell sheet, that too without the use of enzymes. Their research into biomaterials for more than two decades has yielded a thermo-responsive polymer, the poly(N-isopropylacrylamide (PIPAAm [1] coated culture dish for cell sheet engineering. In their technology, PIPAAm is polymerized and grafted to tissue culture polystyrene (TCPS dishes. Cells have been found to grow confluent on PIPAAm-TCPS at 37 °C. Once confluent as a monolayer, by merely reducing the temperature of the PIPAAm-TCPS to 20 °C, it

  10. Enumeration of viable and non-viable larvated Ascaris eggs with quantitative PCR.

    Science.gov (United States)

    Raynal, Maria; Villegas, Eric N; Nelson, Kara L

    2012-12-01

    The goal of this study was to further develop an incubation-quantitative polymerase chain reaction (qPCR) method for quantifying viable Ascaris eggs by characterizing the detection limit and number of template copies per egg, determining the specificity of the method, and testing the method with viable and inactivated larvated eggs. The number of template copies per cell was determined by amplifying DNA from known numbers of eggs at different development stages; the value was estimated to be 32 copies. The specificity of the method was tested against a panel of bacteria, fungi, protozoa and helminths, and no amplification was found with non-target DNA. Finally, fully larvated eggs were inactivated by four different treatments: 254 nm ultraviolet light, 2,000 ppm NH(3)-N at pH 9, moderate heat (48 °C) and high heat (70 °C). Concentrations of treated eggs were measured by direct microscopy and incubation-qPCR. The qPCR signal decreased following all four treatments, and was in general agreement with the decrease in viable eggs determined by microscopy. The incubation-qPCR method for enumerating viable Ascaris eggs is a promising approach that can produce results faster than direct microscopy, and may have benefits for applications such as assessing biosolids.

  11. Does the preference of peripheral versus central venous access in peripheral blood stem cell collection/yield change stem cell kinetics in autologous stem cell transplantation?

    Science.gov (United States)

    Dogu, Mehmet Hilmi; Kaya, Ali Hakan; Berber, Ilhami; Sari, İsmail; Tekgündüz, Emre; Erkurt, Mehmet Ali; Iskender, Dicle; Kayıkçı, Ömur; Kuku, Irfan; Kaya, Emin; Keskin, Ali; Altuntaş, Fevzi

    2016-02-01

    Central venous access is often used during apheresis procedure in stem cell collection. The aim of the present study was to evaluate whether central or peripheral venous access has an effect on stem cell yield and the kinetics of the procedure and the product in patients undergoing ASCT after high dose therapy. A total of 327 patients were retrospectively reviewed. The use of peripheral venous access for stem cell yield was significantly more frequent in males compared to females (p = 0.005). Total volume of the product was significantly lower in central venous access group (p = 0.046). As being a less invasive procedure, peripheral venous access can be used for stem cell yield in eligible selected patients. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Influence of milk somatic cell content on Parmigiano-Reggiano cheese yield.

    Science.gov (United States)

    Summer, Andrea; Franceschi, Piero; Formaggioni, Paolo; Malacarne, Massimo

    2015-05-01

    The aim of this study was to determine the influence of the somatic cell content (SCC) of milk on Parmigiano-Reggiano cheese yield, produced in commercial cheese factories under field conditions. The study was carried out following the production of 56 batches of Parmigiano-Reggiano in 13 commercial cheese factories by processing milk collected from Italian Friesian cattle herds. The vat-milk (V-milk) used for making each cheese batch was obtained by mixing evening milk (partially skimmed following spontaneous separation of fat overnight, natural creaming) and morning milk. The batches of cheese produced were divided into 5 classes according to the SCC value of the evening milk determined prior to natural creaming (class 1, from 0 to 200,000; 2, 201,000-300,000; 3, 301,000-400,000; 4, 401,000-500,000; 5, over 501,000 cells/ml). The cheese yield was calculated as the amount of 24-h cheese, expressed in kilograms, obtained from 100 kg of V-milk (24 h ACY). The values of fat, crude protein, true protein, casein and 24 h ACY of V-milk were negatively correlated with the somatic cell score (SCS) of the evening milk. Conversely, a positive correlation was observed between chloride and SCS. Fat, protein fractions (crude protein, casein and whey proteins), P and titratable acidity of V-milk were positively correlated with its 24 h ACY, while chloride, pH and SCS showed a negative correlation. A significant drop in 24 h ACY was observed in classes 3, 4 and 5, therefore when the SCC of the evening milk exceeded 300,000 cells/ml. Finally a lower recovery of milk fat in cheese was observed as SCC of evening milk increase.

  13. The effect of bone marrow aspiration strategy on the yield and quality of human mesenchymal stem cells

    NARCIS (Netherlands)

    Fennema, E.M.; Renard, Auke J.S.; Mentink-Leusink, Anouk; van Blitterswijk, Clemens; de Boer, Jan

    2009-01-01

    Introduction Large inter-donor differences exist in human mesenchymal stem cell (hMSC) yield and the response of these cells to osteogenic stimuli. The source of these differences may be clinical differences in stem cell characteristics between individuals or the aspiration procedure itself. Methods

  14. Influence of scrotal bipartition on spermatogenesis yield and sertoli cell efficiency in sheep

    Directory of Open Access Journals (Sweden)

    Ramon T.G.A. Rodrigues

    2016-04-01

    Full Text Available Abstract With the objective to assess the effect of scrotal bipartition on spermatogenesis in sheep, the testes were used from 12 crossbred rams of sheep farms in the municipality of Patos, Paraíba, Brazil, distributed into two groups: GI with six rams with scrotal bipartition, and GII with six rams without scrotal bipartition. The testicular biometry was measured and the testes were collected, fixed in Bouin and fragments were processed to obtain histological slides. The spermatogenesis yield and the Sertoli cell efficiency was estimated by counting the cells of the spermatogenetic line at stage one of the seminiferous epithelium cycle and the Sertoli cells. The results were submitted to analysis of variance with the ASSISTAT v.7.6 program and the mean values were compared by the Student-Newman-Keuls test (SNK at 5% significance. The testicular biometric parameters did not show statistical difference (p>0.05 between the groups. The meiotic, spermatogenetic and Sertoli cell efficiency were higher in bipartitioned rams (p0.05 between GI and GII. The results indicated that there is superiority in the spermatogenetic parameters of bi-partitioned rams, suggesting that these sheep present, as reported in goats, indication of better reproductive indices.

  15. Optimizing Viable Leukocyte Sampling from the Female Genital Tract for Clinical Trials: An International Multi-Site Study

    Science.gov (United States)

    De Rosa, Stephen C.; Martinson, Jeffrey A.; Plants, Jill; Brady, Kirsten E.; Gumbi, Pamela P.; Adams, Devin J.; Vojtech, Lucia; Galloway, Christine G.; Fialkow, Michael; Lentz, Gretchen; Gao, Dayong; Shu, Zhiquan; Nyanga, Billy; Izulla, Preston; Kimani, Joshua; Kimwaki, Steve; Bere, Alfred; Moodie, Zoe; Landay, Alan L.; Passmore, Jo-Ann S.; Kaul, Rupert; Novak, Richard M.; McElrath, M. Juliana; Hladik, Florian

    2014-01-01

    Background Functional analysis of mononuclear leukocytes in the female genital mucosa is essential for understanding the immunologic effects of HIV vaccines and microbicides at the site of HIV exposure. However, the best female genital tract sampling technique is unclear. Methods and Findings We enrolled women from four sites in Africa and the US to compare three genital leukocyte sampling methods: cervicovaginal lavages (CVL), endocervical cytobrushes, and ectocervical biopsies. Absolute yields of mononuclear leukocyte subpopulations were determined by flow cytometric bead-based cell counting. Of the non-invasive sampling types, two combined sequential cytobrushes yielded significantly more viable mononuclear leukocytes than a CVL (pbiopsies. Sample yields were consistent between sites. In a subgroup analysis, we observed significant reproducibility between replicate same-day biopsies (r = 0.89, p = 0.0123). Visible red blood cells in cytobrushes increased leukocyte yields more than three-fold (p = 0.0078), but did not change their subpopulation profile, indicating that these leukocytes were still largely derived from the mucosa and not peripheral blood. We also confirmed that many CD4+ T cells in the female genital tract express the α4β7 integrin, an HIV envelope-binding mucosal homing receptor. Conclusions CVL sampling recovered the lowest number of viable mononuclear leukocytes. Two cervical cytobrushes yielded comparable total numbers of viable leukocytes to one biopsy, but cytobrushes and biopsies were biased toward macrophages and T lymphocytes, respectively. Our study also established the feasibility of obtaining consistent flow cytometric analyses of isolated genital cells from four study sites in the US and Africa. These data represent an important step towards implementing mucosal cell sampling in international clinical trials of HIV prevention. PMID:24454917

  16. The Chemically Synthesized Ageladine A-Derivative LysoGlow84 Stains Lysosomes in Viable Mammalian Brain Cells and Specific Structures in the Marine Flatworm Macrostomum lignano

    Directory of Open Access Journals (Sweden)

    Thorsten Mordhorst

    2015-02-01

    Full Text Available Based on the chemical structure and the known chemical synthesis of the marine sponge alkaloid ageladine A, we synthesized the ageladine A-derivative 4-(naphthalene-2-yl-1H-imidazo[4,5-c]pyridine trifluoroacetate (LysoGlow84. The two-step synthesis started with the Pictet-Spengler reaction of histamine and naphthalene-2-carbaldehyde to a tetrahydropyridine intermediate, which was dehydrogenated with activated manganese (IV oxide to LysoGlow84. Structure and purity of the synthesized LysoGlow84 were confirmed by NMR spectroscopy and mass spectrometry. The fluorescence intensity emitted by LysoGlow84 depended strongly on the pH of the solvent with highest fluorescence intensity recorded at pH 4. The fluorescence maximum (at 315 nm excitation was observed at 440 nm. Biocompatibility of LysoGlow84 was investigated using cultured rat brain astrocytes and the marine flatworm Macrostomum lignano. Exposure of the astrocytes for up to 6 h to micromolar concentrations of LysoGlow84 did not compromise cell viability, as demonstrated by several viability assays, but revealed a promising property of this compound for staining of cellular vesicles. Conventional fluorescence microscopy as well as confocal scanning microscopy of LysoGlow84-treated astrocytes revealed co-localization of LysoGlow84 fluorescence with that of LysoTracker® Red DND-99. LysoGlow84 stained unclear structures in Macrostomum lignano, which were identified as lysosomes by co-staining with LysoTracker. Strong fluorescence staining by LysoGlow84 was further observed around the worms’ anterior gut and the female genital pore which were not counterstained by LysoTracker Red. Thus, LysoGlow84 is a new promising dye that stains lysosomes and other acidic compartments in cultured cells and in worms.

  17. The chemically synthesized ageladine A-derivative LysoGlow84 stains lysosomes in viable mammalian brain cells and specific structures in the marine flatworm Macrostomum lignano.

    Science.gov (United States)

    Mordhorst, Thorsten; Awal, Sushil; Jordan, Sebastian; Petters, Charlotte; Sartoris, Linda; Dringen, Ralf; Bickmeyer, Ulf

    2015-02-11

    Based on the chemical structure and the known chemical synthesis of the marine sponge alkaloid ageladine A, we synthesized the ageladine A-derivative 4-(naphthalene-2-yl)-1H-imidazo[4,5-c]pyridine trifluoroacetate (LysoGlow84). The two-step synthesis started with the Pictet-Spengler reaction of histamine and naphthalene-2-carbaldehyde to a tetrahydropyridine intermediate, which was dehydrogenated with activated manganese (IV) oxide to LysoGlow84. Structure and purity of the synthesized LysoGlow84 were confirmed by NMR spectroscopy and mass spectrometry. The fluorescence intensity emitted by LysoGlow84 depended strongly on the pH of the solvent with highest fluorescence intensity recorded at pH 4. The fluorescence maximum (at 315 nm excitation) was observed at 440 nm. Biocompatibility of LysoGlow84 was investigated using cultured rat brain astrocytes and the marine flatworm Macrostomum lignano. Exposure of the astrocytes for up to 6 h to micromolar concentrations of LysoGlow84 did not compromise cell viability, as demonstrated by several viability assays, but revealed a promising property of this compound for staining of cellular vesicles. Conventional fluorescence microscopy as well as confocal scanning microscopy of LysoGlow84-treated astrocytes revealed co-localization of LysoGlow84 fluorescence with that of LysoTracker® Red DND-99. LysoGlow84 stained unclear structures in Macrostomum lignano, which were identified as lysosomes by co-staining with LysoTracker. Strong fluorescence staining by LysoGlow84 was further observed around the worms' anterior gut and the female genital pore which were not counterstained by LysoTracker Red. Thus, LysoGlow84 is a new promising dye that stains lysosomes and other acidic compartments in cultured cells and in worms.

  18. Comparing milk yield, chemical properties and somatic cell count from organic and conventional mountain farming systems

    Directory of Open Access Journals (Sweden)

    Marcello Bianchi

    2010-01-01

    Full Text Available A study was undertaken to investigate the effects of farming systems (organic vs. conventional, diet (hay/concentrate vs. pasture and their interaction on milk yield, gross composition and fatty acid (FA profile of dairy cows bred in mountainous areas. For this purpose four dairy farms (two organic and two conventional were chosen in the alpine territory of Aosta Valley (NW Italy; individual milk yield was recorded daily and bulk milk samples were collected monthly from February to September 2007 to cover dietary variations. Higher levels of milk production (P<0.05 and lower milk protein amounts (P<0.01 were observed in the organic farms with respect to the conventional ones, while no significant differences were noticed in milk fat and lactose contents and in somatic cell count. Concerning fatty acids, only small differences were detected between organic and conventional milk and such differences seemed to be related mainly to the stabled period. Diet affected almost all variables studied: pasture feeding provided a significant improvement in the fatty acid composition in both organic and conventional systems leading to lower hypercholesterolemic saturated fatty acids, higher mono- and polyunsaturated fatty acids and conjugated linoleic acid amounts (P<0.001.

  19. Effect of somatic cell count on milk yield and composition of first and second lactation dairy cows

    Directory of Open Access Journals (Sweden)

    Mahmut Cinar

    2015-03-01

    Full Text Available This study was carried out to investigate the effect of somatic cell count (SCC on milk yield and milk composition in first and second lactation Holstein dairy cows. Thirty cows in first lactation and 49 cows in second lactation were used in the study. Animals were 15±9.87 days in milk. Individual milk samples were collected monthly from June 2009 to March 2010, and somatic cell counts, milk protein, milk fat, lactose and milk urea-N were determined. Four SCC groups were formed for determining effect of SCC on milk yield and composition. These groups were as follows: ≤200, 201-500, 501-999, and ≥1.000x103 cell/mL. It was observed that SCC had a high significant effect on milk yield, milk protein, milk lactose (P0.05. This study indicates that high SCC negatively affects not only milk yield but also milk composition and quality.

  20. Viable Syntax: Rethinking Minimalist Architecture

    Directory of Open Access Journals (Sweden)

    Ken Safir

    2010-03-01

    Full Text Available Hauser et al. (2002 suggest that the human language faculty emerged as a genetic innovation in the form of what is called here a ‘keystone factor’—a single, simple, formal mental capability that, interacting with the pre-existing faculties of hominid ancestors, caused a cascade of effects resulting in the language faculty in modern humans. They take Merge to be the keystone factor, but instead it is posited here that Merge is the pre-existing mechanism of thought made viable by a principle that permits relations interpretable at the interfaces to be mapped onto c-command. The simplified minimalist architecture proposed here respects the keystone factor as closely as possible, but is justified on the basis of linguistic analyses it makes available, including a relativized intervention theory applicable across Case, scope, agreement, selection and linearization, a derivation of the A/A’-distinction from Case theory, and predictions such as why in situ wh-interpretation is island-insensitive, but susceptible to intervention effects.

  1. Ultrasmall Organic Nanoparticles with Aggregation-Induced Emission and Enhanced Quantum Yield for Fluorescence Cell Imaging.

    Science.gov (United States)

    Xu, Suying; Bai, Xilin; Ma, Jingwen; Xu, Minmin; Hu, Gaofei; James, Tony D; Wang, Leyu

    2016-08-02

    The use of fluorescence probes for biomedical imaging has attracted significant attention over recent years owing to their high resolution at cellular level. The probes are available in many formats including small particle size based imaging agents which are considered to be promising candidates, due to their excellent stabilities. Yet, concerns over the potential cytotoxicity effects of inorganic luminescent particles have led to questions about their suitability for imaging applications. Exploration of alternatives inspired us to use organic fluorophores with aggregation-induced emission (AIE), prepared by functionalizing the amine group on tetraphenylethene with 3,5-bis(trifluoromethyl)phenyl isocyanate. The as-synthesized novel AIE fluorophore (TPE-F) display enhanced quantum yield and longer lifetime as compared with its counterparts (4,4',4″,4‴-(ethene-1,1,2,2-tetrayl)tetraaniline, TPE-AM). Furthermore, the TPE-F was encapsulated into small-size organic nanoparticles (NPs; dynamic light scattering size, ∼10 nm) with polysuccinimide (PSI). The biocompatibility, excellent stability, bright fluorescence, and selective cell targeting of these NPs enable the as-prepared TPE-F NPs to be suitable for specific fluorescence cell imaging.

  2. Increasing Hematopoietic Stem Cell Yield to Develop Mice with Human Immune Systems

    Directory of Open Access Journals (Sweden)

    Juan-Carlos Biancotti

    2013-01-01

    Full Text Available Hematopoietic stem cells (HSCs are unique in their capacity to give rise to all mature cells of the immune system. For years, HSC transplantation has been used for treatment of genetic and neoplastic diseases of the hematopoietic and immune systems. The sourcing of HSCs from human umbilical cord blood has salient advantages over isolation from mobilized peripheral blood. However, poor sample yield has prompted development of methodologies to expand HSCs ex vivo. Cytokines, trophic factors, and small molecules have been variously used to promote survival and proliferation of HSCs in culture, whilst strategies to lower the concentration of inhibitors in the culture media have recently been applied to promote HSC expansion. In this paper, we outline strategies to expand HSCs in vitro, and to improve engraftment and reconstitution of human immune systems in immunocompromised mice. To the extent that these “humanized” mice are representative of the endogenous human immune system, they will be invaluable tools for both basic science and translational medicine.

  3. Experimental design for the optimization of propidium monoazide treatment to quantify viable and non-viable bacteria in piggery effluents.

    Science.gov (United States)

    Desneux, Jérémy; Chemaly, Marianne; Pourcher, Anne-Marie

    2015-08-16

    Distinguishing between viable and dead bacteria in animal and urban effluents is a major challenge. Among existing methods, propidium monoazide (PMA)-qPCR is a promising way to quantify viable cells. However, its efficiency depends on the composition of the effluent, particularly on total suspended solids (TSS)) and on methodological parameters. The aim of this study was evaluate the influence of three methodological factors (concentration of PMA, incubation time and photoactivation time) on the efficiency of PMA-qPCR to quantify viable and dead cells of Listeria monocytogenes used as a microorganism model, in two piggery effluents (manure and lagoon effluent containing 20 and 0.4 TSS g.kg(-1), respectively). An experimental design strategy (Doehlert design and desirability function) was used to identify the experimental conditions to achieve optimal PMA-qPCR results. The quantification of viable cells of L. monocytogenes was mainly influenced by the concentration of PMA in the manure and by the duration of photoactivation in the lagoon effluent. Optimal values differed with the matrix: 55 μM PMA, 5 min incubation and 56 min photoactivation for manure and 20 μM PMA, 20 min incubation and 30 min photoactivation for lagoon effluent. Applied to five manure and four lagoon samples, these conditions resulted in satisfactory quantification of viable and dead cells. PMA-qPCR can be used on undiluted turbid effluent with high levels of TSS, provided preliminary tests are performed to identify the optimal conditions.

  4. Large-scale plasmid DNA processing: evidence that cell harvesting and storage methods affect yield of supercoiled plasmid DNA.

    Science.gov (United States)

    Kong, Simyee; Rock, Cassandra F; Booth, Andrew; Willoughby, Nicholas; O'Kennedy, Ronan D; Relton, Julian; Ward, John M; Hoare, Mike; Levy, M Susana

    2008-09-01

    The effect of bacterial-cell centrifugation and handling on the initial stages of plasmid processing was investigated. Escherichia coli cells containing either a 6 or 20 kb plasmid were grown in 75- and 450-litre bioreactors, and the process yield of the early recovery stages was characterized in terms of SC pDNA (supercoiled plasmid DNA) recovered. In all cases, the cells were totally recovered using either a continuous-feed, intermittent-solids-discharge, disc-stack centrifuge or a continuous-feed, batch-discharge, solid-bowl centrifuge. The cells were then either processed immediately or stored frozen. The centrifugation method considerably affected the yield of SC pDNA, and there was evidence that the intermittent discharge of cells from a centrifuge operating at high speed led to a sediment containing lysed cells and degraded pDNA. This led to estimated plasmid yield losses of up to 40% as compared with cells recovered from laboratory or solid-bowl centrifuges, where there is evidently no cell stress on discharge. By inference, the cell stress on feed to either of the continuous centrifuges studied was not implicated in product loss. Freezing of the recovered cells gives a convenient hold stage prior to further processing. In all cases, this extra freeze-thaw stage led to loss of SC pDNA, and this was in addition to the loss attributed to cell lysis during centrifugation discharge. Only average yields can be gained from pilot plant-scale studies; separate laboratory-based experiments indicated that this loss of SC pDNA is determined by the time and temperature for which the resuspended cells are held.

  5. Cell-type-specific predictive network yields novel insights into mouse embryonic stem cell self-renewal and cell fate.

    Directory of Open Access Journals (Sweden)

    Karen G Dowell

    Full Text Available Self-renewal, the ability of a stem cell to divide repeatedly while maintaining an undifferentiated state, is a defining characteristic of all stem cells. Here, we clarify the molecular foundations of mouse embryonic stem cell (mESC self-renewal by applying a proven Bayesian network machine learning approach to integrate high-throughput data for protein function discovery. By focusing on a single stem-cell system, at a specific developmental stage, within the context of well-defined biological processes known to be active in that cell type, we produce a consensus predictive network that reflects biological reality more closely than those made by prior efforts using more generalized, context-independent methods. In addition, we show how machine learning efforts may be misled if the tissue specific role of mammalian proteins is not defined in the training set and circumscribed in the evidential data. For this study, we assembled an extensive compendium of mESC data: ∼2.2 million data points, collected from 60 different studies, under 992 conditions. We then integrated these data into a consensus mESC functional relationship network focused on biological processes associated with embryonic stem cell self-renewal and cell fate determination. Computational evaluations, literature validation, and analyses of predicted functional linkages show that our results are highly accurate and biologically relevant. Our mESC network predicts many novel players involved in self-renewal and serves as the foundation for future pluripotent stem cell studies. This network can be used by stem cell researchers (at http://StemSight.org to explore hypotheses about gene function in the context of self-renewal and to prioritize genes of interest for experimental validation.

  6. Cell-type-specific predictive network yields novel insights into mouse embryonic stem cell self-renewal and cell fate.

    Science.gov (United States)

    Dowell, Karen G; Simons, Allen K; Wang, Zack Z; Yun, Kyuson; Hibbs, Matthew A

    2013-01-01

    Self-renewal, the ability of a stem cell to divide repeatedly while maintaining an undifferentiated state, is a defining characteristic of all stem cells. Here, we clarify the molecular foundations of mouse embryonic stem cell (mESC) self-renewal by applying a proven Bayesian network machine learning approach to integrate high-throughput data for protein function discovery. By focusing on a single stem-cell system, at a specific developmental stage, within the context of well-defined biological processes known to be active in that cell type, we produce a consensus predictive network that reflects biological reality more closely than those made by prior efforts using more generalized, context-independent methods. In addition, we show how machine learning efforts may be misled if the tissue specific role of mammalian proteins is not defined in the training set and circumscribed in the evidential data. For this study, we assembled an extensive compendium of mESC data: ∼2.2 million data points, collected from 60 different studies, under 992 conditions. We then integrated these data into a consensus mESC functional relationship network focused on biological processes associated with embryonic stem cell self-renewal and cell fate determination. Computational evaluations, literature validation, and analyses of predicted functional linkages show that our results are highly accurate and biologically relevant. Our mESC network predicts many novel players involved in self-renewal and serves as the foundation for future pluripotent stem cell studies. This network can be used by stem cell researchers (at http://StemSight.org) to explore hypotheses about gene function in the context of self-renewal and to prioritize genes of interest for experimental validation.

  7. Detection of viable toxigenic Vibrio cholerae and virulent Shigella ...

    African Journals Online (AJOL)

    A rapid and sensitive assay was developed for the detection of low numbers of viable Vibrio cholerae and Shigella spp. cells in environmental and drinking water samples. Water samples were filtered, and the filters were enriched in a non-selective medium. The enrichment cultures were prepared for polymerase chain ...

  8. Detection of viable toxigenic Vibrio cholerae and virulent Shigella ...

    African Journals Online (AJOL)

    DRINIE

    2003-04-02

    Apr 2, 2003 ... A rapid and sensitive assay was developed for the detection of low numbers of viable Vibrio cholerae and Shigella spp. cells in environmental and drinking water samples. Water samples were filtered, and the filters were enriched in a non-selective medium. The enrichment cultures were prepared for ...

  9. Immunocytochemical assessment of p53 protein to detect malignancy in increased cell-yield brush cytology from the biliopancreatic tree.

    Science.gov (United States)

    Villanacci, Vincenzo; Cestari, Renzo; Giulini, Stefano; Cengia, Paolo; Missale, Guido; Berenzi, Angiola; Rossi, Elisa; Bonardi, Massimo; Baiocchi, Luca; Bassotti, Gabrio

    2009-04-01

    Malignancies arising from the biliopancreatic tree are often diagnostic challenges for the gastroenterologist and the pathologist, especially when strictures without masses are present. To evaluate the diagnostic yield of p53 immunocytology for the detection of malignancies in material obtained by biliopancreatic tree brushing by means of an increased cell-yield procedure. Cytologic specimens obtained from biliary and pancreatic tree brushing in 24 patients with biliary strictures suspected for malignancy were assessed by conventional Papanicolau staining and p53 immunocytochemistry. Papanicolau staining detected 67% and p53 87% of the malignancies in the study group. p53 immunocytology displayed excellent sensitivity, specificity, and diagnostic accuracy. p53 immunocytology may represent a useful diagnostic tool in the detection of malignancies from biliary and pancreatic tree brushing, especially when using an increasing cell-yield procedure.

  10. Genetic correlation patterns between somatic cell score and protein yield in the Italian Holstein-Friesian Population

    NARCIS (Netherlands)

    Samore, A.; Groen, A.F.; Boettcher, P.; Jamrozik, J.; Canavesi, F.; Bagnato, A.

    2008-01-01

    Genetic parameters for somatic cell score (SCS) in the Italian Holstein-Friesian population were estimated addressing the pattern of genetic correlation with protein yield in different parities (first, second, and third) and on different days in milk within each parity. Three approaches for

  11. Reassortment of high-yield influenza viruses in vero cells and safety assessment as candidate vaccine strains.

    Science.gov (United States)

    Zhou, Jian; Yang, Fan; Yang, Jinghui; Ma, Lei; Cun, Yina; Song, Shaohui; Liao, Guoyang

    2017-01-02

    Vaccination is the practiced and accessible measure for preventing influenza infection. Because chicken embryos used for vaccine production have various insufficiencies, more efficient methods are needed. African green monkey kidney (Vero) cells are recommended by the World Health Organization (WHO) as a safe substitute for influenza vaccine production for humans. However, the influenza virus usually had low-yield in Vero cells, which limits the usage of Vero cellular vaccines. This study used 2 high-yield influenza viruses in Vero cells: A/Yunnan/1/2005Va (H3N2) and B/Yunnan/2/2005Va (B) as donor viruses. It used 3 wild strain viruses to reassort new adaptation viruses, including: A/Tianjin/15/2009(H1N1), A/Fujian/196/2009(H3N2), and B/Chongqing/1384/2010(B). These three new viruses could maintain the characteristic of high-yield in Vero cells. Furthermore, they could keep the immunogenic characteristics of the original wild influenza viruses. Importantly, these viruses were shown as safe in chicken embryo and guinea pigs assessment systems. These results provide an alternative method to produce influenza vaccine based on Vero cells.

  12. Biosolar cells: global artificial photosynthesis needs responsive matrices with quantum coherent kinetic control for high yield.

    Science.gov (United States)

    Purchase, R L; de Groot, H J M

    2015-06-06

    This contribution discusses why we should consider developing artificial photosynthesis with the tandem approach followed by the Dutch BioSolar Cells consortium, a current operational paradigm for a global artificial photosynthesis project. We weigh the advantages and disadvantages of a tandem converter against other approaches, including biomass. Owing to the low density of solar energy per unit area, artificial photosynthetic systems must operate at high efficiency to minimize the land (or sea) area required. In particular, tandem converters are a much better option than biomass for densely populated countries and use two photons per electron extracted from water as the raw material into chemical conversion to hydrogen, or carbon-based fuel when CO2 is also used. For the average total light sum of 40 mol m(-2) d(-1) for The Netherlands, the upper limits are many tons of hydrogen or carbon-based fuel per hectare per year. A principal challenge is to forge materials for quantitative conversion of photons to chemical products within the physical limitation of an internal potential of ca 2.9 V. When going from electric charge in the tandem to hydrogen and back to electricity, only the energy equivalent to 1.23 V can be stored in the fuel and regained. A critical step is then to learn from nature how to use the remaining difference of ca 1.7 V effectively by triple use of one overpotential for preventing recombination, kinetic stabilization of catalytic intermediates and finally generating targeted heat for the release of oxygen. Probably the only way to achieve this is by using bioinspired responsive matrices that have quantum-classical pathways for a coherent conversion of photons to fuels, similar to what has been achieved by natural selection in evolution. In appendix A for the expert, we derive a propagator that describes how catalytic reactions can proceed coherently by a convergence of time scales of quantum electron dynamics and classical nuclear dynamics. We

  13. Biosolar cells: global artificial photosynthesis needs responsive matrices with quantum coherent kinetic control for high yield

    Science.gov (United States)

    Purchase, R. L.; de Groot, H. J. M.

    2015-01-01

    This contribution discusses why we should consider developing artificial photosynthesis with the tandem approach followed by the Dutch BioSolar Cells consortium, a current operational paradigm for a global artificial photosynthesis project. We weigh the advantages and disadvantages of a tandem converter against other approaches, including biomass. Owing to the low density of solar energy per unit area, artificial photosynthetic systems must operate at high efficiency to minimize the land (or sea) area required. In particular, tandem converters are a much better option than biomass for densely populated countries and use two photons per electron extracted from water as the raw material into chemical conversion to hydrogen, or carbon-based fuel when CO2 is also used. For the average total light sum of 40 mol m−2 d−1 for The Netherlands, the upper limits are many tons of hydrogen or carbon-based fuel per hectare per year. A principal challenge is to forge materials for quantitative conversion of photons to chemical products within the physical limitation of an internal potential of ca 2.9 V. When going from electric charge in the tandem to hydrogen and back to electricity, only the energy equivalent to 1.23 V can be stored in the fuel and regained. A critical step is then to learn from nature how to use the remaining difference of ca 1.7 V effectively by triple use of one overpotential for preventing recombination, kinetic stabilization of catalytic intermediates and finally generating targeted heat for the release of oxygen. Probably the only way to achieve this is by using bioinspired responsive matrices that have quantum–classical pathways for a coherent conversion of photons to fuels, similar to what has been achieved by natural selection in evolution. In appendix A for the expert, we derive a propagator that describes how catalytic reactions can proceed coherently by a convergence of time scales of quantum electron dynamics and classical nuclear dynamics

  14. Identification of Cell Wall Synthesis Regulatory Genes Controlling Biomass Characteristics and Yield in Rice (Oryza Sativa)

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Zhaohua PEng [Mississippi State University; Ronald, Palmela [UC-Davis; Wang, Guo-Liang [The Ohio State University

    2013-04-26

    This project aims to identify the regulatory genes of rice cell wall synthesis pathways using a cell wall removal and regeneration system. We completed the gene expression profiling studies following the time course from cell wall removal to cell wall regeneration in rice suspension cells. We also completed, total proteome, nuclear subproteome and histone modification studies following the course from cell wall removal and cell wall regeneration process. A large number of differentially expressed regulatory genes and proteins were identified. Meanwhile, we generated RNAi and over-expression transgenic rice for 45 genes with at least 10 independent transgenic lines for each gene. In addition, we ordered T-DNA and transposon insertion mutants for 60 genes from Korea, Japan, and France and characterized the mutants. Overall, we have mutants and transgenic lines for over 90 genes, exceeded our proposed goal of generating mutants for 50 genes. Interesting Discoveries a) Cell wall re-synthesis in protoplasts may involve a novel cell wall synthesis mechanism. The synthesis of the primary cell wall is initiated in late cytokinesis with further modification during cell expansion. Phragmoplast plays an essential role in cell wall synthesis. It services as a scaffold for building the cell plate and formation of a new cell wall. Only one phragmoplast and one new cell wall is produced for each dividing cell. When the cell wall was removed enzymatically, we found that cell wall re-synthesis started from multiple locations simultaneously, suggesting that a novel mechanism is involved in cell wall re-synthesis. This observation raised many interesting questions, such as how the starting sites of cell wall synthesis are determined, whether phragmoplast and cell plate like structures are involved in cell wall re-synthesis, and more importantly whether the same set of enzymes and apparatus are used in cell wall re-synthesis as during cytokinesis. Given that many known cell wall

  15. Milk yield and somatic cell count during the following lactation after selective treatment of cows at dry-off.

    Science.gov (United States)

    Rajala-Schultz, Päivi J; Torres, Audrey H; Degraves, Fred J

    2011-11-01

    Selective dry cow therapy (SDCT) has received increasing attention in recent years owing to global concerns over agricultural use of antimicrobial drugs and development of antimicrobial resistance. The objective of this study was to evaluate the effect of SDCT on milk yield and somatic cell count (SCC) in dairy herds in the USA. Cows in four Ohio dairy herds were categorized into two groups (low-SCC and high-SCC) at dry-off based on their SCC and clinical mastitis (CM) history during the lactation preceding the dry-off. Low-SCC cows were randomly assigned to receive or not to receive intramammary antibiotics at dry-off. Milk yield and SCC of these cows during the following lactation were compared using linear mixed effects models, adjusting for parity, calving season, stage of lactation, previous lactation milk yield and herd. Milk yield of untreated and treated low-SCC cows at dry-off did not differ significantly during the following lactation. Overall, treated low-SCC cows had 16% lower SCC (approximately 35 000 cells/ml, P = 0·0267) than the untreated cows during the following lactation; however, the effect was variable in different herds. Moreover the impact of treatment, or the lack thereof, on milk yield varied considerably between herds. The results suggested that in some herds treating all cows at dry-off may be beneficial while in other herds leaving healthy cows without antibiotic dry cow treatment has no negative impact on milk yield or milk quality (SCC), and in fact, may be beneficial. Further studies are needed to identify characteristics of herds where treating all cows routinely at dry-off may be needed for maintaining good udder health and where switching to selective treatment of cows at dry-off would be the optimal approach to achieve best results.

  16. Pure Quantum Interpretations Are not Viable

    Science.gov (United States)

    Schmelzer, I.

    2011-02-01

    Pure interpretations of quantum theory, which throw away the classical part of the Copenhagen interpretation without adding new structure to its quantum part, are not viable. This is a consequence of a non-uniqueness result for the canonical operators.

  17. Models to Estimate Lactation Curves of Milk Yield and Somatic Cell Count in Dairy Cows at the Herd Level for the Use in Simulations and Predictive Models

    DEFF Research Database (Denmark)

    Græsbøll, Kaare; Kirkeby, Carsten Thure; Nielsen, Søren Saxmose

    2016-01-01

    of dairy herds. Data from 610 Danish Holstein herds were used to determine parity traits in milk production regarding milk yield and somatic cell count of individual cows. Parity was stratified in first, second, and third and higher for milk, and first to sixth and higher for somatic cell count. Fitting....... The correlation between lactations for milk yield and somatic cell count was 0.2–0.6 and significant on more than 95% of farms. The variation in the daily milk yield was observed to be a source of variation to the somatic cell count, and the total somatic cell count was less correlated with the milk production...... than somatic cells per milliliter. A positive correlation was found between relative levels of the total somatic cell count and the milk yield. The variation of lactation and somatic cell count curves between farms highlights the importance of a herd level approach. The one-parameter per cow model...

  18. Enhancing Protein Production Yield from Chinese Hamster Ovary Cells by CRISPR Interference.

    Science.gov (United States)

    Shen, Chih-Che; Sung, Li-Yu; Lin, Shih-Yeh; Lin, Mei-Wei; Hu, Yu-Chen

    2017-08-18

    Chinese hamster ovary (CHO) cells are an important host for biopharmaceutical production. Generation of stable CHO cells typically requires cointegration of dhfr and a foreign gene into chromosomes and subsequent methotrexate (MTX) selection for coamplification of dhfr and foreign gene. CRISPR interference (CRISPRi) is an emerging system that effectively suppresses gene transcription through the coordination of dCas9 protein and guide RNA (gRNA). However, CRISPRi has yet to be exploited in CHO cells. Here we constructed vectors expressing the functional CRISPRi system and proved effective CRISPRi-mediated suppression of dhfr transcription in CHO cells. We next generated stable CHO cell clones coexpressing DHFR, the model protein (EGFP), dCas9 and gRNA targeting dhfr. Combined with MTX selection, CRISPRi-mediated repression of dhfr imparted extra selective pressure to force CHO cells to coamplify more copies of dhfr and egfp genes. Compared with the traditional method relying on MTX selection (up to 250 nM), the CRISPRi approach increased the dhfr copy number ∼3-fold, egfp copy number ∼3.6-fold and enhanced the EGFP expression ∼3.8-fold, without impeding the cell growth. Furthermore, we exploited the CRISPRi approach to enhance the productivity of granulocyte colony stimulating factor (G-CSF) ∼2.3-fold. Our data demonstrate, for the first time, the application of CRISPRi in CHO cells to enhance recombinant protein production and may pave a new avenue to CHO cell engineering.

  19. Charge yield potential of indoor-operated solar cells incorporated into Product Integrated Photovoltaic (PIPV)

    NARCIS (Netherlands)

    Reich, N.H.; van Sark, W.G.J.H.M.; Turkenburg, W.C.

    2010-01-01

    Solar cell performance parameters (open circuit voltage, short circuit current, fill factor and efficiency) are derived for different solar cell types for the irradiance range 0.1–1000 W/m2. Also it is demonstrated how spectral mismatch factors for indoor lighting conditions are calculated. The

  20. Cell surface engineering of Bacillus subtilis improves production yields of heterologously expressed α-amylases

    NARCIS (Netherlands)

    Cao, Haojie; van Heel, Auke J; Ahmed, Hifza; Mols, Maarten; Kuipers, Oscar P

    2017-01-01

    BACKGROUND: Bacillus subtilis is widely used as a cell factory for numerous heterologous proteins of commercial value and medical interest. To explore the possibility of further enhancing the secretion potential of this model bacterium, a library of engineered strains with modified cell surface

  1. Energy Yield Determination of Concentrator Solar Cells using Laboratory Measurements: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Geisz, John F.; Garcia, Ivan; McMahon, William E.; Steiner, Myles A.; Ochoa, Mario; France, Ryan M.; Habte, Aron; Friedman, Daniel J.

    2015-09-14

    The annual energy conversion efficiency is calculated for a four junction inverted metamorphic solar cell that has been completely characterized in the laboratory at room temperature using measurements fit to a comprehensive optoelectronic model of the multijunction solar cells. A simple model of the temperature dependence is used to predict the performance of the solar cell under varying temperature and spectra characteristic of Golden, CO for an entire year. The annual energy conversion efficiency is calculated by integrating the predicted cell performance over the entire year. The effects of geometric concentration, CPV system thermal characteristics, and luminescent coupling are highlighted. temperature and spectra characteristic of Golden, CO for an entire year. The annual energy conversion efficiency is calculated by integrating the predicted cell performance over the entire year. The effects of geometric concentration, CPV system thermal characteristics, and luminescent coupling are highlighted.

  2. High yield production of influenza virus in Madin Darby canine kidney (MDCK cells with stable knockdown of IRF7.

    Directory of Open Access Journals (Sweden)

    Itsuki Hamamoto

    Full Text Available Influenza is a serious public health problem that causes a contagious respiratory disease. Vaccination is the most effective strategy to reduce transmission and prevent influenza. In recent years, cell-based vaccines have been developed with continuous cell lines such as Madin-Darby canine kidney (MDCK and Vero. However, wild-type influenza and egg-based vaccine seed viruses will not grow efficiently in these cell lines. Therefore, improvement of virus growth is strongly required for development of vaccine seed viruses and cell-based influenza vaccine production. The aim of our research is to develop novel MDCK cells supporting highly efficient propagation of influenza virus in order to expand the capacity of vaccine production. In this study, we screened a human siRNA library that involves 78 target molecules relating to three major type I interferon (IFN pathways to identify genes that when knocked down by siRNA lead to enhanced production of influenza virus A/Puerto Rico/8/1934 in A549 cells. The siRNAs targeting 23 candidate genes were selected to undergo a second screening pass in MDCK cells. We examined the effects of knockdown of target genes on the viral production using newly designed siRNAs based on sequence analyses. Knockdown of the expression of a canine gene corresponding to human IRF7 by siRNA increased the efficiency of viral production in MDCK cells through an unknown process that includes the mechanisms other than inhibition of IFN-α/β induction. Furthermore, the viral yield greatly increased in MDCK cells stably transduced with the lentiviral vector for expression of short hairpin RNA against IRF7 compared with that in control MDCK cells. Therefore, we propose that modified MDCK cells with lower expression level of IRF7 could be useful not only for increasing the capacity of vaccine production but also facilitating the process of seed virus isolation from clinical specimens for manufacturing of vaccines.

  3. Slow light enhanced singlet exciton fission solar cells with a 126% yield of electrons per photon

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, Nicholas J.; Congreve, Daniel N.; Baldo, Marc A., E-mail: vmenon@qc.cuny.edu, E-mail: baldo@mit.edu [Energy Frontier Research Center for Excitonics, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 (United States); Goldberg, David; Menon, Vinod M., E-mail: vmenon@qc.cuny.edu, E-mail: baldo@mit.edu [Department of Physics, Queens College and Graduate Center, The City University of New York, Flushing, New York 11367 (United States)

    2013-12-23

    Singlet exciton fission generates two triplet excitons per absorbed photon. It promises to increase the power extracted from sunlight without increasing the number of photovoltaic junctions in a solar cell. We demonstrate solar cells with an external quantum efficiency of 126% by enhancing absorption in thin films of the singlet exciton fission material pentacene. The device structure exploits the long photon dwell time at the band edge of a distributed Bragg reflector to achieve enhancement over a broad range of angles. Measuring the reflected light from the solar cell establishes a lower bound of 137% for the internal quantum efficiency.

  4. Grazing of particle-associated bacteria-an elimination of the non-viable fraction.

    Science.gov (United States)

    Gonsalves, Maria-Judith; Fernandes, Sheryl Oliveira; Priya, Madasamy Lakshmi; LokaBharathi, Ponnapakkam Adikesavan

    Quantification of bacteria being grazed by microzooplankton is gaining importance since they serve as energy subsidies for higher trophic levels which consequently influence fish production. Hence, grazing pressure on viable and non-viable fraction of free and particle-associated bacteria in a tropical estuary controlled mainly by protist grazers was estimated using the seawater dilution technique. In vitro incubations over a period of 42h showed that at the end of 24h, growth coefficient (k) of particle-associated bacteria was 9 times higher at 0.546 than that of free forms. Further, 'k' value of viable cells on particles was double that of free forms at 0.016 and 0.007, respectively. While bacteria associated with particles were grazed (coefficient of removal (g)=0.564), the free forms were relatively less grazed indicating that particle-associated bacteria were exposed to grazers in these waters. Among the viable and non-viable forms, 'g' of non-viable fraction (particle-associated bacteria=0.615, Free=0.0086) was much greater than the viable fraction (particle-associated bacteria=0.056, Free=0.068). Thus, grazing on viable cells was relatively low in both the free and attached states. These observations suggest that non-viable forms of particle-associated bacteria were more prone to grazing and were weeded out leaving the viable cells to replenish the bacterial standing stock. Particle colonization could thus be a temporary refuge for the "persistent variants" where the viable fraction multiply and release their progeny. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  5. EFFECT OF DIFFERENT STAPHYLOCOCCUS AUREUS MASTITIS VACCINES ON THE MILK YIELD, FAT, PROTEIN AND SOMATIC CELL COUNT IN BUFFALOES

    Directory of Open Access Journals (Sweden)

    A. SHAKOOR, M. ATHAR, G. MUHAMMAD, S. U. RAHMAN1, A. A. BUTT2, I. HUSSAIN1 AND R. AHMAD3

    2006-04-01

    Full Text Available Four Staph. aureus mastitis vaccines were evaluated in five different groups of non-mastitic healthy pregnant buffaloes. These vaccines [live attenuated (Group B1, simple bacterin (Group B2, dextran sulphate adjuvanted (Group B3 and oil adjuvanted (Group B4] were administered to 20 non-mastitic healthy pregnant buffaloes, while the group 5 was kept as unvaccinated control. Each vaccine was administered twice @ 5 ml IM at 60 and 30 days pre-partum. The effect of these vaccines on milk yield, fat, protein, and somatic cell count was studied till 4th month post partum. There was a significant difference in the milk yield, fat and protein percentage between the vaccinated and non-vaccinated groups, while difference of these parameters among the vaccinated groups of buffaloes was non significant. All the vaccines reduced the somatic cell count significantly as compared to control group. Linear somatic cell count scouring method indicated the highest milk loss/lactation in the control group (1086 litres, followed by bacterin (703 litres, dextran sulphate (549 litres, live attenuated (526 litres and oil based (577 litres vaccines.

  6. Optimizing culture medium composition to improve oligodendrocyte progenitor cell yields in vitro from subventricular zone-derived neural progenitor cell neurospheres.

    Science.gov (United States)

    Franco, Paula G; Pasquini, Juana M; Silvestroff, Lucas

    2015-01-01

    Neural Stem and Progenitor Cells (NSC/NPC) are gathering tangible recognition for their uses in cell therapy and cell replacement therapies for human disease, as well as a model system to continue research on overall neural developmental processes in vitro. The Subventricular Zone is one of the largest NSC/NPC niches in the developing mammalian Central Nervous System, and persists through to adulthood. Oligodendrocyte progenitor cell (OPC) enriched cultures are usefull tools for in vitro studies as well as for cell replacement therapies for treating demyelination diseases. We used Subventricular Zone-derived NSC/NPC primary cultures from newborn mice and compared the effects of different growth factor combinations on cell proliferation and OPC yield. The Platelet Derived Growth Factor-AA and BB homodimers had a positive and significant impact on OPC generation. Furthermore, heparin addition to the culture media contributed to further increase overall culture yields. The OPC generated by this protocol were able to mature into Myelin Basic Protein-expressing cells and to interact with neurons in an in vitro co-culture system. As a whole, we describe an optimized in vitro method for increasing OPC.

  7. Using Generic Examples to Make Viable Arguments

    Science.gov (United States)

    Adams, Anne E.; Ely, Rob; Yopp, David

    2017-01-01

    The twenty-first century has seen an increased call to train students to craft mathematical arguments. The third of the Common Core's (CCSS) Standards for Mathematical Practice (SMP 3) (CCSSI 2010) calls for all mathematically proficient students to "construct viable arguments" to support the truth of their ideas and to "critique…

  8. Improved poliovirus d-antigen yields by application of different Vero cell cultivation methods

    NARCIS (Netherlands)

    Thomassen, Y.E.; Rubingh, O.; Wijffels, R.H.; Pol, van der L.A.

    2014-01-01

    Vero cells were grown adherent to microcarriers (Cytodex 1; 3 g L-1) using animal component free media in stirred-tank type bioreactors. Different strategies for media refreshment, daily media replacement (semi-batch), continuous media replacement (perfusion) and recirculation of media, were

  9. Optimisation of the Factor VIII yield in mammalian cell cultures by reducing the membrane bound fraction

    DEFF Research Database (Denmark)

    Kolind, Mille Petersen; Nørby, Peder Lisby; Berchtold, Martin Werner

    2011-01-01

    of active membrane bound rFVIII to the culture medium. Moreover, the attachment of rFVIII to cell membranes of un-transfected HEK293 cells was studied in the presence of compounds that competes for interactions between rFVIII and PS. Competitive assays between iodinated rFVIII (¹²5I-rFVIII) and annexin V......In vivo, clotting Factor VIII (FVIII) circulates in plasma bound to von Willebrand factor (vWF), and the vWF:FVIII complex prevents binding of FVIII to phosphatidylserine (PS). Activation of FVIII by thrombin releases FVIII from vWF, and subsequently FVIII binds to PS exposed on activated platelets...... of the production cells. Recently, we showed that as much as 90% of secreted rFVIII is bound to transiently transfected production cells during serum free conditions. In this study, we investigated the effect of including vWF in the serum free medium, and demonstrate that addition of vWF results in release...

  10. Lost in transition: start-up of glycolysis yields subpopulations of nongrowing cells.

    NARCIS (Netherlands)

    van Heerden, J.H.; Wortel, M.T.; Bruggeman, F.J.; Heijnen, J.J.; Bollen, Y.J.M.; Planque, R.; Hulshof, J.; O'Toole, T.G.; Wahl, S.A.; Teusink, B.

    2014-01-01

    Cells need to adapt to dynamic environments. Yeast that fail to cope with dynamic changes in the abundance of glucose can undergo growth arrest. We show that this failure is caused by imbalanced reactions in glycolysis, the essential pathway in energy metabolism in most organisms. The imbalance

  11. Removal of viable bacteria and endotoxins by Electro Deionization (EDI).

    Science.gov (United States)

    Harada, Norimitsu; Otomo, Teruo; Watabe, Tomoichi; Ase, Tomonobu; Takemura, Takuto; Sato, Toshio

    2011-09-01

    Viable bacteria and endotoxins in water sometimes cause problems for human health. Endotoxins are major components of the outer cell wall of gram-negative bacteria (lipopolysaccharides). In medical procedures, especially haemodialysis (HD) and related therapies (haemodiafiltration (HDF), haemofiltration (HF)), endotoxins in the water for haemodialysis can permeate through the haemodialysis membrane and cause microinflammation or various haemodialysis-related illnesses. To decrease such a biological risk, RO and UF membranes are generally used. Also, hot water disinfection or the chemical disinfection is regularly executed to kill bacteria which produce endotoxins. However, simple treatment methods and equipment may be able to decrease the biological risk more efficiently. In our experiments, we confirmed that viable bacteria and endotoxins were removed by Electro Deionization (EDI) technology and also clarified the desorption mechanisms.

  12. Comparing The Energy Yield of (III-V) Multi-Junction Cells With Different Numbers Of Sub-Cells

    Science.gov (United States)

    Lumb, M. P.; Dobbin, A. L.; Bushnell, D. B.; Lee, K. H.; Tibbits, T. N. D.

    2010-10-01

    We examine the performance of III-V multi-junction solar cells with different numbers of cells under diverse spectral conditions. Using both a detailed balance model and a more realistic analytical charge-transport model, we simulate the energy harvesting properties of a range of cell designs with calculated spectra over 1 year. The spectra are generated using the SMARTS2 model with real atmospheric data for three locations with diverse spectral characteristics. The work underlines the importance of matching multi-junction solar cells to their target spectrum in order to maximize their energy harvest potential.

  13. Somatic cell count and milk yield on physicochemical components of milk from free-stall housed cows

    Directory of Open Access Journals (Sweden)

    José Antônio Freitas

    2017-05-01

    Full Text Available Measurement and evaluation of the factors that may influence milk composition has been shown to determine the actual gains in milk quality and quantity. The study was conducted to evaluate the influence of daily milk yield on the physicochemical composition of milk and analyze the correlations between milk yield and somatic cell count (SCC on the fat, protein, lactose, total solids and nitrogen urea contents in Holstein cows’ milk. A total of 3,544 data collected from 467 cows were evaluated. In the analysis, the animals were divided into groups – I (5-15 kg milk day-1, II (15-30 kg milk day-1 and III (above 30 kg milk day -1 – based on their daily production. Animals of Group I had higher SCC (P 0.05 between SCC and fat, protein and total solids contents was observed. A lower milk yield is significantly associated with a higher SCC, which, besides hindering the quality of milk, compromises the cow's udder health.

  14. Controlled-rate freezer cryopreservation of highly concentrated peripheral blood mononuclear cells results in higher cell yields and superior autologous T-cell stimulation for dendritic cell-based immunotherapy.

    Science.gov (United States)

    Buhl, Timo; Legler, Tobias J; Rosenberger, Albert; Schardt, Anke; Schön, Michael P; Haenssle, Holger A

    2012-11-01

    Availability of large quantities of functionally effective dendritic cells (DC) represents one of the major challenges for immunotherapeutic trials against infectious or malignant diseases. Low numbers or insufficient T-cell activation of DC may result in premature termination of treatment and unsatisfying immune responses in clinical trials. Based on the notion that cryopreservation of monocytes is superior to cryopreservation of immature or mature DC in terms of resulting DC quantity and immuno-stimulatory capacity, we aimed to establish an optimized protocol for the cryopreservation of highly concentrated peripheral blood mononuclear cells (PBMC) for DC-based immunotherapy. Cryopreserved cell preparations were analyzed regarding quantitative recovery, viability, phenotype, and functional properties. In contrast to standard isopropyl alcohol (IPA) freezing, PBMC cryopreservation in an automated controlled-rate freezer (CRF) with subsequent thawing and differentiation resulted in significantly higher cell yields of immature and mature DC. Immature DC yields and total protein content after using CRF were comparable with results obtained with freshly prepared PBMC and exceeded results of standard IPA freezing by approximately 50 %. While differentiation markers, allogeneic T-cell stimulation, viability, and cytokine profiles were similar to DC from standard freezing procedures, DC generated from CRF-cryopreserved PBMC induced a significantly higher antigen-specific IFN-γ release from autologous effector T cells. In summary, automated controlled-rate freezing of highly concentrated PBMC represents an improved method for increasing DC yields and autologous T-cell stimulation.

  15. Combinatorial genome and protein engineering yields monoclonal antibodies with hypergalactosylation from CHO cells.

    Science.gov (United States)

    Chung, Cheng-Yu; Wang, Qiong; Yang, Shuang; Ponce, Sean A; Kirsch, Brian J; Zhang, Hui; Betenbaugh, Michael J

    2017-12-01

    One of the key quality attributes of monoclonal antibodies is the glycan pattern and distribution. Two terminal galactose residues typically represent a small fraction of the total glycans from antibodies. However, antibodies with defined glycosylation properties including enhanced galactosylation have been shown to exhibit altered properties for these important biomedical modalities. In this study, the disruption of two α-2,3 sialyltransferases (ST3GAL4 and ST3GAL6) from Chinese Hamster Ovary (CHO) cells was combined with protein engineering of the Fc region to generate an IgG containing 80% bigalactosylated and fucosylated (G2F) glycoforms. Expression of the same single amino acid mutant (F241A) IgG in CHO cells with a triple gene knockout of fucosyltransferase (FUT8) plus ST3GAL4 and ST3GAL6 lowered the galactosylation glycoprofile to 65% bigalactosylated G2 glycans. However, overexpression of IgGs with four amino acid substitutions recovered the G2 glycoform composition approximately 80%. Combining genome and protein engineering in CHO cells will provide a new antibody production platform that enables biotechnologists to generate glycoforms standards for specific biomedical and biotechnology applications. © 2017 Wiley Periodicals, Inc.

  16. Yields and chondrogenic potential of primary synovial mesenchymal stem cells are comparable between rheumatoid arthritis and osteoarthritis patients.

    Science.gov (United States)

    Kohno, Yuji; Mizuno, Mitsuru; Ozeki, Nobutake; Katano, Hisako; Komori, Keiichiro; Fujii, Shizuka; Otabe, Koji; Horie, Masafumi; Koga, Hideyuki; Tsuji, Kunikazu; Matsumoto, Mikio; Kaneko, Haruka; Takazawa, Yuji; Muneta, Takeshi; Sekiya, Ichiro

    2017-05-16

    Mesenchymal stem cells derived from the synovial membrane (synovial MSCs) are a candidate cell source for regenerative medicine of cartilage and menisci due to their high chondrogenic ability. Regenerative medicine can be expected for RA patients with the inflammation well-controlled as well as OA patients and transplantation of synovial MSCs would also be a possible therapeutic treatment. Some properties of synovial MSCs vary dependent on the diseases patients have, and whether or not the pathological condition of RA affects the chondrogenesis of synovial MSCs remains controversial. The purpose of this study was to compare the properties of primary synovial MSCs between RA and OA patients. Human synovial tissue was harvested during total knee arthroplasty from the knee joints of eight patients with RA and OA respectively. Synovial nucleated cells were cultured for 14 days. Total cell yields, surface markers, and differentiation potentials were analyzed for primary synovial MSCs. Nucleated cell number per 1 mg synovium was 8.4 ± 3.9 thousand in RA and 8.0 ± 0.9 thousand in OA. Total cell number after 14-day culture/1 mg synovium was 0.7 ± 0.4 million in RA and 0.5 ± 0.3 million in OA, showing no significant difference between in RA and OA. Cells after 14-day culture were mostly positive for CD44, CD73, CD90, CD105, negative for CD45 both in RA and OA. There was no significant difference for the cartilage pellet weight and sGAG content per pellet between in RA and OA. Both oil red O-positive colony rate and alizarin red-positive colony rate were similar in RA and OA. Yields, surface markers and chondrogenic potential of primary synovial MSCs in RA were comparable to those in OA. Synovium derived from RA patients can be the cell source of MSCs for cartilage and meniscus regeneration.

  17. Regeneration of viable oil palm plants from protoplasts by optimizing media components, growth regulators and cultivation procedures.

    Science.gov (United States)

    Masani, Mat Yunus Abdul; Noll, Gundula; Parveez, Ghulam Kadir Ahmad; Sambanthamurthi, Ravigadevi; Prüfer, Dirk

    2013-09-01

    Oil palm protoplasts are suitable as a starting material for the production of oil palm plants with new traits using approaches such as somatic hybridization, but attempts to regenerate viable plants from protoplasts have failed thus far. Here we demonstrate, for the first time, the regeneration of viable plants from protoplasts isolated from cell suspension cultures. We achieved a protoplast yield of 1.14×10(6) per gram fresh weight with a viability of 82% by incubating the callus in a digestion solution comprising 2% cellulase, 1% pectinase, 0.5% cellulase onuzuka R10, 0.1% pectolyase Y23, 3% KCl, 0.5% CaCl2 and 3.6% mannitol. The regeneration of protoplasts into viable plants required media optimization, the inclusion of plant growth regulators and the correct culture technique. Microcalli derived from protoplasts were obtained by establishing agarose bead cultures using Y3A medium supplemented with 10μM naphthalene acetic acid, 2μM 2,4-dichlorophenoxyacetic acid, 2μM indole-3-butyric acid, 2μM gibberellic acid and 2μM 2-γ-dimethylallylaminopurine. Small plantlets were regenerated from microcalli by somatic embryogenesis after successive subculturing steps in medium with limiting amounts of growth regulators supplemented with 200mg/l ascorbic acid. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  18. Increasing cell biomass in Saccharomyces cerevisiae increases recombinant protein yield: the use of a respiratory strain as a microbial cell factory

    Directory of Open Access Journals (Sweden)

    Hedfalk Kristina

    2010-06-01

    Full Text Available Abstract Background Recombinant protein production is universally employed as a solution to obtain the milligram to gram quantities of a given protein required for applications as diverse as structural genomics and biopharmaceutical manufacture. Yeast is a well-established recombinant host cell for these purposes. In this study we wanted to investigate whether our respiratory Saccharomyces cerevisiae strain, TM6*, could be used to enhance the productivity of recombinant proteins over that obtained from corresponding wild type, respiro-fermentative strains when cultured under the same laboratory conditions. Results Here we demonstrate at least a doubling in productivity over wild-type strains for three recombinant membrane proteins and one recombinant soluble protein produced in TM6* cells. In all cases, this was attributed to the improved biomass properties of the strain. The yield profile across the growth curve was also more stable than in a wild-type strain, and was not further improved by lowering culture temperatures. This has the added benefit that improved yields can be attained rapidly at the yeast's optimal growth conditions. Importantly, improved productivity could not be reproduced in wild-type strains by culturing them under glucose fed-batch conditions: despite having achieved very similar biomass yields to those achieved by TM6* cultures, the total volumetric yields were not concomitantly increased. Furthermore, the productivity of TM6* was unaffected by growing cultures in the presence of ethanol. These findings support the unique properties of TM6* as a microbial cell factory. Conclusions The accumulation of biomass in yeast cell factories is not necessarily correlated with a proportional increase in the functional yield of the recombinant protein being produced. The respiratory S. cerevisiae strain reported here is therefore a useful addition to the matrix of production hosts currently available as its improved biomass

  19. Monotone viable trajectories for functional differential inclusions

    Science.gov (United States)

    Haddad, Georges

    This paper is a study on functional differential inclusions with memory which represent the multivalued version of retarded functional differential equations. The main result gives a necessary and sufficient equations. The main result gives a necessary and sufficient condition ensuring the existence of viable trajectories; that means trajectories remaining in a given nonempty closed convex set defined by given constraints the system must satisfy to be viable. Some motivations for this paper can be found in control theory where F( t, φ) = { f( t, φ, u)} uɛU is the set of possible velocities of the system at time t, depending on the past history represented by the function φ and on a control u ranging over a set U of controls. Other motivations can be found in planning procedures in microeconomics and in biological evolutions where problems with memory do effectively appear in a multivalued version. All these models require viability constraints represented by a closed convex set.

  20. Structure-based redesign of lysostaphin yields potent antistaphylococcal enzymes that evade immune cell surveillance

    Directory of Open Access Journals (Sweden)

    Kristina Blazanovic

    Full Text Available Staphylococcus aureus infections exert a tremendous burden on the health-care system, and the threat of drug-resistant strains continues to grow. The bacteriolytic enzyme lysostaphin is a potent antistaphylococcal agent with proven efficacy against both drug-sensitive and drug-resistant strains; however, the enzyme's own bacterial origins cause undesirable immunogenicity and pose a barrier to clinical translation. Here, we deimmunized lysostaphin using a computationally guided process that optimizes sets of mutations to delete immunogenic T cell epitopes without disrupting protein function. In vitro analyses showed the methods to be both efficient and effective, producing seven different deimmunized designs exhibiting high function and reduced immunogenic potential. Two deimmunized candidates elicited greatly suppressed proliferative responses in splenocytes from humanized mice, while at the same time the variants maintained wild-type efficacy in a staphylococcal pneumonia model. Overall, the deimmunized enzymes represent promising leads in the battle against S. aureus.

  1. Isolation of viable human hepatic progenitors from adult livers is possible even after 48 hours of cold ischemia.

    Science.gov (United States)

    Aupet, Sophie; Simoné, Gael; Heyd, Bruno; Bachellier, Philippe; Vidal, Isabelle; Richert, Lysiane; Martin, Hélène

    2013-07-01

    Liver transplantation, utilized routinely for end-stage liver disease, has been constrained by the paucity of organ donors, and is being complemented by alternative strategies such as liver cell transplantation. One of the most promising forms of liver cell transplantation is hepatic stem cell therapies, as the number of human hepatic stem cells (hHpSCs) and other early hepatic progenitor cells (HPCs) are sufficient to provide treatment for multiple patients from a single liver source. In the present study, human adult livers were exposed to cold ischemia and then processed after numbers, albeit somewhat lower, were obtained from those exposed to 48 h of cold ischemia. The yields are similar to those reported from livers with minimal exposure to ischemia. When cultured on plastic dishes and in Kubota's Medium, a serum-free medium designed for early lineage stage HPCs, colonies of rapidly expanding cells formed. They were confirmed to be probable hHpSCs by their ability to survive and expand on plastic and in Kubota's Medium for months, by co-expression of EpCAM and neural cell adhesion molecule, minimal if any albumin expression, with EpCAM found throughout the cells, and no expression of alpha-fetoprotein. The yields of viable EpCAM(+) cells were surprisingly large, and the numbers from a single donor liver are sufficient to treat approximately 50-100 patients given the numbers of EpCAM(+) cells currently used in hepatic stem cell therapies. Thus, cold ischemic livers for up to 48 h are a new source of cells that might be used for liver cell therapies.

  2. Perovskite solar cells with CuSCN hole extraction layers yield stabilized efficiencies greater than 20%

    Science.gov (United States)

    Arora, Neha; Dar, M. Ibrahim; Hinderhofer, Alexander; Pellet, Norman; Schreiber, Frank; Zakeeruddin, Shaik Mohammed; Grätzel, Michael

    2017-11-01

    Perovskite solar cells (PSCs) with efficiencies greater than 20% have been realized only with expensive organic hole-transporting materials. We demonstrate PSCs that achieve stabilized efficiencies exceeding 20% with copper(I) thiocyanate (CuSCN) as the hole extraction layer. A fast solvent removal method enabled the creation of compact, highly conformal CuSCN layers that facilitate rapid carrier extraction and collection. The PSCs showed high thermal stability under long-term heating, although their operational stability was poor. This instability originated from potential-induced degradation of the CuSCN/Au contact. The addition of a conductive reduced graphene oxide spacer layer between CuSCN and gold allowed PSCs to retain >95% of their initial efficiency after aging at a maximum power point for 1000 hours under full solar intensity at 60°C. Under both continuous full-sun illumination and thermal stress, CuSCN-based devices surpassed the stability of spiro-OMeTAD–based PSCs.

  3. Evaluation of low-cost cathode catalysts for high yield biohydrogen production in microbial electrolysis cell.

    Science.gov (United States)

    Wang, L; Chen, Y; Ye, Y; Lu, B; Zhu, S; Shen, S

    2011-01-01

    As an ideal fuel due to the advantages of no pollution, high combustion heat and abundant sources, hydrogen gas can be produced from organic matter through the electrohydrogenesis process in microbial electrolysis cells. But in many MECs, platinum is often used as catalyst, which limits the practical applications of MECs. To reduce the cost of the MECs, Ni-based alloy cathodes were developed by electrodepositing. In this paper hydrogen production using Ni-W-P cathode was studied for the first time in a single-chamber membrane-free MEC. At an applied voltage of 0.9 V, MECs with Ni-W-P cathodes obtained a hydrogen production rate of 1.09 m3/m3/day with an cathodic hydrogen recovery of 74%, a Coulombic efficiency of 56% and an electrical energy efficiency relative to electrical input of 139%, which was the best result of reports in this study. The Ni-W-P cathode demonstrated a better electrocatalytic activity than the Ni-Ce-P cathode and achieved a comparable performance to the Pt cathode in terms of hydrogen production rate, Coulombic efficiency, cathodic hydrogen recovery and electrical energy efficiency at 0.9 V.

  4. Improvement in Saccharification Yield of Mixed Rumen Enzymes by Identification of Recalcitrant Cell Wall Constituents Using Enzyme Fingerprinting

    Directory of Open Access Journals (Sweden)

    Ajay Badhan

    2015-01-01

    Full Text Available Identification of recalcitrant factors that limit digestion of forages and the development of enzymatic approaches that improve hydrolysis could play a key role in improving the efficiency of meat and milk production in ruminants. Enzyme fingerprinting of barley silage fed to heifers and total tract indigestible fibre residue (TIFR collected from feces was used to identify cell wall components resistant to total tract digestion. Enzyme fingerprinting results identified acetyl xylan esterases as key to the enhanced ruminal digestion. FTIR analysis also suggested cross-link cell wall polymers as principal components of indigested fiber residues in feces. Based on structural information from enzymatic fingerprinting and FTIR, enzyme pretreatment to enhance glucose yield from barley straw and alfalfa hay upon exposure to mixed rumen-enzymes was developed. Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design. Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes. The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

  5. Improvement in Saccharification Yield of Mixed Rumen Enzymes by Identification of Recalcitrant Cell Wall Constituents Using Enzyme Fingerprinting.

    Science.gov (United States)

    Badhan, Ajay; Wang, Yu-Xi; Gruninger, Robert; Patton, Donald; Powlowski, Justin; Tsang, Adrian; McAllister, Tim A

    2015-01-01

    Identification of recalcitrant factors that limit digestion of forages and the development of enzymatic approaches that improve hydrolysis could play a key role in improving the efficiency of meat and milk production in ruminants. Enzyme fingerprinting of barley silage fed to heifers and total tract indigestible fibre residue (TIFR) collected from feces was used to identify cell wall components resistant to total tract digestion. Enzyme fingerprinting results identified acetyl xylan esterases as key to the enhanced ruminal digestion. FTIR analysis also suggested cross-link cell wall polymers as principal components of indigested fiber residues in feces. Based on structural information from enzymatic fingerprinting and FTIR, enzyme pretreatment to enhance glucose yield from barley straw and alfalfa hay upon exposure to mixed rumen-enzymes was developed. Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design. Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes. The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

  6. Air-spore in Cartagena, Spain: viable and non-viable sampling methods.

    Science.gov (United States)

    Elvira-Rendueles, Belen; Moreno, Jose; Garcia-Sanchez, Antonio; Vergara, Nuria; Martinez-Garcia, Maria Jose; Moreno-Grau, Stella

    2013-01-01

    In the presented study the airborne fungal spores of the semiarid city of Cartagena, Spain, are identified and quantified by means of viable or non-viable sampling methods. Airborne fungal samples were collected simultaneously using a filtration method and a pollen and particle sampler based on the Hirst methodology. This information is very useful for elucidating geographical patterns of hay fever and asthma. The qualitative results showed that when the non-viable methodology was employed, Cladosporium, Ustilago, and Alternaria were the most abundant spores identified in the atmosphere of Cartagena, while the viable methodology showed that the most abundant taxa were: Cladosporium, Penicillium, Aspergillus and Alternaria. The quantitative results of airborne fungal spores identified by the Hirst-type air sampler (non-viable method), showed that Deuteromycetes represented 74% of total annual spore counts, Cladosporium being the major component of the fungal spectrum (62.2%), followed by Alternaria (5.3%), and Stemphylium (1.3%). The Basidiomycetes group represented 18.9% of total annual spore counts, Ustilago (7.1%) being the most representative taxon of this group and the second most abundant spore type. Ascomycetes accounted for 6.9%, Nectria (2.3%) being the principal taxon. Oomycetes (0.2%) and Zygomycestes and Myxomycestes (0.06%) were scarce. The prevailing species define our bioaerosol as typical of dry air. The viable methodology was better at identifying small hyaline spores and allowed for the discrimination of the genus of some spore types. However, non-viable methods revealed the richness of fungal types present in the bioaerosol. Thus, the use of both methodologies provides a more comprehensive characterization of the spore profile.

  7. Parejas viables que perduran en el tiempo

    OpenAIRE

    Juan José Cuervo Rodríguez

    2013-01-01

    El presente artículo científico presenta resultados del proceso llevado a cabo en el proyecto de investigación docente "Mecanismos de autorregulación en parejas viables que perduran en el tiempo". Se soporta en una mirada compleja de la psicología basada en una epistemología de la construcción. En el ámbito metodológico, se inscribe en los estudios de terapia familiar desde una perspectiva de la comunicación humana como un todo integrado. Participaron nueve parejas. Los criterios de inclusión...

  8. A rapid biosensor for viable B. anthracis spores.

    Science.gov (United States)

    Baeumner, Antje J; Leonard, Barbara; McElwee, John; Montagna, Richard A

    2004-09-01

    A simple membrane-strip-based biosensor assay has been combined with a nucleic acid sequence-based amplification (NASBA) reaction for rapid (4 h) detection of a small number (ten) of viable B. anthracis spores. The biosensor is based on identification of a unique mRNA sequence from one of the anthrax toxin genes, the protective antigen ( pag), encoded on the toxin plasmid, pXO1, and thus provides high specificity toward B. anthracis. Previously, the anthrax toxins activator ( atxA) mRNA had been used in our laboratory for the development of a biosensor for the detection of a single B. anthracis spore within 12 h. Changing the target sequence to the pag mRNA provided the ability to shorten the overall assay time significantly. The vaccine strain of B. anthracis (Sterne strain) was used in all experiments. A 500-microL sample containing as few as ten spores was mixed with 500 microL growth medium and incubated for 30 min for spore germination and mRNA production. Thus, only spores that are viable were detected. Subsequently, RNA was extracted from lysed cells, selectively amplified using NASBA, and rapidly identified by the biosensor. While the biosensor assay requires only 15 min assay time, the overall process takes 4 h for detection of ten viable B. anthracis spores, and is shortened significantly if more spores are present. The biosensor is based on an oligonucleotide sandwich-hybridization assay format. It uses a membrane flow-through system with an immobilized DNA probe that hybridizes with the target sequence. Signal amplification is provided when the target sequence hybridizes to a second DNA probe that has been coupled to liposomes encapsulating the dye sulforhodamine B. The amount of liposomes captured in the detection zone can be read visually or quantified with a hand-held reflectometer. The biosensor can detect as little as 1 fmol target mRNA (1 nmol L(-1)). Specificity analysis revealed no cross-reactivity with 11 organisms tested, among them closely

  9. Experimental determination of vacuum-level band alignments of SnS-based solar cells by photoelectron yield spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Sugiyama, Mutsumi, E-mail: mutsumi@rs.noda.tus.ac.jp; Shimizu, Tsubasa; Kawade, Daisuke [Department of Electrical Engineering, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda 278-8510 (Japan); Ramya, Kottadi; Ramakrishna Reddy, K. T. [Department of Physics, Sri Venkateswara University, Tirupati 517502 (India)

    2014-02-28

    Energy band offsets of SnS-based solar cell structure using various n-type semiconductors, such as CdS, SnS{sub 2}, In{sub 2}S{sub 3}, ZnIn{sub 2}Se{sub 4}, ZnO, and Mg{sub 0.3}In{sub 0.7}O, are evaluated by photoelectron yield spectroscopy. The valence band discontinuities are estimated to be 1.6 eV for both SnS/CdS and SnS/SnS{sub 2}, 0.9 eV for SnS/In{sub 2}S{sub 3}, 1.7 eV for SnS/ZnIn{sub 2}Se{sub 4}, and 1.8 eV for both SnS/ZnO and SnS/Mg{sub 0.3}Zn{sub 0.7}O. Using the valence band discontinuity values and the corresponding energy bandgaps of the layers, energy band diagrams are developed. This study implied a type-I heterostructure, appropriate for SnS-based solar cell, for the ZnIn{sub 2}Se{sub 4} or Mg{sub x}Zn{sub 1−x}O (0 ≤ x ≤ 0.3) interface and type-II for other junctions.

  10. Experimental determination of vacuum-level band alignments of SnS-based solar cells by photoelectron yield spectroscopy

    Science.gov (United States)

    Sugiyama, Mutsumi; Shimizu, Tsubasa; Kawade, Daisuke; Ramya, Kottadi; Ramakrishna Reddy, K. T.

    2014-02-01

    Energy band offsets of SnS-based solar cell structure using various n-type semiconductors, such as CdS, SnS2, In2S3, ZnIn2Se4, ZnO, and Mg0.3In0.7O, are evaluated by photoelectron yield spectroscopy. The valence band discontinuities are estimated to be 1.6 eV for both SnS/CdS and SnS/SnS2, 0.9 eV for SnS/In2S3, 1.7 eV for SnS/ZnIn2Se4, and 1.8 eV for both SnS/ZnO and SnS/Mg0.3Zn0.7O. Using the valence band discontinuity values and the corresponding energy bandgaps of the layers, energy band diagrams are developed. This study implied a type-I heterostructure, appropriate for SnS-based solar cell, for the ZnIn2Se4 or MgxZn1-xO (0 ≤ x ≤ 0.3) interface and type-II for other junctions.

  11. Skin-derived precursors from human subjects with Type 2 diabetes yield dysfunctional vascular smooth muscle cells.

    Science.gov (United States)

    Steinbach, Sarah K; Yau, Terrence M; Ouzounian, Maral; Abdel-Qadir, Husam; Chandy, Mark; Waddell, Thomas K; Husain, Mansoor

    2017-08-01

    Objective : Few methods enable molecular and cellular studies of vascular aging or Type 2 diabetes (T2D). Here, we report a new approach to studying human vascular smooth muscle cell (VSMC) pathophysiology by examining VSMCs differentiated from progenitors found in skin. Approach and results : Skin-derived precursors (SKPs) were cultured from biopsies ( N =164, ∼1 cm 2 ) taken from the edges of surgical incisions of older adults ( N =158; males 72%; mean age 62.7 ± 13 years) undergoing cardiothoracic surgery, and differentiated into VSMCs at high efficiency (>80% yield). The number of SKPs isolated from subjects with T2D was ∼50% lower than those without T2D (cells/g: 0.18 ± 0.03, N =58 versus 0.40 ± 0.05, N =100, P <0.05). Importantly, SKP-derived VSMCs from subjects with T2D had higher Fluo-5F-determined baseline cytosolic Ca 2+ concentrations (AU: 1,968 ± 160, N =7 versus 1,386 ± 170, N =13, P <0.05), and a trend toward greater Ca 2+ cycling responses to norepinephrine (NE) (AUC: 177,207 ± 24,669, N =7 versus 101,537 ± 15,881, N =20, P <0.08) despite a reduced frequency of Ca 2+ cycling (events s -1 cell -1 : 0.011 ± 0.004, N =8 versus 0.021 ± 0.003, N =19, P <0.05) than those without T2D. SKP-derived VSMCs from subjects with T2D also manifest enhanced sensitivity to phenylephrine (PE) in an impedance-based assay (EC 50 nM: 72.3 ± 63.6, N =5 versus 3,684 ± 3,122, N =9, P <0.05), and impaired wound closure in vitro (% closure: 21.9 ± 3.6, N =4 versus 67.0 ± 10.3, N =4, P <0.05). Compared with aortic- and saphenous vein-derived primary VSMCs, SKP-derived VSMCs are functionally distinct, but mirror defects of T2D also exhibited by primary VSMCs. Skin biopsies from older adults yield sufficient SKPs to differentiate VSMCs, which reveal abnormal phenotypes of T2D that survive differentiation and persist even after long-term normoglycemic culture. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  12. Prospective clinical and radiographic evaluation of an allogeneic bone matrix containing stem cells (Trinity Evolution® Viable Cellular Bone Matrix) in patients undergoing two-level anterior cervical discectomy and fusion.

    Science.gov (United States)

    Peppers, Timothy A; Bullard, Dennis E; Vanichkachorn, Jed S; Stanley, Scott K; Arnold, Paul M; Waldorff, Erik I; Hahn, Rebekah; Atkinson, Brent L; Ryaby, James T; Linovitz, Raymond J

    2017-04-26

    Trinity Evolution® (TE), a viable cellular bone allograft, previously demonstrated high fusion rates and no safety-related concerns after single-level anterior cervical discectomy and fusion (ACDF) procedures. This prospective multicenter clinical study was performed to assess the radiographic and clinical outcomes of TE in subjects undergoing two-level ACDF procedures. In a prospective, multicenter study, 40 subjects that presented with symptomatic cervical degeneration at two adjacent vertebral levels underwent instrumented ACDF using TE autograft substitute in a polyetherethereketone (PEEK) cage. At 12 months, radiographic fusion status was evaluated by dynamic motion plain radiographs and thin cut CT with multiplanar reconstruction by a panel that was blinded to clinical outcome. Fusion success was defined by angular motion (≤4°) and the presence of bridging bone across the adjacent vertebral endplates. Clinical pain and function assessments included the Neck Disability Index (NDI), neck and arm pain as evaluated by visual analog scales (VAS), and SF-36 at both 6 and 12 months. At both 6 and 12 months, all clinical outcome scores (SF-36, NDI, and VAS pain) improved significantly (p Trinity Evolution in Anterior Cervical Disectomy and Fusion (ACDF) NCT00951938.

  13. PMA-Linked Fluorescence for Rapid Detection of Viable Bacterial Endospores

    Science.gov (United States)

    LaDuc, Myron T.; Venkateswaran, Kasthuri; Mohapatra, Bidyut

    2012-01-01

    The most common approach for assessing the abundance of viable bacterial endospores is the culture-based plating method. However, culture-based approaches are heavily biased and oftentimes incompatible with upstream sample processing strategies, which make viable cells/spores uncultivable. This shortcoming highlights the need for rapid molecular diagnostic tools to assess more accurately the abundance of viable spacecraft-associated microbiota, perhaps most importantly bacterial endospores. Propidium monoazide (PMA) has received a great deal of attention due to its ability to differentiate live, viable bacterial cells from dead ones. PMA gains access to the DNA of dead cells through compromised membranes. Once inside the cell, it intercalates and eventually covalently bonds with the double-helix structures upon photoactivation with visible light. The covalently bound DNA is significantly altered, and unavailable to downstream molecular-based manipulations and analyses. Microbiological samples can be treated with appropriate concentrations of PMA and exposed to visible light prior to undergoing total genomic DNA extraction, resulting in an extract comprised solely of DNA arising from viable cells. This ability to extract DNA selectively from living cells is extremely powerful, and bears great relevance to many microbiological arenas.

  14. Models to Estimate Lactation Curves of Milk Yield and Somatic Cell Count in Dairy Cows at the Herd Level for the Use in Simulations and Predictive Models.

    Science.gov (United States)

    Græsbøll, Kaare; Kirkeby, Carsten; Nielsen, Søren Saxmose; Halasa, Tariq; Toft, Nils; Christiansen, Lasse Engbo

    2016-01-01

    Typically, central milk recording data from dairy herds are recorded less than monthly. Over-fitting early in lactation periods is a challenge, which we explored in different ways by reducing the number of parameters needed to describe the milk yield and somatic cell count of individual cows. Furthermore, we investigated how the parameters of lactation models correlate between parities and from dam to offspring. The aim of the study was to provide simple and robust models for cow level milk yield and somatic cell count for fitting to sparse data to parameterize herd- and cow-specific simulation of dairy herds. Data from 610 Danish Holstein herds were used to determine parity traits in milk production regarding milk yield and somatic cell count of individual cows. Parity was stratified in first, second, and third and higher for milk, and first to sixth and higher for somatic cell count. Fitting of herd level parameters allowed for cow level lactation curves with three, two, or one parameters per lactation. Correlations of milk yield and somatic cell count were estimated between lactations and between dam and offspring. The shape of the lactation curves varied markedly between farms. The correlation between lactations for milk yield and somatic cell count was 0.2-0.6 and significant on more than 95% of farms. The variation in the daily milk yield was observed to be a source of variation to the somatic cell count, and the total somatic cell count was less correlated with the milk production than somatic cells per milliliter. A positive correlation was found between relative levels of the total somatic cell count and the milk yield. The variation of lactation and somatic cell count curves between farms highlights the importance of a herd level approach. The one-parameter per cow model using a herd level curve allows for estimating the cow production level from first the recording in the parity, while a two-parameter model requires more recordings for a credible

  15. Models to Estimate Lactation Curves of Milk Yield and Somatic Cell Count in Dairy Cows at the Herd Level for the Use in Simulations and Predictive Models

    Science.gov (United States)

    Græsbøll, Kaare; Kirkeby, Carsten; Nielsen, Søren Saxmose; Halasa, Tariq; Toft, Nils; Christiansen, Lasse Engbo

    2016-01-01

    Typically, central milk recording data from dairy herds are recorded less than monthly. Over-fitting early in lactation periods is a challenge, which we explored in different ways by reducing the number of parameters needed to describe the milk yield and somatic cell count of individual cows. Furthermore, we investigated how the parameters of lactation models correlate between parities and from dam to offspring. The aim of the study was to provide simple and robust models for cow level milk yield and somatic cell count for fitting to sparse data to parameterize herd- and cow-specific simulation of dairy herds. Data from 610 Danish Holstein herds were used to determine parity traits in milk production regarding milk yield and somatic cell count of individual cows. Parity was stratified in first, second, and third and higher for milk, and first to sixth and higher for somatic cell count. Fitting of herd level parameters allowed for cow level lactation curves with three, two, or one parameters per lactation. Correlations of milk yield and somatic cell count were estimated between lactations and between dam and offspring. The shape of the lactation curves varied markedly between farms. The correlation between lactations for milk yield and somatic cell count was 0.2–0.6 and significant on more than 95% of farms. The variation in the daily milk yield was observed to be a source of variation to the somatic cell count, and the total somatic cell count was less correlated with the milk production than somatic cells per milliliter. A positive correlation was found between relative levels of the total somatic cell count and the milk yield. The variation of lactation and somatic cell count curves between farms highlights the importance of a herd level approach. The one-parameter per cow model using a herd level curve allows for estimating the cow production level from first the recording in the parity, while a two-parameter model requires more recordings for a credible

  16. Roots of success: cultivating viable community forestry

    Energy Technology Data Exchange (ETDEWEB)

    MacQueen, Duncan

    2009-05-15

    Is community forestry emerging from the shadows? The evidence shows that locally controlled enterprises can be economically viable, and often build on stronger social and environmental foundations than the big private-sector players. Certainly this is an industry in need of a shakeup. Many forests have become flashpoints where agro-industry, large-scale logging concerns and conservation interests clash, while forest-dependent communities are left out in the cold. Meanwhile, governments – driven by concerns over the climate impacts of deforestation – are having to gear up for legal, sustainable forestry production. Community forestry could be crucial to solving many of these challenges. By building on local core capabilities and developing strategic partnerships, they are forging key new business models that could transform the sector.

  17. Factors affecting the CD34+ cell yields from the second donations of healthy donors: The steady-state lymphocyte count is a good predictive factor.

    Science.gov (United States)

    Guo, Zhi-Ping; Wang, Tao; Xu, Lan-Ping; Zhang, Xiao-Hui; Wang, Yu; Huang, Xiao-Jun; Chang, Ying-Jun

    2016-12-01

    A second allogeneic hematopoietic stem-cell transplantation and donor lymphocyte infusion using cells from the same donor is a therapeutic option in the case of stem-cell graft failure or disease relapse, but little is known about the factors associated with the CD34+ cell yields from second donations. One-hundred healthy donors who underwent a second mobilization treatment and peripheral blood stem-cell (PBSC) collection were studied. For both mobilization processes, 5 µg of granulocyte colony-stimulating factor per kg per day was administered. The blood counts of the donors were monitored during the processes. The second donations from the same donors provided lower apheresis yields than did the initial collections. The number of CD34+ cells collected from normal donors after a second cycle of PBSC mobilization was associated with their steady-state lymphocyte counts and the intertransplantation interval. Female sex negatively affected the CD34+ cell yields. The cutoff value for the steady-state absolute lymphocyte count was 2.055 × 109/L. To harvest greater numbers of CD34+ cells from second collections, male donors and those with intervals of longer than 9 months between donations should be selected. The lymphocyte counts prior to the first donations may predict the content of CD34+ cells in the allografts prepared using the second donations. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Polymerase chain reaction-based discrimination of viable from non-viable Mycoplasma gallisepticum

    Directory of Open Access Journals (Sweden)

    Ching Giap Tan

    2014-02-01

    Full Text Available The present study was based on the reverse transcription polymerase chain reaction (RT-PCR of the 16S ribosomal nucleic acid (rRNA of Mycoplasma for detection of viable Mycoplasma gallisepticum. To determine the stability of M. gallisepticum 16S rRNA in vitro, three inactivation methods were used and the suspensions were stored at different temperatures. The 16S rRNA of M. gallisepticum was detected up to approximately 20–25 h at 37 °C, 22–25 h at 16 °C, and 23–27 h at 4 °C. The test, therefore, could detect viable or recently dead M. gallisepticum (< 20 h. The RT-PCR method was applied during an in vivo study of drug efficacy under experimental conditions, where commercial broiler-breeder eggs were inoculated with M. gallisepticum into the yolk. Hatched chicks that had been inoculated in ovo were treated with Macrolide 1. The method was then applied in a flock of day 0 chicks with naturally acquired vertical transmission of M. gallisepticum, treated with Macrolide 2. Swabs of the respiratory tract were obtained for PCR and RT-PCR evaluations to determine the viability of M. gallisepticum. This study proved that the combination of both PCR and RT-PCR enables detection and differentiation of viable from non-viable M. gallisepticum.

  19. Improvement of kinetics, yield, and colloidal stability of biogenic gold nanoparticles using living cells of Euglena gracilis microalga

    Science.gov (United States)

    Dahoumane, Si Amar; Yéprémian, Claude; Djédiat, Chakib; Couté, Alain; Fiévet, Fernand; Coradin, Thibaud; Brayner, Roberta

    2016-03-01

    Recent years have witnessed a boom in the biosynthesis of a large variety of nanomaterials using different biological resources among which algae-based entities have been gaining much more attention within the community of material scientists worldwide. In our previously published findings, we explored some factors that governed the biofabrication of gold nanoparticles using living cultures of microalgae, such as the utilized microalgal genera, the phylum they belong to, and the impact of tetrachloroauric acid concentrations on the ability of these strains to perform the biosynthesis of gold nanoparticles once in contact with these cations. As a follow-up, we present in this paper an improvement of the features of bioproduced gold colloids using living cells of Euglena gracilis microalga when this species is grown under either mixotrophic or autotrophic conditions, i.e., exposed to light and grown in an organic carbon-enriched culture medium versus under autotrophic conditions. As an outcome to this alteration, the growth rate of this photosynthetic microorganism is multiplied 7-8 times when grown under mixotrophic conditions compared to autotrophic ones. Therefore, the yield, the kinetics, and the colloidal stability of the biosynthesized gold nanoparticles are dramatically enhanced. Moreover, the shape and the size of the as-produced nano-objects via this biological method are affected. In addition to round-shaped gold nanoparticles, particular shapes, such as triangles and hexagons, appear. These findings add up to the amassed knowledge toward the design of photobioreactors for the scalable and sustainable production of interesting nanomaterials.

  20. Improvement of kinetics, yield, and colloidal stability of biogenic gold nanoparticles using living cells of Euglena gracilis microalga

    Energy Technology Data Exchange (ETDEWEB)

    Dahoumane, Si Amar, E-mail: sa.dahoumane@gmail.com [Paris-Diderot University, Sorbonne Paris Cité, Interfaces, Traitements, Organisation et Dynamique des Systèmes (ITODYS), UMR 7086, CNRS (France); Yéprémian, Claude; Djédiat, Chakib; Couté, Alain [Muséum National d’Histoire Naturelle, Département RDDM, UMR 7245, Unité MCAM (France); Fiévet, Fernand [Paris-Diderot University, Sorbonne Paris Cité, Interfaces, Traitements, Organisation et Dynamique des Systèmes (ITODYS), UMR 7086, CNRS (France); Coradin, Thibaud, E-mail: thibaud.coradin@upmc.fr [UPMC—Paris 06, CNRS, Chimie de la Matière Condensée de Paris, Collège de France (France); Brayner, Roberta, E-mail: roberta.brayner@univ-paris-diderot.fr [Paris-Diderot University, Sorbonne Paris Cité, Interfaces, Traitements, Organisation et Dynamique des Systèmes (ITODYS), UMR 7086, CNRS (France)

    2016-03-15

    Recent years have witnessed a boom in the biosynthesis of a large variety of nanomaterials using different biological resources among which algae-based entities have been gaining much more attention within the community of material scientists worldwide. In our previously published findings, we explored some factors that governed the biofabrication of gold nanoparticles using living cultures of microalgae, such as the utilized microalgal genera, the phylum they belong to, and the impact of tetrachloroauric acid concentrations on the ability of these strains to perform the biosynthesis of gold nanoparticles once in contact with these cations. As a follow-up, we present in this paper an improvement of the features of bioproduced gold colloids using living cells of Euglena gracilis microalga when this species is grown under either mixotrophic or autotrophic conditions, i.e., exposed to light and grown in an organic carbon-enriched culture medium versus under autotrophic conditions. As an outcome to this alteration, the growth rate of this photosynthetic microorganism is multiplied 7–8 times when grown under mixotrophic conditions compared to autotrophic ones. Therefore, the yield, the kinetics, and the colloidal stability of the biosynthesized gold nanoparticles are dramatically enhanced. Moreover, the shape and the size of the as-produced nano-objects via this biological method are affected. In addition to round-shaped gold nanoparticles, particular shapes, such as triangles and hexagons, appear. These findings add up to the amassed knowledge toward the design of photobioreactors for the scalable and sustainable production of interesting nanomaterials.

  1. Genetic correlations among milk yield, morphology, performance test traits and somatic cells in dual-purpose Rendena breed.

    Science.gov (United States)

    Sartori, C; Guzzo, N; Mazza, S; Mantovani, R

    2017-10-17

    Selection in native local breeds needs great carefulness due to the small population size and the risk of inbreeding. Furthermore, most breeds are dual-purpose, and milk and beef attitudes are antagonistic. For preservation purposes functional traits need to be considered. Focusing on the small local Rendena cattle, this study aimed to analyse the genetic correlations among milk, beef and udder health traits and the response to selection predicted under different scenarios. The study considered milk, fat and protein yields (MY), factor scores for udder volume (UV), conformation (UC) and muscularity obtained from type traits scored on primiparous cows, and performance test traits (PT) measured on young bulls at test station: average daily gain, in vivo SEUROP fleshiness, in vivo dressing percentage. Somatic cell score (SCS) was considered as a functional trait, with a possibility of restricting its genetic gain to zero. The study considered 281 497 MY test-day data collected on 16 974 cows, and data from linear type evaluation on 11 992 primiparous cows for factor scores. The PT data were recorded on 1428 young bulls, and SCS obtained from cell counts at milk recording. Bi-trait restricted maximum likelihood animal model analyses were performed to assess genetic parameters. Heritability varied from 0.157 (fat) to 0.442 (dressing percentage). Udder volume and MY resulted positively genetically correlated (average correlation 0.427), whereas the low-negative genetic correlation between MY and UC (-0.141) suggested a negative impact of milk gain on udder form. Beef traits of factor muscularity and PT showed medium-high favourable genetic correlations (from 0.357 to 0.984), excluding a null correlation between daily gain and muscularity. The genetic correlation MY v. muscularity was unfavourable (-0.328 on average), whereas null correlations were found in MY v. PT, apart from fat v. dressing percentage (-0.151). Somatic cell score showed low unfavourable correlations

  2. Reverse transcriptase real-time PCR for detection and quantification of viable Campylobacter jejuni directly from poultry faecal samples

    DEFF Research Database (Denmark)

    Bui, Thanh Xuan; Wolff, Anders; Madsen, Mogens

    2012-01-01

    and quantification of viable Campylobacter jejuni directly from chicken faecal samples. The results of this method anda DNA-based quantitative real-time PCR (qPCR) method were compared with those of a bacterial culture method. Using bacterial culture andRT-qPCR methods, viable C. jejuni cells could be detected...

  3. The Conceptual Mechanism for Viable Organizational Learning Based on Complex System Theory and the Viable System Model

    Science.gov (United States)

    Sung, Dia; You, Yeongmahn; Song, Ji Hoon

    2008-01-01

    The purpose of this research is to explore the possibility of viable learning organizations based on identifying viable organizational learning mechanisms. Two theoretical foundations, complex system theory and viable system theory, have been integrated to provide the rationale for building the sustainable organizational learning mechanism. The…

  4. Is Greenberg's "Macro-Carib" viable?

    Directory of Open Access Journals (Sweden)

    Spike Gildea

    Full Text Available In his landmark work Language in the Americas, Greenberg (1987 proposed that Macro-Carib was one of the major low-level stocks of South America, which together with Macro-Panoan and Macro-Ge-Bororo were claimed to comprise the putative Ge-Pano-Carib Phylum. His Macro-Carib includes the isolates Andoke and Kukura, and the Witotoan, Peba-Yaguan, and Cariban families. Greenberg's primary evidence came from person-marking paradigms in individual languages, plus scattered words from individual languages collected into 79 Macro-Carib 'etymologies' and another 64 Amerind 'etymologies'. The goal of this paper is to re-evaluate Greenberg's Macro-Carib claim in the light of the much more extensive and reliable language data that has become available largely since 1987. Based on full person-marking paradigms for Proto-Cariban, Yagua, Bora and Andoke, we conclude that Greenberg's morphological claims are unfounded. For our lexical comparison, we created lexical lists for Proto-Cariban, Proto-Witotoan, Yagua and Andoke, for both Greenberg's 143 putative etymologies and for the Swadesh 100 list. From both lists, a total of 23 potential cognates were found, but no consonantal correspondences were repeated even once. We conclude that our greatly expanded and improved database does not provide sufficient evidence to convince the skeptic that the Macro-Carib hypothesis is viable

  5. Economically viable large-scale hydrogen liquefaction

    Science.gov (United States)

    Cardella, U.; Decker, L.; Klein, H.

    2017-02-01

    The liquid hydrogen demand, particularly driven by clean energy applications, will rise in the near future. As industrial large scale liquefiers will play a major role within the hydrogen supply chain, production capacity will have to increase by a multiple of today’s typical sizes. The main goal is to reduce the total cost of ownership for these plants by increasing energy efficiency with innovative and simple process designs, optimized in capital expenditure. New concepts must ensure a manageable plant complexity and flexible operability. In the phase of process development and selection, a dimensioning of key equipment for large scale liquefiers, such as turbines and compressors as well as heat exchangers, must be performed iteratively to ensure technological feasibility and maturity. Further critical aspects related to hydrogen liquefaction, e.g. fluid properties, ortho-para hydrogen conversion, and coldbox configuration, must be analysed in detail. This paper provides an overview on the approach, challenges and preliminary results in the development of efficient as well as economically viable concepts for large-scale hydrogen liquefaction.

  6. Effects of ovarian disaggregation on adult murine follicle yield and viability.

    Science.gov (United States)

    Young, Fiona; Drummond, John; Akers, Emma; Bartle, Louise; Kennedy, David; Asaduzzaman, Mohammad

    2017-11-01

    Follicles are isolated from ovaries for numerous reasons, including IVM, but adult murine yields are GraphPad Prism. Col-IV disaggregation yielded the highest number of follicles (17±10 folliclesmg-1 ovarian tissue). All disaggregation methods released more secondary follicles (86±20 per ovary; P<0.05) than any other size cohort. Mechanical and Col-IV disaggregation yielded similar numbers of morphologically intact follicles, whereas AOF disaggregation caused more damage (P<0.01). As the morphological disruption increased, DAPI and CMXRos staining decreased (P<0.05), and tubulin localisation became more heterogeneous. Col-IV disaggregation gave the best yield of morphologically intact follicles containing viable granulosa cells. In conclusion, we improved adult murine follicle yields and applied molecular markers to assess follicle morphology, cellular cytoskeleton and mitochondrial function.

  7. Is telomerase a viable target in cancer?

    Science.gov (United States)

    Buseman, C.M.; Wright, W.E.; Shay, J.W.

    2012-01-01

    The ideal cancer treatment would specifically target cancer cells yet have minimal or no adverse effects on normal somatic cells. Telomerase, the ribonucleoprotein reverse transcriptase that maintains the ends of human chromosome, is an attractive cancer therapeutic target for exactly this reason [1]. Telomerase is expressed in more than 85% of cancer cells, making it a nearly universal cancer marker, while the majority of normal somatic cells are telomerase negative. Telomerase activity confers limitless replicative potential to cancer cells, a hallmark of cancer which must be attained for the continued growth that characterizes almost all advanced neoplasms [2]. In this review we will summarize the role of telomeres and telomerase in cancer cells, and how properties of telomerase are being exploited to create targeted cancer therapies including telomerase inhibitors, telomerase-targeted immunotherapies and telomerase-driven virotherapies. A frank and balanced assessment of the current state of telomerase inhibitors with caveats and potential limitations will be included. PMID:21802433

  8. yield indicators

    African Journals Online (AJOL)

    YIELD INDICATORS. P. NTAWURUHUNGA, P.R. RUBAIHAYOI, J.B.A. WHYTE, A.G.O. DIXONZ and use. osnzu1. International Institute of Tropical Agriculture, East and Southern Africa, Centre, PO. Box 7878, l Kampala ... most important sources of food energy in several ... efficiency in selecting and identifying cassava.

  9. Parejas viables que perduran en el tiempo

    Directory of Open Access Journals (Sweden)

    Juan José Cuervo Rodríguez

    2013-01-01

    Full Text Available El presente artículo científico presenta resultados del proceso llevado a cabo en el proyecto de investigación docente "Mecanismos de autorregulación en parejas viables que perduran en el tiempo". Se soporta en una mirada compleja de la psicología basada en una epistemología de la construcción. En el ámbito metodológico, se inscribe en los estudios de terapia familiar desde una perspectiva de la comunicación humana como un todo integrado. Participaron nueve parejas. Los criterios de inclusión fueron: cinco o más años de convivencia, participación voluntaria, no presentar (ni haber presentado problemáticas especiales que ameriten intervención psicoterapéutica y la obtención de un porcentaje significativo en el uso de estrategias de comunicación asertiva en la resolución de conflictos. El método general utilizado fue el análisis de la comunicación en tarea de conversación. Los principales hallazgos señalan una estrecha relación entre el contexto de desarrollo de las parejas, la emergencia de códigos comunicacionales propios y la posibilidad de perdurar en el tiempo; también, se resalta el tipo de comunicación asertiva o constructiva, la construcción de valores como el respeto y la aceptación de las diferencias, y el deseo por vivir y construir bienestar común, como elementos constitutivos de su identidad como pareja.

  10. Predictors of mother and child DNA yields in buccal cell samples collected in pediatric cancer epidemiologic studies: a report from the Children’s Oncology group

    Science.gov (United States)

    2013-01-01

    Background Collection of high-quality DNA is essential for molecular epidemiology studies. Methods have been evaluated for optimal DNA collection in studies of adults; however, DNA collection in young children poses additional challenges. Here, we have evaluated predictors of DNA quantity in buccal cells collected for population-based studies of infant leukemia (N = 489 mothers and 392 children) and hepatoblastoma (HB; N = 446 mothers and 412 children) conducted through the Children’s Oncology Group. DNA samples were collected by mail using mouthwash (for mothers and some children) and buccal brush (for children) collection kits and quantified using quantitative real-time PCR. Multivariable linear regression models were used to identify predictors of DNA yield. Results Median DNA yield was higher for mothers in both studies compared with their children (14 μg vs. <1 μg). Significant predictors of DNA yield in children included case–control status (β = −0.69, 50% reduction, P = 0.01 for case vs. control children), brush collection type, and season of sample collection. Demographic factors were not strong predictors of DNA yield in mothers or children in this analysis. Conclusions The association with seasonality suggests that conditions during transport may influence DNA yield. The low yields observed in most children in these studies highlight the importance of developing alternative methods for DNA collection in younger age groups. PMID:23937514

  11. High-yield production of a stable Vero cell-based vaccine candidate against the highly pathogenic avian influenza virus H5N1

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Fangye; Zhou, Jian; Ma, Lei; Song, Shaohui; Zhang, Xinwen; Li, Weidong; Jiang, Shude [No. 5, Department of Bioproducts, Institute of Medical Biology, Chinese Academy of Medical Science and Pecking Union Medical College, Jiaoling Avenue 935, Kunming, Yunnan Province 650102, People' s Republic of China (China); Wang, Yue, E-mail: euy-tokyo@umin.ac.jp [National Institute for Viral Disease Control and Prevention, China Center for Disease Control and Prevention, Yingxin Lane 100, Xicheng District, Beijing 100052, People' s Republic of China (China); Liao, Guoyang, E-mail: liaogy@21cn.com [No. 5, Department of Bioproducts, Institute of Medical Biology, Chinese Academy of Medical Science and Pecking Union Medical College, Jiaoling Avenue 935, Kunming, Yunnan Province 650102, People' s Republic of China (China)

    2012-05-18

    Highlights: Black-Right-Pointing-Pointer Vero cell-based HPAI H5N1 vaccine with stable high yield. Black-Right-Pointing-Pointer Stable high yield derived from the YNVa H3N2 backbone. Black-Right-Pointing-Pointer H5N1/YNVa has a similar safety and immunogenicity to H5N1delta. -- Abstract: Highly pathogenic avian influenza (HPAI) viruses pose a global pandemic threat, for which rapid large-scale vaccine production technology is critical for prevention and control. Because chickens are highly susceptible to HPAI viruses, the supply of chicken embryos for vaccine production might be depleted during a virus outbreak. Therefore, developing HPAI virus vaccines using other technologies is critical. Meeting vaccine demand using the Vero cell-based fermentation process has been hindered by low stability and yield. In this study, a Vero cell-based HPAI H5N1 vaccine candidate (H5N1/YNVa) with stable high yield was achieved by reassortment of the Vero-adapted (Va) high growth A/Yunnan/1/2005(H3N2) (YNVa) virus with the A/Anhui/1/2005(H5N1) attenuated influenza vaccine strain (H5N1delta) using the 6/2 method. The reassorted H5N1/YNVa vaccine maintained a high hemagglutination (HA) titer of 1024. Furthermore, H5N1/YNVa displayed low pathogenicity and uniform immunogenicity compared to that of the parent virus.

  12. Maintenance of pathogenicity during entry into and resuscitation from viable but nonculturable state in Aeromonas hydrophila exposed to natural seawater at low temperature.

    Science.gov (United States)

    Maalej, S; Gdoura, R; Dukan, S; Hammami, A; Bouain, A

    2004-01-01

    To investigate the fate of Aeromonas hydrophila pathogenicity when cells switch, in nutrient-poor filtered sterilized seawater, between the culturable and nonculturable state. Aeromonas hydrophila ATCC 7966, rendered non culturable within 50-55 days of exposure to marine stress conditions, was tested for its ability to maintain haemolysin and to adhere to McCoy cells. Results showed that pathogenicity was lost concomitantly with culturability, whereas cell viability remained undamaged, as determined by the Kogure cell elongation test. However, this loss is only temporary because, following temperature shift from 5 to 23 degrees C, multiple biological activities of recovered Aer. hydrophila cells, which include their ability to lyse human erythrocytes and to attach and destroy McCoy cells were regained. During the temperature-induced resuscitation, constant total cell counts were observed. Moreover, no significant improvement in recovery yield was obtained on brain-heart infusion (BHI) agar plates amended with catalase. We suggest that in addition to the growth of the few undetected culturable cells, there is repair and growth of some mildly injured viable but nonculturable cells. The possibility that nonculturable cells of normally culturable Aer. hydrophila in natural marine environment may constitute a source of infectious diseases posing a public health problem was demonstrated. These experiments may mimic what happens when Aer. hydrophila cells are released in natural seawater with careful attention to the conditions in which surrounding waters gradually become warmer in late summer/early autumn. Copyright 2004 The Society for Applied Microbiology

  13. Multi-institutional investigation of the prognostic value of lymph nodel yield in advanced-stage oral cavity squamous cell carcinoma.

    Science.gov (United States)

    Jaber, James J; Zender, Chad A; Mehta, Vikas; Davis, Kara; Ferris, Robert L; Lavertu, Pierre; Rezaee, Rod; Feustel, Paul J; Johnson, Jonas T

    2014-10-01

    Although existing literature provides surgical recommendations for treating occult disease (cN0) in early-stage oral cavity squamous cell carcinoma (SCC), a focus on late-stage oral cavity SCC is less pervasive. The medical records of 162 patients with late-stage oral cavity SCC pN0 who underwent primary neck dissections were reviewed. Lymph node yield as a prognosticator was examined. Despite being staged pN0, patients that had a higher lymph node yield had an improved regional/distant control rates, disease-free survival (DFS), disease-specific survival (DSS), and overall survival (OS). Lymph node yield consistently outperformed all other standard variables as being the single best prognostic factor with a tight risk ratio range (RR = 0.95-0.98) even when correcting for the number of lymph nodes examined. The results of this study showed that lower regional recurrence rates and improved survival outcomes were seen as lymph node yield increased for advanced T classification oral cavity SCC pN0. This suggests that increasing lymph node yield with an extended cervical lymphadenectomy may result in lower recurrence rates and improved survival outcomes for this advanced stage group. © 2014 Wiley Periodicals, Inc.

  14. Determination of viable Salmonellae from potable and source water through PMA assisted qPCR.

    Science.gov (United States)

    Singh, Gulshan; Vajpayee, Poornima; Bhatti, Saurabh; Ronnie, Nirmala; Shah, Nimish; McClure, Peter; Shanker, Rishi

    2013-07-01

    Resource constrained countries identified as endemic zones for pathogenicity of Salmonella bear an economic burden due to recurring expenditure on medical treatment. qPCR used for Salmonella detection could not discriminate between viable and nonviable cells. Propidium monoazide (PMA) that selectively penetrates nonviable cells to cross-link their DNA, was coupled with ttr gene specific qPCR for quantifying viable salmonellae in source/potable waters collected from a north Indian city. Source water (raw water for urban potable water supply) and urban potable water exhibited viable salmonellae in the range of 2.1×10(4)-2.6×10(6) and 2-7160CFU/100mL, respectively. Potable water at water works exhibited DNA from dead cells but no viable cells were detected. PMA assisted qPCR could specifically detect low numbers of live salmonellae in Source and potable waters. This strategy can be used in surveillance of urban potable water distribution networks to map contamination points for better microbial risk management. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. A novel fed-batch based cultivation method provides high cell-density and improves yield of soluble recombinant proteins in shaken cultures

    Science.gov (United States)

    2010-01-01

    Background Cultivations for recombinant protein production in shake flasks should provide high cell densities, high protein productivity per cell and good protein quality. The methods described in laboratory handbooks often fail to reach these goals due to oxygen depletion, lack of pH control and the necessity to use low induction cell densities. In this article we describe the impact of a novel enzymatically controlled fed-batch cultivation technology on recombinant protein production in Escherichia coli in simple shaken cultures. Results The enzymatic glucose release system together with a well-balanced combination of mineral salts and complex medium additives provided high cell densities, high protein yields and a considerably improved proportion of soluble proteins in harvested cells. The cultivation method consists of three steps: 1) controlled growth by glucose-limited fed-batch to OD600 ~10, 2) addition of growth boosters together with an inducer providing efficient protein synthesis within a 3 to 6 hours period, and 3) a slow growth period (16 to 21 hours) during which the recombinant protein is slowly synthesized and folded. Cell densities corresponding to 10 to 15 g l-1 cell dry weight could be achieved with the developed technique. In comparison to standard cultures in LB, Terrific Broth and mineral salt medium, we typically achieved over 10-fold higher volumetric yields of soluble recombinant proteins. Conclusions We have demonstrated that by applying the novel EnBase® Flo cultivation system in shaken cultures high cell densities can be obtained without impairing the productivity per cell. Especially the yield of soluble (correctly folded) proteins was significantly improved in comparison to commonly used LB, Terrific Broth or mineral salt media. This improvement is thought to result from a well controlled physiological state during the whole process. The higher volumetric yields enable the use of lower culture volumes and can thus significantly reduce

  16. Asouzu's Complementary Ontology as a Foundation for a Viable ...

    African Journals Online (AJOL)

    This paper on “Asouzu's Complementary Ontology as a foundation for a viable Ethic of the Environment”, posits that an ethic of the environment can be seen as viable if it considers the whole of reality as ontologically relevant. This point of view would free environmental ethics of anthropocentric bias and its attendant ...

  17. Models to estimate lactation curves of milk yield and somatic cell count in dairy cows at the herd level for the use in simulations and predictive models

    Directory of Open Access Journals (Sweden)

    Kaare Græsbøll

    2016-12-01

    Full Text Available Typically, central milk recording data from dairy herds are recorded less than monthly. Over-fitting early in lactation periods is a challenge, which we explored in different ways by reducing the number of parameters needed to describe the milk yield and somatic cell count of individual cows. Furthermore, we investigated how the parameters of lactation models correlate between parities and from dam to offspring. The aim of the study was to provide simple and robust models for cow level milk yield and somatic cell count (SCC for fitting to sparse data to parameterise herd- and cow-specific simulation of dairy herds.Data from 610 Danish Holstein herds were used to determine parity traits in milk production regarding milk yield and SCC of individual cows. Parity was stratified in first, second and third and higher for milk, and first to sixth and higher for SCC. Fitting of herd level parameters allowed for cow level lactation curves with three, two or one-parameters per lactation. Correlations of milk yield and SCC were estimated between lactations and between dam and offspring.The shape of the lactation curves varied markedly between farms. The correlation between lactations for milk yield and SCC were 0.2-0.6 and significant on more than 95% of farms. The variation in the daily milk yield was observed to be a source of variation to the SCC, and the total SCC was less correlated with the milk production than somatic cells per ml. A positive correlation was found between relative levels of the total SCC and the milk yield.The variation of lactation and SCC curves between farms highlights the importance of a herd level approach. The one-parameter per cow model using a herd level curve allows for estimating the cow production level from first the recording in the parity, while a two-parameter model requires more recordings for a credible estimate, but may more precisely predict persistence, and given the independence of parameters, these can be

  18. An invention of thermo-responsive polymer surface, yielding cell sheet based regenerative therapies in cardiology and ophthalmology

    OpenAIRE

    Sawa Y; KM Cherian; Mohanty S; Ganesh JS; Abraham S

    2015-01-01

    The Invention: In vitro cell culture methodologies provide a conducive environment for the cells taken out of their native environment to grow and proliferate in a non-physiological environment, the culture dish. Research experiments have been focusing on various criteria for assessing how far it is possible to recapitulate the native extra-cellular environment in vitro. Scaffolds, culture media, growth factors and cell surface modified culture dishes are some of the components that provi...

  19. Separation of viable and non-viable tomato (Solanum lycopersicum L.) seeds using single seed near-infrared spectroscopy

    DEFF Research Database (Denmark)

    Shrestha, Santosh; Deleuran, Lise Christina; Gislum, René

    2017-01-01

    -viable tomato seeds of two cultivars using chemometrics. The data exploration were performed by principal component analysis (PCA). Subsequently, viable and non-viable seeds were classified by partial least squares-discriminant analysis (PLS-DA) and interval PLS-DA (iPLS-DA). The indication of clustering...... of viable and non-viable seeds were observed in the PCA of each cultivar and the pooled samples. However, the PCA did not exhibit a pattern of separation among the early, normal and late germinated tomato seeds. The NIR spectral regions of 1160–1170, 1383–1397, 1647–1666, 1860–1884 and 1915–1940 nm were...... identified as important for classification of viable and non-viable tomato seeds by iPLS-DA. The sensitivity i.e. ability to correctly identify the positive samples and specificity i.e. ability to reject the negative samples of the (iPLS-DA) model on identified spectral regions for prediction of viable...

  20. The yield and quality of cellular and bacterial DNA extracts from human oral rinse samples are variably affected by the cell lysis methodology.

    Science.gov (United States)

    Sohrabi, Mohsen; Nair, Raj G; Samaranayake, Lakshman P; Zhang, Li; Zulfiker, Abu Hasanat Md; Ahmetagic, Adnan; Good, David; Wei, Ming Q

    2016-03-01

    Recent culture-independent studies have enabled detailed mapping of human microbiome that has not been hitherto achievable by culture-based methods. DNA extraction is a key element of bacterial culture-independent studies that critically impacts on the outcome of the detected microbial profile. Despite the variations in DNA extraction methods described in the literature, no standardized technique is available for the purpose of microbiome profiling. Hence, standardization of DNA extraction methods is urgently needed to yield comparable data from different studies. We examined the effect of eight different cell lysis protocols on the yield and quality of the extracted DNA from oral rinse samples. These samples were exposed to cell lysis techniques based on enzymatic, mechanical, and a combination of enzymatic-mechanical methods. The outcome measures evaluated were total bacterial population, Firmicutes levels and human DNA contamination (in terms of surrogate GAPDH levels). We noted that all three parameters were significantly affected by the method of cell lysis employed. Although the highest yield of gDNA was obtained using lysozyme-achromopeptidase method, the lysozyme-zirconium beads method yielded the peak quantity of total bacterial DNA and Firmicutes with a lower degree of GAPDH contamination compared with the other methods. Taken together our data clearly points to an urgent need for a consensus, standardized DNA extraction technique to evaluate the oral microbiome using oral rinse samples. Further, if Firmicutes levels are the focus of investigation in oral rinse microbiome analyses then the lysozyme-zirconium bead method would be the method of choice in preference to others. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Effects of bovine subclinical mastitis caused by Corynebacterium spp. on somatic cell count, milk yield and composition by comparing contralateral quarters.

    Science.gov (United States)

    Gonçalves, Juliano Leonel; Tomazi, Tiago; Barreiro, Juliana Regina; Beuron, Daniele Cristine; Arcari, Marcos André; Lee, Sarah Hwa In; Martins, Cristian Marlon de Magalhães Rodrigues; Araújo Junior, João Pessoa; dos Santos, Marcos Veiga

    2016-03-01

    Subclinical mastitis caused by Corynebacterium spp. (as a group and at the species level) was investigated by evaluating contralateral (healthy and infected) mammary quarters for somatic cell count (SCC), milk yield and composition. Selection of cows with subclinical mastitis caused by Corynebacterium spp. was performed by microbiological culture of composite samples collected from 1242 dairy cows from 21 dairy herds. For each of the selected cows, milk yield was measured and milk samples were collected at the mammary quarter level (i.e., 1140 mammary samples collected from 285 cows) for analysis of milk composition and SCC. The identification of Corynebacterium spp. isolates was performed by 16S rRNA gene sequencing. One hundred and eighty Corynebacterium spp. isolates were identified, of which 167 (92.77%) were C.bovis and eight (4.44%) non-C.bovis; for five of the Corynebacterium spp. isolates (2.77%), sequencing of 16S rRNA genes did not allow identification at the species level. Mammary quarters infected with Corynebacterium spp. as a group had a higher geometric mean SCC (197,900 cells/mL) than healthy contralateral mammary quarters (85,800 cells/mL). Species of Corynebacterium non-C.bovis were infrequently isolated and did not change SCC, milk yield or milk solid contents when evaluated at the contralateral quarter level. Although C.bovis infection showed no effect on milk yield, fat, protein, casein or total solids in milk, it increased SCC and decreased lactose and milk solids non-fat content. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Predictors of mother and child DNA yields in buccal cell samples collected in pediatric cancer epidemiologic studies: a report from the Children's Oncology group.

    Science.gov (United States)

    Poynter, Jenny N; Ross, Julie A; Hooten, Anthony J; Langer, Erica; Blommer, Crystal; Spector, Logan G

    2013-08-12

    Collection of high-quality DNA is essential for molecular epidemiology studies. Methods have been evaluated for optimal DNA collection in studies of adults; however, DNA collection in young children poses additional challenges. Here, we have evaluated predictors of DNA quantity in buccal cells collected for population-based studies of infant leukemia (N = 489 mothers and 392 children) and hepatoblastoma (HB; N = 446 mothers and 412 children) conducted through the Children's Oncology Group. DNA samples were collected by mail using mouthwash (for mothers and some children) and buccal brush (for children) collection kits and quantified using quantitative real-time PCR. Multivariable linear regression models were used to identify predictors of DNA yield. Median DNA yield was higher for mothers in both studies compared with their children (14 μg vs. mothers or children in this analysis. The association with seasonality suggests that conditions during transport may influence DNA yield. The low yields observed in most children in these studies highlight the importance of developing alternative methods for DNA collection in younger age groups.

  3. Reproducible high yields of recombinant adeno-associated virus produced using invertebrate cells in 0.02- to 200-liter cultures.

    Science.gov (United States)

    Cecchini, Sylvain; Virag, Tamas; Kotin, Robert M

    2011-08-01

    The large amounts of recombinant adeno-associated virus (rAAV) vector needed for clinical trials and eventual commercialization require robust, economical, reproducible, and scalable production processes compatible with current good manufacturing practice. rAAV produced using baculovirus and insect cells satisfies these conditions; however, recovering rAAV particles from 200-liter bioreactors is more complicated than bench-scale vector preparations. Using a variety of processing media, we developed a reliable and routine downstream procedure for rAAV production that is scalable from 0.02- to 200-liter cultures. To facilitate the upstream process, we adapted the titerless infected-cell preservation and scale-up process for rAAV production. Single-use aliquots of cryopreserved baculovirus-infected insect cells (BIIC) are thawed and added to the suspension culture to achieve the desired ratio of BIIC to rAAV-producer cells. By using conditions established with small-scale cultures, rAAV was produced in larger volume cultures. Strikingly consistent rAAV yields were attained in cultures ranging from 10 liters to 200 liters. Based on the final yield, each cell produced 18,000 ± 6,800 particles of purified rAAV in 10-, 20-, 100-, and 200-liter cultures. Thus, with an average cell density of 4.32 × 10(6) cells/ml, ≥ 10(16) purified rAAV particles are produced from 100 to 200 liters. The downstream process resulted in about 20% recovery estimated from comparing the quantities of capsid protein antigen in the crude bioreactor material and in the final, purified product. The ease and reproducibility of rAAV production in 200-liter bioreactors suggest that the limit has not been reached, and 500-liter productions are planned.

  4. Oxidative Damage and Mutagenic Potency of Fast Neutron and UV-B Radiation in Pollen Mother Cells and Seed Yield of Vicia faba L.

    Directory of Open Access Journals (Sweden)

    Ekram Abdel Haliem

    2013-01-01

    Full Text Available In recent years, there has been a great deal of attention toward free radicals, reactive oxygen species (ROS generated by exposure of crop plant cells to physical radiations. Henceforth, the current study was planned to compare oxidative stress and mutagenic potential of different irradiation doses of fast neutron (FN and UV-B on meiotic-pollen mother cells (PMCs, pollen grains (PGs and seeds yielded from irradiated faba beans seedlings. On the cytogenetic level, each irradiation type had special interference with DNA of PMC and exhibited wide range of mutagenic action on the frequency and type of chromosomal anomalies, fertility of PGs and seed yield productivity based on the irradiation exposure dose and radiation sensitivity of faba bean plants compared with un-irradiated ones. On the molecular level, SDS-PAGE and RPAD-PCR analyses of seeds yielded from irradiated seedlings exhibited distinctive polymorphisms based on size, intensity, appearance, and disappearance of polypeptides bands compared with un-irradiated ones. The total values of protein and DNA polymorphisms reached 88% and 90.80% respectively. The neutron fluency (2.3 × 106 n/cm2 and UV-B dose for 1 hr were recorded as bio-positive effects. The present study proved that genetic variations revealed by cytogenetic test could be supported by gene expression (alterations in RAPD and protein profiles.

  5. Characterization of the Viable but Nonculturable (VBNC State in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Mohammad Salma

    Full Text Available The Viable But Non Culturable (VBNC state has been thoroughly studied in bacteria. In contrast, it has received much less attention in other microorganisms. However, it has been suggested that various yeast species occurring in wine may enter in VBNC following sulfite stress.In order to provide conclusive evidences for the existence of a VBNC state in yeast, the ability of Saccharomyces cerevisiae to enter into a VBNC state by applying sulfite stress was investigated. Viable populations were monitored by flow cytometry while culturable populations were followed by plating on culture medium. Twenty-four hours after the application of the stress, the comparison between the culturable population and the viable population demonstrated the presence of viable cells that were non culturable. In addition, removal of the stress by increasing the pH of the medium at different time intervals into the VBNC state allowed the VBNC S. cerevisiae cells to "resuscitate". The similarity between the cell cycle profiles of VBNC cells and cells exiting the VBNC state together with the generation rate of cells exiting VBNC state demonstrated the absence of cellular multiplication during the exit from the VBNC state. This provides evidence of a true VBNC state. To get further insight into the molecular mechanism pertaining to the VBNC state, we studied the involvement of the SSU1 gene, encoding a sulfite pump in S. cerevisiae. The physiological behavior of wild-type S. cerevisiae was compared to those of a recombinant strain overexpressing SSU1 and null Δssu1 mutant. Our results demonstrated that the SSU1 gene is only implicated in the first stages of sulfite resistance but not per se in the VBNC phenotype. Our study clearly demonstrated the existence of an SO2-induced VBNC state in S. cerevisiae and that the stress removal allows the "resuscitation" of VBNC cells during the VBNC state.

  6. High-Yield Method for Isolation and Culture of Endothelial Cells from Rat Coronary Blood Vessels Suitable for Analysis of Intracellular Calcium and Nitric Oxide Biosynthetic Pathways

    Directory of Open Access Journals (Sweden)

    Nistri Silvia

    2002-01-01

    Full Text Available We describe here a method for isolating endothelial cells from rat heart blood vessels by means of coronary microperfusion with collagenase. This methods makes it possible to obtain high amounts of endothelial cells in culture which retain the functional properties of their in vivo counterparts, including the ability to uptake fluorescently-labeled acetylated low-density lipoproteins and to respond to vasoactive agents by modulating intracellular calcium and by upregulating intrinsic nitric oxide generation. The main advantages of our technique are: (i good reproducibility, (ii accurate sterility that can be maintained throughout the isolation procedure and (iii high yield of pure endothelial cells, mainly due to microperfusion and temperature-controlled incubation with collagenase which allow an optimal distribution of this enzyme within the coronary vascular bed.

  7. Non-viable Borrelia burgdorferi induce inflammatory mediators and apoptosis in human oligodendrocytes.

    Science.gov (United States)

    Parthasarathy, Geetha; Fevrier, Helene B; Philipp, Mario T

    2013-11-27

    In previous studies, exposure to live Borrelia burgdorferi was shown to induce inflammation and apoptosis of human oligodendrocytes. In this study we assessed the ability of non-viable bacteria (heat killed or sonicated) to induce inflammatory mediators and cell death. Both heat-killed and sonicated bacteria induced release of CCL2, IL-6, and CXCL8 from oligodendrocytes in a dose dependent manner. In addition, non-viable B. burgdorferi also induced cell death as evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and another cell viability assay. These results suggest that spirochetal residues left after bacterial demise, due to treatment or otherwise, may continue to be pathogenic to the central nervous system. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  8. Display of cellulases on the cell surface of Saccharomyces cerevisiae for high yield ethanol production from high-solid lignocellulosic biomass.

    Science.gov (United States)

    Matano, Yuki; Hasunuma, Tomohisa; Kondo, Akihiko

    2012-03-01

    Economically feasible processes for industrial cellulosic ethanol production requires increasing the final ethanol titer during fermentation due to the high energy demands of the subsequent ethanol distillation. In the present study, high-yield ethanol production was achieved by short-term liquefaction and fermentation of lignocellulose biomass in a novel drum-type rotary fermentation system using a yeast strain developed for cell-surface display of fungal endoglucanase, cellobiohydrolase, and β-glucosidase. In the presence of 10 FPU/g-biomass cellulase added, the recombinant cellulolytic strain produced 1.4-fold higher ethanol (89% of theoretical yield) from high-solid (200 g-dry weight/L) rice straw within 72 h of fermentation than wild type strain. Cell-surface engineering successfully reduced the amount of commercial enzyme required for the fermentation of cellulose. This study demonstrates that cellulases displayed on the yeast cell surface are capable of hydrolyzing cellulose that was not hydrolyzed by commercial cellulases, leading to increased sugar utilization for improved ethanol production. Copyright © 2012. Published by Elsevier Ltd.

  9. CHO-S antibody titers >1 gram/liter using flow electroporation-mediated transient gene expression followed by rapid migration to high-yield stable cell lines.

    Science.gov (United States)

    Steger, Krista; Brady, James; Wang, Weili; Duskin, Meg; Donato, Karen; Peshwa, Madhusudan

    2015-04-01

    In recent years, researchers have turned to transient gene expression (TGE) as an alternative to CHO stable cell line generation for early-stage antibody development. Despite advances in transfection methods and culture optimization, the majority of CHO-based TGE systems produce insufficient antibody titers for extensive use within biotherapeutic development pipelines. Flow electroporation using the MaxCyte STX Scalable Transfection System is a highly efficient, scalable means of CHO-based TGE for gram-level production of antibodies without the need for specialized expression vectors or genetically engineered CHO cell lines. CHO cell flow electroporation is easily scaled from milligram to multigram quantities without protocol reoptimization while maintaining transfection performance and antibody productivity. In this article, data are presented that demonstrate the reproducibility, scalability, and antibody production capabilities of CHO-based TGE using the MaxCyte STX. Data show optimization of posttransfection parameters such as cell density, media composition, and feed strategy that result in secreted antibody titers >1 g/L and production of multiple grams of antibody within 2 weeks of a single CHO-S cell transfection. In addition, data are presented to demonstrate the application of scalable electroporation for the rapid generation of high-yield stable CHO cell lines to bridge the gap between early- and late-stage antibody development activities. © 2014 Society for Laboratory Automation and Screening.

  10. The effects of milking frequency in early lactation on milk yield, mammary cell turnover, and secretory activity in grazing dairy cows.

    Science.gov (United States)

    Murney, R; Stelwagen, K; Wheeler, T T; Margerison, J K; Singh, K

    2015-01-01

    In dairy cows, short-term changes of milking frequency in early lactation have been shown to produce an immediate and a long-term effect on milk yield in stall-fed cows. The effect is controlled locally within mammary glands and could be a function of either secretory mammary epithelial cell number or activity. To resolve this and determine its applicability in other feed management systems, a unilateral milking frequency experiment was conducted with udder halves of 17 multiparous, pasture-fed dairy cows milked either 4 times (4×) or once a day (1×) for 14d from 5±2d in milk. Mean half-udder milk yield during the treatment period was higher from the 4× compared with 1× udder halves and continued to be higher until 200d in milk once returned to twice a day milking. Mammary biopsies were obtained on d 14 of treatment from both udder halves of 10 cows. Proliferation of mammary cells was higher in 4× udder halves compared with 1×, whereas no difference in apoptosis levels was detected. Abundance of αS1-casein, β-casein, α-lactalbumin, and β-lactoglobulin mRNA was higher in tissue samples from 4× udder halves compared with 1×, whereas lactoferrin mRNA abundance was lower in 4× udder halves. In summary, change in milking frequency during early lactation affects proliferation of mammary cells as well as expression of the major milk protein genes, which both contribute to the observed changes in milk yield during and after unilateral milking frequency treatment. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  11. Rapid Changes in Cell Wall Yielding of Elongating Begonia argenteo-guttata L. Leaves in Response to Changes in Plant Water Status.

    Science.gov (United States)

    Serpe, M D; Matthews, M A

    1992-12-01

    Elongation and epidermal cell turgor (P) of Begonia argenteoguttata L. leaves were simultaneously measured to determine the wall-yielding behavior of growing leaf cells in response to changes in plant water status. Rapid changes in plant water status were imposed by irrigating the rooting media with solutions of -0.20 and -0.30 MPa mannitol. These treatments caused decreases in P of 0.09 and 0.17 MPa, respectively. The decreases in P were complete within 10 min, and P did not change thereafter. Following treatments, leaf elongation was nil for periods of 25 to 38 min. Subsequently, elongation recovered to steady rates that were 45 or 75% lower than in the well-watered controls. Leaves of plants that were pretreated with -0.30 MPa of mannitol and rewatered showed an increase in P of 0.19 MPa, which was complete within 15 min; P did not change thereafter. Rewatering caused a several-fold increase in leaf elongation rates, which subsequently declined while P was increasing, to reach steady rates similar to that of the controls. Several estimates of elastic deformation indicated that most of the elongation responses to altered P were due to changes in irreversible deformation. The results showed that the initial effects of changes in P on leaf elongation were partially compensated for by changes in the cell wall-yielding properties. We conclude that linear relationships between P and adjusted growth rates are not necessarily indicative of constant wall-yielding properties. Instead, these relationships may reflect the effect of P on wall-loosening processes.

  12. Rapid Changes in Cell Wall Yielding of Elongating Begonia argenteo-guttata L. Leaves in Response to Changes in Plant Water Status 1

    Science.gov (United States)

    Serpe, Marcelo D.; Matthews, Mark A.

    1992-01-01

    Elongation and epidermal cell turgor (P) of Begonia argenteoguttata L. leaves were simultaneously measured to determine the wall-yielding behavior of growing leaf cells in response to changes in plant water status. Rapid changes in plant water status were imposed by irrigating the rooting media with solutions of −0.20 and −0.30 MPa mannitol. These treatments caused decreases in P of 0.09 and 0.17 MPa, respectively. The decreases in P were complete within 10 min, and P did not change thereafter. Following treatments, leaf elongation was nil for periods of 25 to 38 min. Subsequently, elongation recovered to steady rates that were 45 or 75% lower than in the well-watered controls. Leaves of plants that were pretreated with −0.30 MPa of mannitol and rewatered showed an increase in P of 0.19 MPa, which was complete within 15 min; P did not change thereafter. Rewatering caused a several-fold increase in leaf elongation rates, which subsequently declined while P was increasing, to reach steady rates similar to that of the controls. Several estimates of elastic deformation indicated that most of the elongation responses to altered P were due to changes in irreversible deformation. The results showed that the initial effects of changes in P on leaf elongation were partially compensated for by changes in the cell wall-yielding properties. We conclude that linear relationships between P and adjusted growth rates are not necessarily indicative of constant wall-yielding properties. Instead, these relationships may reflect the effect of P on wall-loosening processes. PMID:16653208

  13. PMA-PhyloChip DNA Microarray to Elucidate Viable Microbial Community Structure

    Science.gov (United States)

    Venkateswaran, Kasthuri J.; Stam, Christina N.; Andersen, Gary L.; DeSantis, Todd

    2011-01-01

    Since the Viking missions in the mid-1970s, traditional culture-based methods have been used for microbial enumeration by various NASA programs. Viable microbes are of particular concern for spacecraft cleanliness, for forward contamination of extraterrestrial bodies (proliferation of microbes), and for crew health/safety (viable pathogenic microbes). However, a "true" estimation of viable microbial population and differentiation from their dead cells using the most sensitive molecular methods is a challenge, because of the stability of DNA from dead cells. The goal of this research is to evaluate a rapid and sensitive microbial detection concept that will selectively estimate viable microbes. Nucleic acid amplification approaches such as the polymerase chain reaction (PCR) have shown promise for reducing time to detection for a wide range of applications. The proposed method is based on the use of a fluorescent DNA intercalating agent, propidium monoazide (PMA), which can only penetrate the membrane of dead cells. The PMA-quenched reaction mixtures can be screened, where only the DNA from live cells will be available for subsequent PCR reaction and microarray detection, and be identified as part of the viable microbial community. An additional advantage of the proposed rapid method is that it will detect viable microbes and differentiate from dead cells in only a few hours, as opposed to less comprehensive culture-based assays, which take days to complete. This novel combination approach is called the PMA-Microarray method. DNA intercalating agents such as PMA have previously been used to selectively distinguish between viable and dead bacterial cells. Once in the cell, the dye intercalates with the DNA and, upon photolysis under visible light, produces stable DNA adducts. DNA cross-linked in this way is unavailable for PCR. Environmental samples suspected of containing a mixture of live and dead microbial cells/spores will be treated with PMA, and then incubated

  14. Flow cytometry to estimate the cell disruption yield and biomass release of Chlorella sp. during bead milling

    NARCIS (Netherlands)

    Günerken, Emre; Hondt, D' Els; Eppink, Michel; Elst, Kathy; Wijffels, Rene

    2017-01-01

    A number of visual, chemical and fluorescence-based methods are generally employed for monitoring of algae cell growth, culture health and biomass concentration. These methods are often time-consuming, demand destructive and high volume sampling. Rapid, efficient, cost-effective and automated

  15. Genetic correlations between the cumulative pseudo-survival rate, milk yield, and somatic cell score during lactation in Holstein cattle in Japan using a random regression model.

    Science.gov (United States)

    Sasaki, O; Aihara, M; Nishiura, A; Takeda, H

    2017-09-01

    Trends in genetic correlations between longevity, milk yield, and somatic cell score (SCS) during lactation in cows are difficult to trace. In this study, changes in the genetic correlations between milk yield, SCS, and cumulative pseudo-survival rate (PSR) during lactation were examined, and the effect of milk yield and SCS information on the reliability of estimated breeding value (EBV) of PSR were determined. Test day milk yield, SCS, and PSR records were obtained for Holstein cows in Japan from 2004 to 2013. A random subset of the data was used for the analysis (825 herds, 205,383 cows). This data set was randomly divided into 5 subsets (162-168 herds, 83,389-95,854 cows), and genetic parameters were estimated in each subset independently. Data were analyzed using multiple-trait random regression animal models including either the residual effect for the whole lactation period (H0), the residual effects for 5 lactation stages (H5), or both of these residual effects (HD). Milk yield heritability increased until 310 to 351 d in milk (DIM) and SCS heritability increased until 330 to 344 DIM. Heritability estimates for PSR increased with DIM from 0.00 to 0.05. The genetic correlation between milk yield and SCS increased negatively to under -0.60 at 455 DIM. The genetic correlation between milk yield and PSR increased until 342 to 355 DIM (0.53-0.57). The genetic correlation between the SCS and PSR was -0.82 to -0.83 at around 180 DIM, and decreased to -0.65 to -0.71 at 455 DIM. The reliability of EBV of PSR for sires with 30 or more recorded daughters was 0.17 to 0.45 when the effects of correlated traits were ignored. The maximum reliability of EBV was observed at 257 (H0) or 322 (HD) DIM. When the correlations of PSR with milk yield and SCS were considered, the reliabilities of PSR estimates increased to 0.31-0.76. The genetic parameter estimates of H5 were the same as those for HD. The rank correlation coefficients of the EBV of PSR between H0 and H5 or HD were

  16. Five QTL hotspots for yield in short rotation coppice bioenergy poplar: The Poplar Biomass Loci

    Directory of Open Access Journals (Sweden)

    Harris Nicole

    2009-02-01

    Full Text Available Abstract Background Concern over land use for non-food bioenergy crops requires breeding programmes that focus on producing biomass on the minimum amount of land that is economically-viable. To achieve this, the maximum potential yield per hectare is a key target for improvement. For long lived tree species, such as poplar, this requires an understanding of the traits that contribute to biomass production and their genetic control. An important aspect of this for long lived plants is an understanding of genetic interactions at different developmental stages, i.e. how genes or genetic regions impact on yield over time. Results QTL mapping identified regions of genetic control for biomass yield. We mapped consistent QTL across multiple coppice cycles and identified five robust QTL hotspots on linkage groups III, IV, X, XIV and XIX, calling these 'Poplar Biomass Loci' (PBL 1–5. In total 20% of the variation in final harvest biomass yield was explained by mapped QTL. We also investigated the genetic correlations between yield related traits to identify 'early diagnostic' indicators of yield showing that early biomass was a reasonable predictor of coppice yield and that leaf size, cell number and stem and sylleptic branch number were also valuable traits. Conclusion These findings provide insight into the genetic control of biomass production and correlation to 'early diagnostic' traits determining yield in poplar SRC for bioenergy. QTL hotspots serve as useful targets for directed breeding for improved biomass productivity that may also be relevant across additional poplar hybrids.

  17. Five QTL hotspots for yield in short rotation coppice bioenergy poplar: the Poplar Biomass Loci.

    Science.gov (United States)

    Rae, Anne M; Street, Nathaniel Robert; Robinson, Kathryn Megan; Harris, Nicole; Taylor, Gail

    2009-02-26

    Concern over land use for non-food bioenergy crops requires breeding programmes that focus on producing biomass on the minimum amount of land that is economically-viable. To achieve this, the maximum potential yield per hectare is a key target for improvement. For long lived tree species, such as poplar, this requires an understanding of the traits that contribute to biomass production and their genetic control. An important aspect of this for long lived plants is an understanding of genetic interactions at different developmental stages, i.e. how genes or genetic regions impact on yield over time. QTL mapping identified regions of genetic control for biomass yield. We mapped consistent QTL across multiple coppice cycles and identified five robust QTL hotspots on linkage groups III, IV, X, XIV and XIX, calling these 'Poplar Biomass Loci' (PBL 1-5). In total 20% of the variation in final harvest biomass yield was explained by mapped QTL. We also investigated the genetic correlations between yield related traits to identify 'early diagnostic' indicators of yield showing that early biomass was a reasonable predictor of coppice yield and that leaf size, cell number and stem and sylleptic branch number were also valuable traits. These findings provide insight into the genetic control of biomass production and correlation to 'early diagnostic' traits determining yield in poplar SRC for bioenergy. QTL hotspots serve as useful targets for directed breeding for improved biomass productivity that may also be relevant across additional poplar hybrids.

  18. Fate of viable but non-culturable Listeria monocytogenes in pig manure microcosms

    Directory of Open Access Journals (Sweden)

    Jeremy eDesneux

    2016-03-01

    Full Text Available The fate of two strains of L. monocytogenes and their ability to become viable but non-culturable (VBNC was investigated in microcosms containing piggery effluents (two raw manures and two biologically treated manures stored for two months at 8°C and 20°C. Levels of L. monocytogenes were estimated using the culture method, qPCR, and propidium monoazide treatment combined with qPCR (qPCRPMA. The chemical composition and the microbial community structure of the manures were also analysed. The strains showed similar decline rates and persisted up to 63 days. At day zero, the percentage of VBNC cells among viable cells was higher in raw manures (81.5-94.8% than in treated manures (67.8-79.2%. The changes in their proportion over time depended on the temperature and on the type of effluent: the biggest increase was observed in treated manures at 20°C and the smallest increase in raw manures at 8°C. The chemical parameters had no influence on the behaviour of the strains, but decrease of the persistence of viable cells was associated with an increase in the microbial richness of the manures. This study demonstrated that storing manure altered the culturability of L. monocytogenes, which rapidly entered the VBNC state, and underlines the importance of including VBNC cells when estimating the persistence of the pathogens in farm effluents.

  19. Glioma Surgical Aspirate: A Viable Source of Tumor Tissue for Experimental Research

    Directory of Open Access Journals (Sweden)

    Perry F. Bartlett

    2013-04-01

    Full Text Available Brain cancer research has been hampered by a paucity of viable clinical tissue of sufficient quality and quantity for experimental research. This has driven researchers to rely heavily on long term cultured cells which no longer represent the cancers from which they were derived. Resection of brain tumors, particularly at the interface between normal and tumorigenic tissue, can be carried out using an ultrasonic surgical aspirator (CUSA that deposits liquid (blood and irrigation fluid and resected tissue into a sterile bottle for disposal. To determine the utility of CUSA-derived glioma tissue for experimental research, we collected 48 CUSA specimen bottles from glioma patients and analyzed both the solid tissue fragments and dissociated tumor cells suspended in the liquid waste fraction. We investigated if these fractions would be useful for analyzing tumor heterogeneity, using IHC and multi-parameter flow cytometry; we also assessed culture generation and orthotopic xenograft potential. Both cell sources proved to be an abundant, highly viable source of live tumor cells for cytometric analysis, animal studies and in-vitro studies. Our findings demonstrate that CUSA tissue represents an abundant viable source to conduct experimental research and to carry out diagnostic analyses by flow cytometry or other molecular diagnostic procedures.

  20. Genome-wide association mapping in winter barley for grain yield and culm cell wall polymer content using the high-throughput CoMPP technique.

    Directory of Open Access Journals (Sweden)

    Andrea Bellucci

    Full Text Available A collection of 112 winter barley varieties (Hordeum vulgare L. was grown in the field for two years (2008/09 and 2009/10 in northern Italy and grain and straw yields recorded. In the first year of the trial, a severe attack of barley yellow mosaic virus (BaYMV strongly influenced final performances with an average reduction of ~ 50% for grain and straw harvested in comparison to the second year. The genetic determination (GD for grain yield was 0.49 and 0.70, for the two years respectively, and for straw yield GD was low in 2009 (0.09 and higher in 2010 (0.29. Cell wall polymers in culms were quantified by means of the monoclonal antibodies LM6, LM11, JIM13 and BS-400-3 and the carbohydrate-binding module CBM3a using the high-throughput CoMPP technique. Of these, LM6, which detects arabinan components, showed a relatively high GD in both years and a significantly negative correlation with grain yield (GYLD. Overall, heritability (H2 was calculated for GYLD, LM6 and JIM and resulted to be 0.42, 0.32 and 0.20, respectively. A total of 4,976 SNPs from the 9K iSelect array were used in the study for the analysis of population structure, linkage disequilibrium (LD and genome-wide association study (GWAS. Marker-trait associations (MTA were analyzed for grain yield and cell wall determination by LM6 and JIM13 as these were the traits showing significant correlations between the years. A single QTL for GYLD containing three MTAs was found on chromosome 3H located close to the Hv-eIF4E gene, which is known to regulate resistance to BaYMV. Subsequently the QTL was shown to be tightly linked to rym4, a locus for resistance to the virus. GWAs on arabinans quantified by LM6 resulted in the identification of major QTLs closely located on 3H and hypotheses regarding putative candidate genes were formulated through the study of gene expression levels based on bioinformatics tools.

  1. Generation of High-Yielding Influenza A Viruses in African Green Monkey Kidney (Vero) Cells by Reverse Genetics

    OpenAIRE

    Ozaki, Hiroichi; Govorkova, Elena A.; Li, Chenghong; Xiong, Xiaoping; Webster, Robert G.; Webby, Richard J.

    2004-01-01

    Influenza A viruses are the cause of annual epidemics of human disease with occasional outbreaks of pandemic proportions. The zoonotic nature of the disease and the vast viral reservoirs in the aquatic birds of the world mean that influenza will not easily be eradicated and that vaccines will continue to be needed. Recent technological advances in reverse genetics methods and limitations of the conventional production of vaccines by using eggs have led to a push to develop cell-based strategi...

  2. Microbial fuel cell coupled to biohydrogen reactor: a feasible technology to increase energy yield from cheese whey.

    Science.gov (United States)

    Wenzel, J; Fuentes, L; Cabezas, A; Etchebehere, C

    2017-06-01

    An important pollutant produced during the cheese making process is cheese whey which is a liquid by-product with high content of organic matter, composed mainly by lactose and proteins. Hydrogen can be produced from cheese whey by dark fermentation but, organic matter is not completely removed producing an effluent rich in volatile fatty acids. Here we demonstrate that this effluent can be further used to produce energy in microbial fuel cells. Moreover, current production was not feasible when using raw cheese whey directly to feed the microbial fuel cell. A maximal power density of 439 mW/m 2 was obtained from the reactor effluent which was 1000 times more than when using raw cheese whey as substrate. 16S rRNA gene amplicon sequencing showed that potential electroactive populations (Geobacter, Pseudomonas and Thauera) were enriched on anodes of MFCs fed with reactor effluent while fermentative populations (Clostridium and Lactobacillus) were predominant on the MFC anode fed directly with raw cheese whey. This result was further demonstrated using culture techniques. A total of 45 strains were isolated belonging to 10 different genera including known electrogenic populations like Geobacter (in MFC with reactor effluent) and known fermentative populations like Lactobacillus (in MFC with cheese whey). Our results show that microbial fuel cells are an attractive technology to gain extra energy from cheese whey as a second stage process during raw cheese whey treatment by dark fermentation process.

  3. Prior knowledge transfer across transcriptional data sets and technologies using compositional statistics yields new mislabelled ovarian cell line.

    Science.gov (United States)

    Blayney, Jaine K; Davison, Timothy; McCabe, Nuala; Walker, Steven; Keating, Karen; Delaney, Thomas; Greenan, Caroline; Williams, Alistair R; McCluggage, W Glenn; Capes-Davis, Amanda; Harkin, D Paul; Gourley, Charlie; Kennedy, Richard D

    2016-09-30

    Here, we describe gene expression compositional assignment (GECA), a powerful, yet simple method based on compositional statistics that can validate the transfer of prior knowledge, such as gene lists, into independent data sets, platforms and technologies. Transcriptional profiling has been used to derive gene lists that stratify patients into prognostic molecular subgroups and assess biomarker performance in the pre-clinical setting. Archived public data sets are an invaluable resource for subsequent in silico validation, though their use can lead to data integration issues. We show that GECA can be used without the need for normalising expression levels between data sets and can outperform rank-based correlation methods. To validate GECA, we demonstrate its success in the cross-platform transfer of gene lists in different domains including: bladder cancer staging, tumour site of origin and mislabelled cell lines. We also show its effectiveness in transferring an epithelial ovarian cancer prognostic gene signature across technologies, from a microarray to a next-generation sequencing setting. In a final case study, we predict the tumour site of origin and histopathology of epithelial ovarian cancer cell lines. In particular, we identify and validate the commonly-used cell line OVCAR-5 as non-ovarian, being gastrointestinal in origin. GECA is available as an open-source R package. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  4. Reduced voltage losses yield 10% efficient fullerene free organic solar cells with >1 V open circuit voltages

    KAUST Repository

    Baran, D.

    2016-11-09

    Optimization of the energy levels at the donor-acceptor interface of organic solar cells has driven their efficiencies to above 10%. However, further improvements towards efficiencies comparable with inorganic solar cells remain challenging because of high recombination losses, which empirically limit the open-circuit voltage (Voc) to typically less than 1 V. Here we show that this empirical limit can be overcome using non-fullerene acceptors blended with the low band gap polymer PffBT4T-2DT leading to efficiencies approaching 10% (9.95%). We achieve Voc up to 1.12 V, which corresponds to a loss of only Eg/q - Voc = 0.5 ± 0.01 V between the optical bandgap Eg of the polymer and Voc. This high Voc is shown to be associated with the achievement of remarkably low non-geminate and non-radiative recombination losses in these devices. Suppression of non-radiative recombination implies high external electroluminescence quantum efficiencies which are orders of magnitude higher than those of equivalent devices employing fullerene acceptors. Using the balance between reduced recombination losses and good photocurrent generation efficiencies achieved experimentally as a baseline for simulations of the efficiency potential of organic solar cells, we estimate that efficiencies of up to 20% are achievable if band gaps and fill factors are further optimized. © The Royal Society of Chemistry 2016.

  5. Preliminary findings on the use of pulsatile machine reperfusion of a placenta to improve the cord blood collection yield including primitive hematopoietic stem cell fractions.

    Science.gov (United States)

    Takebe, Naoko; Gage, Fred; Cheng, Xiangfei; Lauw, Marietya I S

    2009-09-01

    Umbilical cord blood (CB) use is limited in most adults since the mean amount of hematopoietic stem cells (HSCs) collected is generally insufficient to engraft successfully. This study demonstrates for the first time the feasibility of pulsatile machine placenta reperfusion (PMPR), which significantly improves CB collection quantity and quality compared to standard collection methods. PMPR was performed on eight delivered placentas up to 39 hours after venipuncture-based collection. PMPR was performed on average for 26 (20-30) minutes, 17 hours after delivery (6.25-39), using perfusate approved for donated organ reperfusion. Both PMPR and conventional collection-derived CB cells were analyzed using immunophenotyping and colony assays. The combination of PMPR and the conventional venipuncture method yielded a mean of 1.5-, 4.9-, 11-, 7.5-, 7.5-, and 7.7-fold increased number of total mononuclear, CD34+, CD34+/CD38-, CD133+, CD133+/CD34+, and CD133+/CD34- cells, respectively, compared to conventional venipuncture alone. CB cells obtained by PMPR alone generally demonstrated a significant increase in percentages of primitive HSC phenotype with equivalent cell viability between PMPR and venipuncture. This article demonstrates for the first time that CB can be collected up to 39 hours after delivery with PMPR while maximizing CB collection including HSCs with primitive phenotypes.

  6. Effect of gradual or abrupt cessation of milking at dry off on milk yield and somatic cell score in the subsequent lactation.

    Science.gov (United States)

    Gott, P N; Rajala-Schultz, P J; Schuenemann, G M; Proudfoot, K L; Hogan, J S

    2017-03-01

    The objective of this study was to assess the effect of milk cessation method (abrupt or gradual) at dry off on milk yield and somatic cell score (SCS) up to 120 d in milk during the subsequent lactation. Data from 428 cows from 8 dairy herds in Ohio were analyzed. Abrupt cessation cows kept the farm's regular milking schedule (2 or 3 times) through dry off and gradual cessation cows were milked once daily for the final week of lactation. Milk yield and SCS were collected using Dairy Herd Improvement Association test-day records. Aseptic quarter milk samples were collected approximately 1 wk before dry off, at dry off, and within 1 wk after calving for bacterial culture to determine the presence of intramammary infections. Overall, milk cessation method was not significantly associated with either milk yield or SCS in early lactation; however, interaction between the milk cessation method and herd was highly significant. Cows producing greater amounts of milk around dry off had significantly higher SCS in the following lactation. Shorter dry periods were significantly associated with decreased milk yield in the following lactation, especially among abruptly dried off cows. Additionally, as expected, several other factors, such as parity of cows and stage of lactation, were significantly associated with both outcomes. No interactions between the milk cessation method and the other explanatory variables in the final models were significant. The results of the current study suggest that higher milk yield at dry off was associated with higher SCS in the following lactation, even though milk cessation method at the end of lactation had a varying effect on test-day milk yield and SCS in different herds during the first 120 d in milk in the following lactation. The specific herd characteristics influencing this could not be identified within this study, warranting further research. The Authors. Published by the Federation of Animal Science Societies and Elsevier Inc. on

  7. Performance of Pd on activated carbon as hydrogen electrode with respect to hydrogen yield in a single cell proton exchange membrane (PEM) water electrolyser

    Energy Technology Data Exchange (ETDEWEB)

    Naga Mahesh, K.; Sarada Prasad, J.; Venkateswer Rao, M.; Himabindu, V. [Centre for Environment, Institute of Science and Technology, Jawaharlal Nehru Technological University Hyderabad, Kukatpally, Hyderabad 500085 (A.P.) (India); Yerramilli, Anjaneyulu [TLGVRC, JSU Box 18739, Jackson State University, Jackson, MS 32917 - 0939 (United States); Raghunathan Rao, P. [Fuel cell section, Heavy Water Division, Bhabha Atomic Research Centre, Trombay, Mumbai - 400 085 (India)

    2009-08-15

    Palladium (Pd) on activated carbon is used as electrocatalyst coated on Nafion 115 membrane as Hydrogen electrode and RuO{sub 2} is coated on other side of membrane used as oxygen electrode. 5 wt% and 10 wt% Pd on activated carbon is prepared as membrane electrode assembly (MEA) and investigated the performance of the same using inhouse prepared 10 cm{sup 2} single cell. The performance of the single cell assembly and the hydrogen yield are reported during electrolysis operation at temperatures 27 C, 45 C and 65 C at 0.1, 0.2, 0.3, 0.4, 0.5 A/cm{sup 2} current densities with respect to voltages. (author)

  8. Probing the importance of clonality: Single cell subcloning of clonally derived CHO cell lines yields widely diverse clones differing in growth, productivity, and product quality.

    Science.gov (United States)

    Ko, Peggy; Misaghi, Shahram; Hu, Zhilan; Zhan, Dejin; Tsukuda, Joni; Yim, Mandy; Sanford, Mark; Shaw, David; Shiratori, Masaru; Snedecor, Brad; Laird, Michael; Shen, Amy

    2017-12-11

    In the past few decades, a large variety of therapeutic antibodies and proteins have been expressed in Chinese hamster ovary (CHO) cells. This mammalian expression system is robust, scalable, relatively inexpensive, and importantly allows for post-translational modifications that are important for some therapeutic proteins. Historically, CHO cell lines were derived from colonies of cells grown in semi-solid or liquid plates using either serum-containing or serum-free media. Current advancements in cell sorting and imaging technologies have allowed for isolating and imaging single cell progenitors at the seeding step, significantly increasing the probability of isolating clonally derived cell lines. However, it is debatable how much population heterogeneity can be eliminated when clonally derived cell lines, originated from a single cell progenitor, are scaled up. To further investigate this phenomenon, we subcloned two different clonally derived (day 0 imaged and visually inspected) cell lines expressing antibody-X. The results showed that when six randomly chosen subclones of each line were evaluated in a production assay, these subclones displayed a range of variation in titer, specific productivity, growth, and product quality attributes. Some subclones displayed variations in transgene copy numbers. Additionally, clonal derivation did not assure stability of the derived cell lines. Our findings show that cell heterogeneity exists in a population even when derived from a single cell progenitor. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 2017. © 2017 American Institute of Chemical Engineers.

  9. Simulations of DSB Yields and Radiation-induced Chromosomal Aberrations in Human Cells Based on the Stochastic Track Structure Induced by HZE Particles

    Science.gov (United States)

    Ponomarev, Artem; Plante, Ianik; George, Kerry; Wu, Honglu

    2014-01-01

    The formation of double-strand breaks (DSBs) and chromosomal aberrations (CAs) is of great importance in radiation research and, specifically, in space applications. We are presenting a new particle track and DNA damage model, in which the particle stochastic track structure is combined with the random walk (RW) structure of chromosomes in a cell nucleus. The motivation for this effort stems from the fact that the model with the RW chromosomes, NASARTI (NASA radiation track image) previously relied on amorphous track structure, while the stochastic track structure model RITRACKS (Relativistic Ion Tracks) was focused on more microscopic targets than the entire genome. We have combined chromosomes simulated by RWs with stochastic track structure, which uses nanoscopic dose calculations performed with the Monte-Carlo simulation by RITRACKS in a voxelized space. The new simulations produce the number of DSBs as function of dose and particle fluence for high-energy particles, including iron, carbon and protons, using voxels of 20 nm dimension. The combined model also calculates yields of radiation-induced CAs and unrejoined chromosome breaks in normal and repair deficient cells. The joined computational model is calibrated using the relative frequencies and distributions of chromosomal aberrations reported in the literature. The model considers fractionated deposition of energy to approximate dose rates of the space flight environment. The joined model also predicts of the yields and sizes of translocations, dicentrics, rings, and more complex-type aberrations formed in the G0/G1 cell cycle phase during the first cell division after irradiation. We found that the main advantage of the joined model is our ability to simulate small doses: 0.05-0.5 Gy. At such low doses, the stochastic track structure proved to be indispensable, as the action of individual delta-rays becomes more important.

  10. Simulations of DSB Yields and Radiation-induced Chromosomal Aberrations in Human Cells Based on the Stochastic Track Structure iIduced by HZE Particles

    Science.gov (United States)

    Ponomarev, Artem; Plante, Ianik; George, Kerry; Wu, Honglu

    2014-01-01

    The formation of double-strand breaks (DSBs) and chromosomal aberrations (CAs) is of great importance in radiation research and, specifically, in space applications. We are presenting a new particle track and DNA damage model, in which the particle stochastic track structure is combined with the random walk (RW) structure of chromosomes in a cell nucleus. The motivation for this effort stems from the fact that the model with the RW chromosomes, NASARTI (NASA radiation track image) previously relied on amorphous track structure, while the stochastic track structure model RITRACKS (Relativistic Ion Tracks) was focused on more microscopic targets than the entire genome. We have combined chromosomes simulated by RWs with stochastic track structure, which uses nanoscopic dose calculations performed with the Monte-Carlo simulation by RITRACKS in a voxelized space. The new simulations produce the number of DSBs as function of dose and particle fluence for high-energy particles, including iron, carbon and protons, using voxels of 20 nm dimension. The combined model also calculates yields of radiation-induced CAs and unrejoined chromosome breaks in normal and repair deficient cells. The joined computational model is calibrated using the relative frequencies and distributions of chromosomal aberrations reported in the literature. The model considers fractionated deposition of energy to approximate dose rates of the space flight environment. The joined model also predicts of the yields and sizes of translocations, dicentrics, rings, and more complex-type aberrations formed in the G0/G1 cell cycle phase during the first cell division after irradiation. We found that the main advantage of the joined model is our ability to simulate small doses: 0.05-0.5 Gy. At such low doses, the stochastic track structure proved to be indispensable, as the action of individual delta-rays becomes more important.

  11. Development of a radiation track structure clustering algorithm for the prediction of DNA DSB yields and radiation induced cell death in Eukaryotic cells.

    Science.gov (United States)

    Douglass, Michael; Bezak, Eva; Penfold, Scott

    2015-04-21

    The preliminary framework of a combined radiobiological model is developed and calibrated in the current work. The model simulates the production of individual cells forming a tumour, the spatial distribution of individual ionization events (using Geant4-DNA) and the stochastic biochemical repair of DNA double strand breaks (DSBs) leading to the prediction of survival or death of individual cells. In the current work, we expand upon a previously developed tumour generation and irradiation model to include a stochastic ionization damage clustering and DNA lesion repair model. The Geant4 code enabled the positions of each ionization event in the cells to be simulated and recorded for analysis. An algorithm was developed to cluster the ionization events in each cell into simple and complex double strand breaks. The two lesion kinetic (TLK) model was then adapted to predict DSB repair kinetics and the resultant cell survival curve. The parameters in the cell survival model were then calibrated using experimental cell survival data of V79 cells after low energy proton irradiation. A monolayer of V79 cells was simulated using the tumour generation code developed previously. The cells were then irradiated by protons with mean energies of 0.76 MeV and 1.9 MeV using a customized version of Geant4. By replicating the experimental parameters of a low energy proton irradiation experiment and calibrating the model with two sets of data, the model is now capable of predicting V79 cell survival after low energy (cell survival probability, the cell survival probability is calculated for each cell in the geometric tumour model developed in the current work. This model uses fundamental measurable microscopic quantities such as genome length rather than macroscopic radiobiological quantities such as alpha/beta ratios. This means that the model can be theoretically used under a wide range of conditions with a single set of input parameters once calibrated for a given cell line.

  12. Strategic Priming with Multiple Antigens Can Yield Memory Cell Phenotypes Optimized for Infection with Mycobacterium tuberculosis: a Computational Study

    Directory of Open Access Journals (Sweden)

    Cordelia eZiraldo

    2016-01-01

    Full Text Available Lack of an effective vaccine results in 9 million new cases of tuberculosis (TB every year and 1.8 million deaths worldwide. Although many infants are vaccinated at birth with BCG (an attenuated M. bovis, this does not prevent infection or development of TB after childhood. Immune responses necessary for prevention of infection or disease are still unknown, making development of effective vaccines against TB challenging. Several new vaccines are ready for human clinical trials, but these trials are difficult and expensive; especially challenging is determining the appropriate cellular response necessary for protection. The magnitude of an immune response is likely key to generating a successful vaccine. Characteristics such as numbers of central memory (CM and effector memory (EM T cells responsive to a diverse set of epitopes are also correlated with protection. Promising vaccines against TB contain mycobacterial subunit antigens (Ag present during both active and latent infection. We hypothesize that protection against different key immunodominant antigens could require a vaccine that produces different levels of EM and CM for each Ag-specific memory population. We created a computational model to explore EM and CM values, and their ratio, within what we term Memory Design Space. Our model captures events involved in T cell priming within lymph nodes and tracks their circulation through blood to peripheral tissues. We used the model to test whether multiple Ag-specific memory cell populations could be generated with distinct locations within Memory Design Space at a specific time point post vaccination. Boosting can further shift memory populations to memory cell ratios unreachable by initial priming events. By strategically varying antigen load, properties of cellular interactions within the LN, and delivery parameters (e.g. number of boosts of multi-subunit vaccines, we can generate multiple Ag-specific memory populations that cover a wide

  13. Strategic Priming with Multiple Antigens can Yield Memory Cell Phenotypes Optimized for Infection with Mycobacterium tuberculosis: A Computational Study.

    Science.gov (United States)

    Ziraldo, Cordelia; Gong, Chang; Kirschner, Denise E; Linderman, Jennifer J

    2015-01-01

    Lack of an effective vaccine results in 9 million new cases of tuberculosis (TB) every year and 1.8 million deaths worldwide. Although many infants are vaccinated at birth with BCG (an attenuated M. bovis), this does not prevent infection or development of TB after childhood. Immune responses necessary for prevention of infection or disease are still unknown, making development of effective vaccines against TB challenging. Several new vaccines are ready for human clinical trials, but these trials are difficult and expensive; especially challenging is determining the appropriate cellular response necessary for protection. The magnitude of an immune response is likely key to generating a successful vaccine. Characteristics such as numbers of central memory (CM) and effector memory (EM) T cells responsive to a diverse set of epitopes are also correlated with protection. Promising vaccines against TB contain mycobacterial subunit antigens (Ag) present during both active and latent infection. We hypothesize that protection against different key immunodominant antigens could require a vaccine that produces different levels of EM and CM for each Ag-specific memory population. We created a computational model to explore EM and CM values, and their ratio, within what we term Memory Design Space. Our model captures events involved in T cell priming within lymph nodes and tracks their circulation through blood to peripheral tissues. We used the model to test whether multiple Ag-specific memory cell populations could be generated with distinct locations within Memory Design Space at a specific time point post vaccination. Boosting can further shift memory populations to memory cell ratios unreachable by initial priming events. By strategically varying antigen load, properties of cellular interactions within the LN, and delivery parameters (e.g., number of boosts) of multi-subunit vaccines, we can generate multiple Ag-specific memory populations that cover a wide range of

  14. Mitochondrial respiration in human viable platelets-Methodology and influence of gender, age and storage

    DEFF Research Database (Denmark)

    Sjövall, Fredrik; Ehinger, Johannes K H; Marelsson, Sigurður E

    2013-01-01

    Studying whole cell preparations with intact mitochondria and respiratory complexes has a clear benefit compared to isolated or disrupted mitochondria due to the dynamic interplay between mitochondria and other cellular compartments. Platelet mitochondria have a potential to serve as a source...... of human viable mitochondria when studying mitochondrial physiology and pathogenic mechanisms, as well as for the diagnostics of mitochondrial diseases. The objective of the present study was to perform a detailed evaluation of platelet mitochondrial respiration using high-resolution respirometry. Further...

  15. Seed priming and sulfur effects on soybean cell membrane stability and yield in saline soil Condicionamento osmótico das sementes e disponibilidade de enxofre na estabilidade da membrana celular e produtividade de soja em solo salino

    National Research Council Canada - National Science Library

    Teymur Khandan Bejandi; Mohammad Sedghi; Raouf Seyed Sharifi; Ali Namvar; Peyman Molaei

    2009-01-01

    The objective of this work was to determine the effects of seed priming and sulfur application on cell membrane characteristics, seedling emergence, chlorophyll content and grain yield of soybean (Glycine max) in saline soil...

  16. Detection of viable Salmonella in lettuce by propidium monoazide real-time PCR.

    Science.gov (United States)

    Liang, Ningjian; Dong, Jin; Luo, Laixin; Li, Yong

    2011-05-01

    Contamination of lettuce by Salmonella has caused serious public health problems. Polymerase chain reaction (PCR) allows rapid detection of pathogenic bacteria in food, but it is inaccurate as it might amplify DNA from dead target cells as well. This study aimed to investigate the stability of DNA of dead Salmonella cells in lettuce and to develop an approach to detecting viable Salmonella in lettuce. Salmonella-free lettuce was inoculated with heat-killed Salmonella Typhimurium cells and stored at 4 °C. Bacterial DNA extracted from the sample was amplified by real-time PCR targeting the invA gene. Our results indicate that DNA from the dead cells remained stable in lettuce for at least 8 d. To overcome this limitation, propidium monoazide (PMA), a dye that can selectively penetrate dead bacterial cells and cross-link their DNA upon light exposure, was combined with real-time PCR. Lettuce samples inoculated with different levels of dead or viable S. Typhimurium cells were treated or untreated with PMA before DNA extraction. Real-time PCR suggests that PMA treatment effectively prevented PCR amplification from as high as 10(8) CFU/g dead S. Typhimurium cells in lettuce. The PMA real-time PCR assay could detect viable Salmonella at as low as 10(2) CFU/mL in pure culture and 10(3) CFU/g in lettuce. With 12-h enrichment, S. Typhimurium of 10(1) CFU/g in lettuce was detectable. In conclusion, the PMA real-time PCR assay provides an alternative to real-time PCR assay for accurate detection of Salmonella in food. © 2011 Institute of Food Technologists®

  17. An advanced PCR method for the specific detection of viable total coliform bacteria in pasteurized milk.

    Science.gov (United States)

    Soejima, Takashi; Minami, Jun-ichi; Yaeshima, Tomoko; Iwatsuki, Keiji

    2012-07-01

    Pasteurized milk is a complex food that contains various inhibitors of polymerase chain reaction (PCR) and may contain a large number of dead bacteria, depending on the milking conditions and environment. Ethidium monoazide bromide (EMA)-PCR is occasionally used to distinguish between viable and dead bacteria in foods other than pasteurized milk. EMA is a DNA-intercalating dye that selectively permeates the compromised cell membranes of dead bacteria and cleaves DNA. Usually, EMA-PCR techniques reduce the detection of dead bacteria by up to 3.5 logs compared with techniques that do not use EMA. However, this difference may still be insufficient to suppress the amplification of DNA from dead Gram-negative bacteria (e.g., total coliform bacteria) if they are present in pasteurized milk in large numbers. Thus, false positives may result. We developed a new method that uses real-time PCR targeting of a long DNA template (16S-23S rRNA gene, principally 2,451 bp) following EMA treatment to completely suppress the amplification of DNA of up to 7 logs (10(7) cells) of dead total coliforms. Furthermore, we found that a low dose of proteinase K (25 U/ml) removed PCR inhibitors and simultaneously increased the signal from viable coliform bacteria. In conclusion, our simple protocol specifically detects viable total coliforms in pasteurized milk at an initial count of ≥1 colony forming unit (CFU)/2.22 ml within 7.5 h of total testing time. This detection limit for viable cells complies with the requirements for the analysis of total coliforms in pasteurized milk set by the Japanese Sanitation Act (which specifies <1 CFU/2.22 ml).

  18. Antigen stimulation of peripheral blood mononuclear cells from Mycobacterium bovis infected cattle yields evidence for a novel gene expression program

    Directory of Open Access Journals (Sweden)

    Zhao Yingdong

    2008-09-01

    Full Text Available Abstract Background Bovine tuberculosis (BTB caused by Mycobacterium bovis continues to cause substantial losses to global agriculture and has significant repercussions for human health. The advent of high throughput genomics has facilitated large scale gene expression analyses that present a novel opportunity for revealing the molecular mechanisms underlying mycobacterial infection. Using this approach, we have previously shown that innate immune genes in peripheral blood mononuclear cells (PBMC from BTB-infected animals are repressed in vivo in the absence of exogenous antigen stimulation. In the present study, we hypothesized that the PBMC from BTB-infected cattle would display a distinct gene expression program resulting from exposure to M. bovis. A functional genomics approach was used to examine the immune response of BTB-infected (n = 6 and healthy control (n = 6 cattle to stimulation with bovine tuberculin (purified protein derivative – PPD-b in vitro. PBMC were harvested before, and at 3 h and 12 h post in vitro stimulation with bovine tuberculin. Gene expression changes were catalogued within each group using a reference hybridization design and a targeted immunospecific cDNA microarray platform (BOTL-5 with 4,800 spot features representing 1,391 genes. Results 250 gene spot features were significantly differentially expressed in BTB-infected animals at 3 h post-stimulation contrasting with only 88 gene spot features in the non-infected control animals (P ≤ 0.05. At 12 h post-stimulation, 56 and 80 gene spot features were differentially expressed in both groups respectively. The results provided evidence of a proinflammatory gene expression profile in PBMC from BTB-infected animals in response to antigen stimulation. Furthermore, a common panel of eighteen genes, including transcription factors were significantly expressed in opposite directions in both groups. Real-time quantitative reverse transcription PCR (qRT-PCR demonstrated

  19. Microfiltration of enzyme treated egg whites for accelerated detection of viable Salmonella.

    Science.gov (United States)

    Ku, Seockmo; Ximenes, Eduardo; Kreke, Thomas; Foster, Kirk; Deering, Amanda J; Ladisch, Michael R

    2016-11-01

    We report detection of egg white within 7 h by concentrating the bacteria using microfiltration through 0.2-μm cutoff polyethersulfone hollow fiber membranes. A combination of enzyme treatment, controlled cross-flow on both sides of the hollow fibers, and media selection were key to controlling membrane fouling so that rapid concentration and the subsequent detection of low numbers of microbial cells were achieved. We leveraged the protective effect of egg white proteins and peptone so that the proteolytic enzymes did not attack the living cells while hydrolyzing the egg white proteins responsible for fouling. The molecular weight of egg white proteins was reduced from about 70 kDa to 15 kDa during hydrolysis. This enabled a 50-fold concentration of the cells when a volume of 525 mL of peptone and egg white, containing 13 CFU of Salmonella, was decreased to a 10 mL volume in 50 min. A 10-min microcentrifugation step further concentrated the viable Salmonella cells by 10×. The final cell recovery exceeded 100%, indicating that microbial growth occurred during the 3-h processing time. The experiments leading to rapid concentration, recovery, and detection provided further insights on the nature of membrane fouling enabling fouling effects to be mitigated. Unlike most membrane processes where protein recovery is the goal, recovery of viable microorganisms for pathogen detection is the key measure of success, with modification of cell-free proteins being both acceptable and required to achieve rapid microfiltration of viable microorganisms. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1464-1471, 2016. © 2016 American Institute of Chemical Engineers.

  20. A direct viable count method for the enumeration of attached bacteria and assessment of biofilm disinfection

    Science.gov (United States)

    Yu, F. P.; Pyle, B. H.; McFeters, G. A.

    1993-01-01

    This report describes the adaptation of an in situ direct viable count (in situ DVC) method in biofilm disinfection studies. The results obtained with this technique were compared to two other enumeration methods, the plate count (PC) and conventional direct viable count (c-DVC). An environmental isolate (Klebsiella pneumoniae Kp1) was used to form biofilms on stainless steel coupons in a stirred batch reactor. The in situ DVC method was applied to directly assess the viability of bacteria in biofilms without disturbing the integrity of the interfacial community. As additional advantages, the results were observed after 4 h instead of the 24 h incubation time required for colony formation and total cell numbers that remained on the substratum were enumerated. Chlorine and monochloramine were used to determine the susceptibilities of attached and planktonic bacteria to disinfection treatment using this novel analytical approach. The planktonic cells in the reactor showed no significant change in susceptibility to disinfectants during the period of biofilm formation. In addition, the attached cells did not reveal any more resistance to disinfection than planktonic cells. The disinfection studies of young biofilms indicated that 0.25 mg/l free chlorine (at pH 7.2) and 1 mg/l monochloramine (at pH 9.0) have comparable disinfection efficiencies at 25 degrees C. Although being a weaker disinfectant, monochloramine was more effective in removing attached bacteria from the substratum than free chlorine. The in situ DVC method always showed at least one log higher viable cell densities than the PC method, suggesting that the in situ DVC method is more efficient in the enumeration of biofilm bacteria. The results also indicated that the in situ DVC method can provide more accurate information regarding the cell numbers and viability of bacteria within biofilms following disinfection.

  1. The search for viable local government system in Nigeria: an ...

    African Journals Online (AJOL)

    The history of the Nigerian local government system has been one long episode of trails and errors aimed at achieving viable local government institution without much success. Local government in the country began its long series of reforms from the colonial period when the colonial government attempted to ...

  2. Comment: Towards a Viable Local Government Structure in Nigeria ...

    African Journals Online (AJOL)

    Local governments are principally established for development at the grassroots and they must be structured in a manner that makes them viable and capable of achieving this purpose. The objective of this comment is to appraise the current local government structure under the Nigerian constitutional framework with a view ...

  3. Cultivation and multiplication of viable axenic Trypanosoma vivax in ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-09-01

    Sep 1, 2009 ... Cultivation and multiplication of viable axenic. Trypanosoma vivax in vitro and in vivo. O. A. Idowu, A. B. Idowu, C. F. Mafiana and S. O. Sam-Wobo*. Parasitology Laboratory, Department of Biological Sciences, University of Agriculture, Abeokuta, Nigeria. Accepted 13 April, 2006. Trypanosoma vivax was ...

  4. Inoculation of cyprinid herpesvirus 3 (CyHV-3) on common carp brain cells-influence of process parameters on virus yield.

    Science.gov (United States)

    Mletzko, A; Amtmann, A; Bergmann, S; Lee, P; Christian, J; Buchholz, R; Becker, A

    2017-08-01

    Research of cyprinid herpesvirus 3 (CyHV-3) is focused on the infection mechanism and disease development in animals using genetic and immunological approaches to improve treatments and diagnostics. In contrast, only few tried to investigate the CyHV-3 replication behaviour in available cell cultures. Whereas, obtaining high virus yields by in vitro replication enables achieving of the mentioned above goals easier and more reliable. The following work presents an attempt to illuminate the KHV replication in common carp brain (CCB) cell cultures from the engineering point of view. The isolate KHV-TP30 was used testing the influence on process parameters, such as multiplicity of infection (MOI), time of infection (TOI) and time of harvest (TOH). Virus concentrations and infectivity at different time points of infection were examined using hydrolyzed probe qPCR (Gilad et al. 2004) and 50% tissue culture infectivity dose (TCID50). The data obtained show that while the amount of the virus DNA remains constant after reaching its maximum, the infectivity of the virus decreases. Thus, especially, TOH can be crucial for generating a high-quality virus stock. Applying optimized parameters improved the infectivity of the harvested virus and reached a robust titre as high as 1.9 × 108 TCID50/mL. To our knowledge, so far, there is no information in the peer-reviewed literature showing comparably high virus titres. Such virus yields not only facilitate conduction of further studies, including stability tests of the virus stock under various supplementation or disinfection trails, but also provide enough virus material to perform more detailed examinations of the infection mechanism.

  5. Genome-wide association analysis to identify genotype × environment interaction for milk protein yield and level of somatic cell score as environmental descriptors in German Holsteins.

    Science.gov (United States)

    Streit, M; Reinhardt, F; Thaller, G; Bennewitz, J

    2013-01-01

    Genotype by environment interaction (G × E) has been widely reported in dairy cattle. If the environment can be measured on a continuous scale, reaction norms can be applied to study G × E. The average herd milk production level has frequently been used as an environmental descriptor because it is influenced by the level of feeding or the feeding regimen. Another important environmental factor is the level of udder health and hygiene, for which the average herd somatic cell count might be a descriptor. In the present study, we conducted a genome-wide association analysis to identify single nucleotide polymorphisms (SNP) that affect intercept and slope of milk protein yield reaction norms when using the average herd test-day solution for somatic cell score as an environmental descriptor. Sire estimates for intercept and slope of the reaction norms were calculated from around 12 million daughter records, using linear reaction norm models. Sires were genotyped for ~54,000 SNP. The sire estimates were used as observations in the association analysis, using 1,797 sires. Significant SNP were confirmed in an independent validation set consisting of 500 sires. A known major gene affecting protein yield was included as a covariable in the statistical model. Sixty (21) SNP were confirmed for intercept with P ≤ 0.01 (P ≤ 0.001) in the validation set, and 28 and 11 SNP, respectively, were confirmed for slope. Most but not all SNP affecting slope also affected intercept. Comparison with an earlier study revealed that SNP affecting slope were, in general, also significant for slope when the environment was modeled by the average herd milk production level, although the two environmental descriptors were poorly correlated. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  6. Histomorphometric evaluation of the neotropical brown brocket deer Mazama gouazoubira testis, with an emphasis on cell population indexes of spermatogenic yield.

    Science.gov (United States)

    Costa, Kyvia Lugate Cardoso; da Matta, Sérgio Luis Pinto; de Lucca Moreira Gomes, Marcos; de Paula, Tarcízio Antônio Rêgo; de Freitas, Karine Moura; de Araújo Resende Carvalho, Fabíola; de Assis Silveira, Juliana; Dolder, Heidi; Chamindrani Mendis-Handagama, S M L

    2011-09-01

    Information on the reproductive biology of neotropical cervids is scarce. Therefore, the aim of this study was to perform biometric, histologic and stereologic analyses of the brown brocket deer Mazama gouazoubira testis, with an emphasis on the intrinsic yield and the Sertoli cell index. Seven adult males kept in captivity were used. The animals were immobilized; anesthetized and testicle fragments were obtained by biopsy incision. The material was fixed, processed and examined by routine histological methods for light microscopy. The average body weight was 17.2kg, from which 0.40% were allocated in gonads and 0.33% in seminiferous tubules, which represented 85.9% of the testis parenchyma. The mean albuginea width and volume were 345.7μm and 3.5mL (5.3% of the testicular weight), respectively. The mean mediastinum volume of both testicles was 1.0mL (1.5% of the testicular weight) and the testicular parenchyma volume corresponded to 93.1% of total testicular weight (64.9g). The seminiferous tubules diameter was 224.4μm, while the epithelium height was 69.6μm. On average, an adult brown brocket deer showed a total of 1418m of seminiferous tubules in both testicles (21.5m per gram of testis). Each stage I seminiferous tubular cross section contained 1.10 type A spermatogonia, 13.4 primary spermatocytes in pre-leptotene/leptotene, 13.7 spermatocytes in pachytene, 48.8 round spermatids and 3.7 Sertoli cells. The general yield of spermatogenesis was 44.7 cells and the Sertoli cell index was 13.2. The qualitative and quantitative description of testicular histology of brown brocket deer help to understand its spermatogenic process and to establish parameters for the reproductive biology of this wild species. Furthermore, the data from the present research will help further studies using other species of Brazilian cervids, especially endangered ones, making an additional effort to the species preservation. Copyright © 2011 Elsevier B.V. All rights reserved.

  7. Permanently Hypoxic Cell Culture Yields Rat Bone Marrow Mesenchymal Cells with Higher Therapeutic Potential in the Treatment of Chronic Myocardial Infarction.

    Science.gov (United States)

    Liu, Yihua; Yang, Xiaoxi; Maureira, Pablo; Falanga, Aude; Marie, Vanessa; Gauchotte, Guillaume; Poussier, Sylvain; Groubatch, Frederique; Marie, Pierre-Yves; Tran, Nguyen

    2017-01-01

    The mismatch between traditional in vitro cell culture conditions and targeted chronic hypoxic myocardial tissue could potentially hamper the therapeutic effects of implanted bone marrow mesenchymal stem cells (BMSCs). This study sought to address (i) the extent of change to BMSC biological characteristics in different in vitro culture conditions and (ii) the effectiveness of permanent hypoxic culture for cell therapy in treating chronic myocardial infarction (MI) in rats. rat BMSCs were harvested and cultured in normoxic (21% O2, n=27) or hypoxic conditions (5% O2, n=27) until Passage 4 (P4). Cell growth tests, flow cytometry, and Bio-Plex assays were conducted to explore variations in the cell proliferation, phenotype, and cytokine expression, respectively. In the in vivo set-up, P3-BMSCs cultured in normoxia (n=6) or hypoxia (n=6) were intramyocardially injected into rat hearts that had previously experienced 1-month-old MI. The impact of cell therapy on cardiac segmental viability and hemodynamic performance was assessed 1 month later by 2-Deoxy-2[18F]fluoro-D-glucose (18F-FDG) positron emission tomography (PET) imaging and pressure-volume catheter, respectively. Additional histomorphological examinations were conducted to evaluate inflammation, fibrosis, and neovascularization. Hypoxic preconditioning significantly enhanced rat BMSC clonogenic potential and proliferation without altering the multipotency. Different profiles of inflammatory, fibrotic, and angiogenic cytokine secretion were also documented, with a marked correlation observed between in vitro and in vivo proangiogenic cytokine expression and tissue neovessels. Hypoxic-preconditioned cells presented a beneficial effect on the myocardial viability of infarct segments and intrinsic contractility. Hypoxic-preconditioned BMSCs were able to benefit myocardial perfusion and contractility, probably by modulating the inflammation and promoting angiogenesis. © 2017 The Author(s). Published by S. Karger AG

  8. Permanently Hypoxic Cell Culture Yields Rat Bone Marrow Mesenchymal Cells with Higher Therapeutic Potential in the Treatment of Chronic Myocardial Infarction

    Directory of Open Access Journals (Sweden)

    Yihua Liu

    2017-11-01

    Full Text Available Background: The mismatch between traditional in vitro cell culture conditions and targeted chronic hypoxic myocardial tissue could potentially hamper the therapeutic effects of implanted bone marrow mesenchymal stem cells (BMSCs. This study sought to address (i the extent of change to BMSC biological characteristics in different in vitro culture conditions and (ii the effectiveness of permanent hypoxic culture for cell therapy in treating chronic myocardial infarction (MI in rats. Methods: rat BMSCs were harvested and cultured in normoxic (21% O2, n=27 or hypoxic conditions (5% O2, n=27 until Passage 4 (P4. Cell growth tests, flow cytometry, and Bio-Plex assays were conducted to explore variations in the cell proliferation, phenotype, and cytokine expression, respectively. In the in vivo set-up, P3-BMSCs cultured in normoxia (n=6 or hypoxia (n=6 were intramyocardially injected into rat hearts that had previously experienced 1-month-old MI. The impact of cell therapy on cardiac segmental viability and hemodynamic performance was assessed 1 month later by 2-Deoxy-2[18F]fluoro-D-glucose (18F-FDG positron emission tomography (PET imaging and pressure-volume catheter, respectively. Additional histomorphological examinations were conducted to evaluate inflammation, fibrosis, and neovascularization. Results: Hypoxic preconditioning significantly enhanced rat BMSC clonogenic potential and proliferation without altering the multipotency. Different profiles of inflammatory, fibrotic, and angiogenic cytokine secretion were also documented, with a marked correlation observed between in vitro and in vivo proangiogenic cytokine expression and tissue neovessels. Hypoxic-preconditioned cells presented a beneficial effect on the myocardial viability of infarct segments and intrinsic contractility. Conclusion: Hypoxic-preconditioned BMSCs were able to benefit myocardial perfusion and contractility, probably by modulating the inflammation and promoting

  9. Molecular approaches for viable bacterial population and transcriptional analyses in a rodent model of dental caries.

    Science.gov (United States)

    Klein, M I; Scott-Anne, K M; Gregoire, S; Rosalen, P L; Koo, H

    2012-10-01

    Culturing methods are the primary approach for microbiological analysis of plaque biofilms in rodent models of dental caries. In this study, we developed strategies for the isolation of DNA and RNA from plaque biofilms formed in vivo to analyse the viable bacterial population and gene expression. Plaque biofilm samples from rats were treated with propidium monoazide to isolate DNA from viable cells, and the purified DNA was used to quantify total bacteria and the Streptococcus mutans population via quantitative polymerase chain reaction (qPCR) and specific primers; the same samples were also analysed by counting colony-forming units (CFU). In parallel, RNA was isolated from plaque-biofilm samples (from the same animals) and used for transcriptional analyses via reverse transcription-qPCR. The viable populations of both S. mutans and total bacteria assessed by qPCR were positively correlated with the CFU data (P  0.8). However, the qPCR data showed higher bacterial cell counts, particularly for total bacteria (vs. CFU). Moreover, S. mutans proportion in the plaque biofilm determined by qPCR analysis showed strong correlation with incidence of smooth-surface caries (P = 0.0022, r = 0.71). The purified RNAs presented high RNA integrity numbers (> 7), which allowed measurement of the expression of genes that are critical for S. mutans virulence (e.g. gtfB and gtfC). Our data show that the viable microbial population and the gene expression can be analysed simultaneously, providing a global assessment of the infectious aspect of dental caries. Our approach could enhance the value of the current rodent model in further understanding the pathophysiology of this disease and facilitating the exploration of novel anti-caries therapies. © 2012 John Wiley & Sons A/S.

  10. Concurrent detection of other respiratory viruses in children shedding viable human respiratory syncytial virus.

    Science.gov (United States)

    Gagliardi, T B; Paula, F E; Iwamoto, M A; Proença-Modena, J L; Santos, A E; Camara, A A; Cervi, M C; Cintra, O A L; Arruda, E

    2013-10-01

    Human respiratory syncytial virus (HRSV) is an important cause of respiratory disease. The majority of studies addressing the importance of virus co-infections to the HRSV-disease have been based on the detection of HRSV by RT-PCR, which may not distinguish current replication from prolonged shedding of remnant RNA from previous HRSV infections. To assess whether co-detections of other common respiratory viruses are associated with increased severity of HRSV illnesses from patients who were shedding viable-HRSV, nasopharyngeal aspirates from children younger than 5 years who sought medical care for respiratory infections in Ribeirão Preto (Brazil) were tested for HRSV by immunofluorescence, RT-PCR and virus isolation in cell culture. All samples with viable-HRSV were tested further by PCR for other respiratory viruses. HRSV-disease severity was assessed by a clinical score scale. A total of 266 samples from 247 children were collected and 111 (42%) were HRSV-positive. HRSV was isolated from 70 (63%), and 52 (74%) of them were positive for at least one additional virus. HRSV-positive diseases were more severe than HRSV-negative ones, but there was no difference in disease severity between patients with viable-HRSV and those HRSV-positives by RT-PCR. Co-detection of other viruses did not correlate with increased disease severity. HRSV isolation in cell culture does not seem to be superior to RT-PCR to distinguish infections associated with HRSV replication in studies of clinical impact of HRSV. A high rate of co-detection of other respiratory viruses was found in samples with viable-HRSV, but this was not associated with more severe HRSV infection. Copyright © 2013 Wiley Periodicals, Inc.

  11. Selective detection of viable seed-borne Acidovorax citrulli by real-time PCR with propidium monoazide

    Science.gov (United States)

    Tian, Qian; Feng, Jian-jun; Hu, Jie; Zhao, Wen-jun

    2016-01-01

    In recent years, use of the DNA-intercalating dye propidium monoazide (PMA) in real-time PCR has been reported as a novel method to detect viable bacteria in different types of samples, such as food, environmental, and microbiological samples. In this study, viable cells of Acidovorax citrulli, the causal agent of bacterial seedling blight and fruit blotch, were selectively detected and differentiated from dead cells by real-time fluorescent polymerase chain reaction amplification after the bacterial solution was treated with the DNA-binding dye PMA. The primers and TaqMan probe were based on the A. citrulli genome (Aave_1909, Gene ID: 4669443) and were highly specific for A. citrulli. The detection threshold of this assay was 103 colony-forming units per mL (CFU/mL) in pure cell suspensions containing viable and dead cells and infected watermelon seeds. Application of this assay enables the selective detection of viable cells of A. citrulli and facilitates monitoring of the pathogen in watermelon and melon seeds. PMID:27739469

  12. Biofilms in Full-Scale Drinking Water Ozone Contactors Contribute Viable Bacteria to Ozonated Water.

    Science.gov (United States)

    Kotlarz, Nadine; Rockey, Nicole; Olson, Terese M; Haig, Sarah-Jane; Sanford, Larry; LiPuma, John J; Raskin, Lutgarde

    2018-02-13

    Concentrations of viable microbial cells were monitored using culture-based and culture-independent methods across multichamber ozone contactors in a full-scale drinking water treatment plant. Membrane-intact and culturable cell concentrations in ozone contactor effluents ranged from 1200 to 3750 cells/mL and from 200 to 3850 colony forming units/mL, respectively. Viable cell concentrations decreased significantly in the first ozone contact chamber, but rose, even as ozone exposure increased, in subsequent chambers. Our results implicate microbial detachment from biofilms on contactor surfaces, and from biomass present within lime softening sediments in a hydraulic dead zone, as a possible reason for increasing cell concentrations in water samples from sequential ozone chambers. Biofilm community structures on baffle walls upstream and downstream from the dead zone were significantly different from each other (p = 0.017). The biofilms downstream of the dead zone contained a significantly (p = 0.036) higher relative abundance of bacteria of the genera Mycobacterium and Legionella than the upstream biofilms. These results have important implications as the effluent from ozone contactors is often treated further in biologically active filters and bacteria in ozonated water continuously seed filter microbial communities.

  13. Survival Strategy of Erwinia amylovora against Copper: Induction of the Viable-but-Nonculturable State

    Science.gov (United States)

    Ordax, Mónica; Marco-Noales, Ester; López, María M.; Biosca, Elena G.

    2006-01-01

    Copper compounds, widely used to control plant-pathogenic bacteria, have traditionally been employed against fire blight, caused by Erwinia amylovora. However, recent studies have shown that some phytopathogenic bacteria enter into the viable-but-nonculturable (VBNC) state in the presence of copper. To determine whether copper kills E. amylovora or induces the VBNC state, a mineral medium without copper or supplemented with 0.005, 0.01, or 0.05 mM Cu2+ was inoculated with 107 CFU/ml of this bacterium and monitored over 9 months. Total and viable cell counts were determined by epifluorescence microscopy using the LIVE/DEAD kit and by flow cytometry with 5-cyano-2,3-ditolyl tetrazolium chloride and SYTO 13. Culturable cells were counted on King's B nonselective solid medium. Changes in the bacterial morphology in the presence of copper were observed by scanning electron microscopy. E. amylovora entered into the VBNC state at all three copper concentrations assayed, much faster when the copper concentration increased. The addition of different agents which complex copper allowed the resuscitation (restoration of culturability) of copper-induced VBNC cells. Finally, copper-induced VBNC cells were virulent only for the first 5 days, while resuscitated cells always regained their pathogenicity on immature fruits over 9 months. These results have shown, for the first time, the induction of the VBNC state in E. amylovora as a survival strategy against copper. PMID:16672494

  14. Determination of viable wine yeast using DNA binding dyes and quantitative PCR.

    Science.gov (United States)

    Andorrà, Imma; Esteve-Zarzoso, Braulio; Guillamón, José M; Mas, Albert

    2010-12-15

    The detection and quantification of wine yeast can be misleading due to under or overestimation of these microorganisms. Underestimation may be caused by variable growing rates of different microorganisms in culture media or the presence of viable but non-cultivable microorganisms. Overestimation may be caused by the lack of discrimination between live and dead microorganisms if quantitative PCR is used to quantify with DNA as the template. However, culture-independent methods that use dyes have been described to remove the DNA from dead cells and then quantify the live microorganisms. Two dyes have been studied in this paper: ethidium monoazide bromide (EMA) and propidium monoazide bromide (PMA). The technique was applied to grape must fermentation and ageing wines. Both dyes presented similar results on yeast monitoring. Membrane cell recovery was necessary when yeasts were originated from ethanol-containing media. When applied to grape must fermentation, differences of up to 1 log unit were seen between the QPCR estimation with or without the dye during the stationary phase. In ageing wines, good agreement was found between plating techniques and QPCR. Most of the viable cells were also culturable and no differences were observed with the methods, except for Zygosaccharomyces bailii and Dekkera bruxellensis where much higher counts were occasionally detected by QPCR. The presence of excess dead cells did not interfere with the quantification of live cells with either of the dyes. Copyright © 2010 Elsevier B.V. All rights reserved.

  15. How Important Is The Resolution Of Atmospheric Data In Calculations Of Spectral Irradiance And Energy Yield For (III-V) Triple-Junction Cells?

    Science.gov (United States)

    Dobbin, A. L.; Lumb, M. P.; Tibbits, T. N. D.

    2010-10-01

    We use the SMARTS 2.9.5 model to simulate hourly spectral irradiance over a 1 year period at two different geographic locations. Hourly, daily, monthly and yearly averages of cloud-screened atmospheric data from the AERONET network were used to generate 4 sets of spectra for each location. A detailed balance model was then used to investigate how differences in the temporal resolution of atmospheric inputs affect calculations of the energy harvest efficiency of a specific III-V multi-junction cell design. The calculations were repeated using a single yearly averaged spectrum. The modelling results show that changing the time resolution of atmospheric data inputs to the SMARTS model does not cause large changes in calculated energy harvest efficiency for the specific cell design used. This suggests that in the absence of high-resolution atmospheric data, efficiency predictions utilizing spectra derived from yearly or monthly averages will yield similar results to those using daily or hourly averaged data. Use of a single yearly averaged spectrum did however result in an increase of nearly 2% in modelled energy harvest efficiency meaning that temporally varying spectra should be used in calculations of energy harvest efficiency and III-V multi-junction tuning rather than annual averages.

  16. Effective lactation yield

    NARCIS (Netherlands)

    Kok, Akke; Middelaar, van C.E.; Engel, B.; Knegsel, van A.T.M.; Hogeveen, H.; Kemp, B.; Boer, de I.J.M.

    2016-01-01

    To compare milk yields between cows or management strategies, lactations are traditionally standardized to 305-d yields. The 305-d yield, however, gives no insight into the combined effect of additional milk yield before calving, decreased milk yield after calving, and a possible shorter calving

  17. Viable group A streptococci in macrophages during acute soft tissue infection.

    Directory of Open Access Journals (Sweden)

    Pontus Thulin

    2006-03-01

    Full Text Available Group A streptococcal severe soft tissue infections, such as necrotizing fasciitis, are rapidly progressive infections associated with high mortality. Group A streptococcus is typically considered an extracellular pathogen, but has been shown to reside intracellularly in host cells.We characterized in vivo interactions between group A streptococci (GAS and cells involved in innate immune responses, using human biopsies (n = 70 collected from 17 patients with soft tissue infections. Immunostaining and in situ image analysis revealed high amounts of bacteria in the biopsies, even in those collected after prolonged antibiotic therapy. Viability of the streptococci was assessed by use of a bacterial viability stain, which demonstrated viable bacteria in 74% of the biopsies. GAS were present both extracellularly and intracellularly within phagocytic cells, primarily within macrophages. Intracellular GAS were predominantly noted in biopsies from newly involved tissue characterized by lower inflammation and bacterial load, whereas purely extracellular GAS or a combination of intra- and extracellular GAS dominated in severely inflamed tissue. The latter tissue was also associated with a significantly increased amount of the cysteine protease streptococcal pyrogenic exotoxin SpeB. In vitro studies confirmed that macrophages serve as reservoirs for viable GAS, and infection with a speB-deletion mutant produced significantly lower frequencies of cells with viable GAS following infection as compared to the wild-type bacteria.This is the first study to demonstrate that GAS survive intracellularly in macrophages during acute invasive infections. This intracellular presence may have evolved as a mechanism to avoid antibiotic eradication, which may explain our finding that high bacterial load is present even in tissue collected after prolonged intravenous antibiotic therapy. This new insight into the pathogenesis of streptococcal soft tissue infections

  18. Viable Group A Streptococci in Macrophages during Acute Soft Tissue Infection.

    Directory of Open Access Journals (Sweden)

    2006-01-01

    Full Text Available BACKGROUND: Group A streptococcal severe soft tissue infections, such as necrotizing fasciitis, are rapidly progressive infections associated with high mortality. Group A streptococcus is typically considered an extracellular pathogen, but has been shown to reside intracellularly in host cells. METHODS AND FINDINGS: We characterized in vivo interactions between group A streptococci (GAS and cells involved in innate immune responses, using human biopsies (n = 70 collected from 17 patients with soft tissue infections. Immunostaining and in situ image analysis revealed high amounts of bacteria in the biopsies, even in those collected after prolonged antibiotic therapy. Viability of the streptococci was assessed by use of a bacterial viability stain, which demonstrated viable bacteria in 74% of the biopsies. GAS were present both extracellularly and intracellularly within phagocytic cells, primarily within macrophages. Intracellular GAS were predominantly noted in biopsies from newly involved tissue characterized by lower inflammation and bacterial load, whereas purely extracellular GAS or a combination of intra- and extracellular GAS dominated in severely inflamed tissue. The latter tissue was also associated with a significantly increased amount of the cysteine protease streptococcal pyrogenic exotoxin SpeB. In vitro studies confirmed that macrophages serve as reservoirs for viable GAS, and infection with a speB-deletion mutant produced significantly lower frequencies of cells with viable GAS following infection as compared to the wild-type bacteria. CONCLUSIONS: This is the first study to demonstrate that GAS survive intracellularly in macrophages during acute invasive infections. This intracellular presence may have evolved as a mechanism to avoid antibiotic eradication, which may explain our finding that high bacterial load is present even in tissue collected after prolonged intravenous antibiotic therapy. This new insight into the pathogenesis

  19. Acupuntura un tratamiento viable para las adicciones en Colombia

    Directory of Open Access Journals (Sweden)

    Hernán López Seuscún

    2013-07-01

    Los tratamientos con auriculoterapia, como el protocolo NADA (National Acupuncture Detoxification Association, son los métodos más usados para las adicciones en el mundo, y aunque no se ha logrado evidenciar su efectividad, por su costo, facilidad y el poco riesgo de efectos adversos se hace viable en un país con pocos recursos económicos como Colombia.

  20. Academic Pediatric Dentistry is a Rewarding, Financially Viable Career Path.

    Science.gov (United States)

    Townsend, Janice A; Chi, Donald L

    2017-09-15

    Newly graduated pediatric dentists have unprecedented levels of debt. High levels of student debt may be perceived as an obstacle to pursue an academic career. However, opportunities exist through faculty compensation models and loan repayment programs that make an academic career financially viable. The purpose of this paper is to outline the benefits of a career in academic dentistry and provide examples of young pediatric dentistry faculty members who have been able to manage student debt while pursuing meaningful and rewarding careers.

  1. How Can We Prevent Violence Becoming a Viable Political Strategy?

    OpenAIRE

    Patricia Justino

    2009-01-01

    A basic issue that conflict analysis investigates is how non-peaceful ways of living and governing become viable political strategies. Macro-level studies provide some important insights but micro-level analysis is vital to understand the mechanisms that make violence possible. This briefing outlines some preliminary findings in this respect from MICROCON, a major research programme analysing violent conflict at the micro level. It also discusses their implications for policies aimed at preve...

  2. Donor body mass index is an important factor that affects peripheral blood progenitor cell yield in healthy donors after mobilization with granulocyte-colony-stimulating factor.

    Science.gov (United States)

    Chen, Jian; Burns, Kevin M; Babic, Aleksandar; Carrum, George; Kennedy, Martha; Segura, Francisco J; Garcia, Salvador; Potts, Sandra; Leveque, Christopher

    2014-01-01

    The use of hematopoietic progenitor cell (HPC) transplantation has rapidly expanded in recent years. Currently, several sources of HPCs are available for transplantation including peripheral blood HPCs (PBPCs), cord blood cells, and marrow cells. Of these, PBPC collection has become the major source of HPCs. An important variable in PBPC collection is the response to PBPC mobilization, which varies significantly and sometime causes mobilization failure. A retrospective study of 69 healthy donors who underwent PBPC donation by leukapheresis was performed. All of these donors received 10 μg/kg/day or more granulocyte-colony-stimulating factor (G-CSF) for 5 days before PBPC harvest. Donor factors were evaluated and correlated with mobilization responses, as indicated by the precollection CD34 count (pre-CD34). Donors with a pre-CD34 of more than 100 × 10(6) /L had higher body mass index (BMI) compared with donors whose pre-CD34 was 38 × 10(6) to 99 × 10(6) /L or less than 38 × 10(6) /L (32.0 ± 1.04 kg/m(2) vs. 28.7 ± 0.93 kg/m(2) vs. 25.9 ± 1.27 kg/m(2) , respectively; p donors with high BMIs had higher pre-CD34 on a per-kilogram-of-body-weight basis compared with donors with low BMIs. BMI is an important factor that affects donor's response to mobilization and consequently the HPC yield. This effect may be due to a relatively high dose of G-CSF administered to donors with higher BMI or due to the presence of unknown intrinsic factors affecting mobilization that correlate with the amount of adipose tissue in each donor. © 2013 American Association of Blood Banks.

  3. Model based analysis of transient fluorescence yield induced by actinic laser flashes in spinach leaves and cells of green alga Chlorella pyrenoidosa Chick.

    Science.gov (United States)

    Belyaeva, N E; Schmitt, F-J; Paschenko, V Z; Riznichenko, G Yu; Rubin, A B; Renger, G

    2014-04-01

    Measurements of Single Flash Induced Transient Fluorescence Yield (SFITFY) on spinach leaves and whole cells of green thermophilic alga Chlorella pyrenoidosa Chick were analyzed for electron transfer (ET) steps and coupled proton transfer (PT) on both the donor and the acceptor side of the reaction center (RC) of photosystem II (PS II). A specially developed PS II model (Belyaeva et al., 2008, 2011a) allowed the determination of ET steps that occur in a hierarchically ordered time scale from nanoseconds to several seconds. Our study demonstrates that our SFITFY data is consistent with the concept of the reduction of P680(+) by YZ in both leaves and algae (studied on spinach leaves and cells of Chlorella pyrenoidosa Chick). The multiphasic P680(+) reduction kinetics by YZ in PS II core complexes with high oxygen evolution capacity was seen in both algae and leaves. Model simulation to fit SFITFY curves for dark adapted species used here gives the rate constants to verify nanosecond kinetic stages of P680(+) reduction by YZ in the redox state S1 of the water oxidizing complex (WOC) shown in Kühn et al. (2004). Then a sequence of relaxation steps in the redox state S1, outlined by Renger (2012), occurs in both algae and leaves as a similar non-adiabatic ET reactions. Coupled PT is discussed briefly to understand a rearrangement of hydrogen bond protons in the protein matrix of the WOC (Umena et al., 2011). On the other hand, present studies showed a slower reoxidation of reduced QA by QB in algal cells as compared with that in a leaf that might be regarded as a consequence of differences of spatial domains at the QB-site in leaves compared to algae. Our comparative study helped to correlate theory with experimental data for molecular photosynthetic mechanisms in thylakoid membranes. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  4. Mice carrying a complete deletion of the talin2 coding sequence are viable and fertile

    Energy Technology Data Exchange (ETDEWEB)

    Debrand, Emmanuel; Conti, Francesco J.; Bate, Neil; Spence, Lorraine; Mazzeo, Daniela; Pritchard, Catrin A.; Monkley, Susan J. [Department of Biochemistry, University of Leicester, Lancaster Road, Leicester LE1 9HN (United Kingdom); Critchley, David R., E-mail: drc@le.ac.uk [Department of Biochemistry, University of Leicester, Lancaster Road, Leicester LE1 9HN (United Kingdom)

    2012-09-21

    Highlights: Black-Right-Pointing-Pointer Mice lacking talin2 are viable and fertile with only a mildly dystrophic phenotype. Black-Right-Pointing-Pointer Talin2 null fibroblasts show no major defects in proliferation, adhesion or migration. Black-Right-Pointing-Pointer Maintaining a colony of talin2 null mice is difficult indicating an underlying defect. -- Abstract: Mice homozygous for several Tln2 gene targeted alleles are viable and fertile. Here we show that although the expression of talin2 protein is drastically reduced in muscle from these mice, other tissues continue to express talin2 albeit at reduced levels. We therefore generated a Tln2 allele lacking the entire coding sequence (Tln2{sup cd}). Tln2{sup cd/cd} mice were viable and fertile, and the genotypes of Tln2{sup cd/+} intercrosses were at the expected Mendelian ratio. Tln2{sup cd/cd} mice showed no major difference in body mass or the weight of the major organs compared to wild-type, although they displayed a mildly dystrophic phenotype. Moreover, Tln2{sup cd/cd} mouse embryo fibroblasts showed no obvious defects in cell adhesion, migration or proliferation. However, the number of Tln2{sup cd/cd} pups surviving to adulthood was variable suggesting that such mice have an underlying defect.

  5. Test-day somatic cell score, fat-to-protein ratio and milk yield as indicator traits for sub-clinical mastitis in dairy cattle.

    Science.gov (United States)

    Jamrozik, J; Schaeffer, L R

    2012-02-01

    Test-day (TD) records of milk, fat-to-protein ratio (F:P) and somatic cell score (SCS) of first-lactation Canadian Holstein cows were analysed by a three-trait finite mixture random regression model, with the purpose of revealing hidden structures in the data owing to putative, sub-clinical mastitis. Different distributions of the data were allowed in 30 intervals of days in milk (DIM), covering the lactation from 5 to 305 days. Bayesian analysis with Gibbs sampling was used for model inferences. Estimated proportion of TD records originated from cows infected with mastitis was 0.66 in DIM from 5 to 15 and averaged 0.2 in the remaining part of lactation. Data from healthy and mastitic cows exhibited markedly different distributions, with respect to both average value and the variance, across all parts of lactation. Heterogeneity of distributions for infected cows was also apparent in different DIM intervals. Cows with mastitis were characterized by smaller milk yield (down to -5 kg) and larger F:P (up to 0.13) and SCS (up to 1.3) compared with healthy contemporaries. Differences in averages between healthy and infected cows for F:P were the most profound at the beginning of lactation, when a dairy cow suffers the strongest energy deficit and is therefore more prone to mammary infection. Residual variances for data from infected cows were substantially larger than for the other mixture components. Fat-to-protein ratio had a significant genetic component, with estimates of heritability that were larger or comparable with milk yield, and was not strongly correlated with milk and SCS on both genetic and environmental scales. Daily milk, F:P and SCS are easily available from milk-recording data for most breeding schemes in dairy cattle. Fat-to-protein ratio can potentially be a valuable addition to SCS and milk yield as an indicator trait for selection against mastitis. © 2011 Blackwell Verlag GmbH.

  6. Sorption and precipitation of Mn2+ by viable and autoclaved Shewanella putrefaciens: Effect of contact time

    KAUST Repository

    Chubar, Natalia

    2013-01-01

    The sorption of Mn(II) by viable and inactivated cells of Shewanella putrefaciens, a non-pathogenic, facultative anaerobic, gram-negative bacterium characterised as a Mn(IV) and Fe(III) reducer, was studied under aerobic conditions, as a function of pH, bacterial density and metal loading. During a short contact time (3-24h), the adsorptive behaviour of live and dead bacteria toward Mn(II) was sufficiently similar, an observation that was reflected in the studies on adsorption kinetics at various metal loadings, effects of pH, bacteria density, isotherms and drifting of pH during adsorption. Continuing the experiment for an additional 2-30days demonstrated that the Mn(II) sorption by suspensions of viable and autoclaved cells differed significantly from one another. The sorption to dead cells was characterised by a rapid equilibration and was described by an isotherm. In contrast, the sorption (uptake) to live bacteria exhibited a complex time-dependent uptake. This uptake began as adsorption and ion exchange processes followed by bioprecipitation, and it was accompanied by the formation of polymeric sugars (EPS) and the release of dissolved organic substances. FTIR, EXAFS/XANES and XPS demonstrated that manganese(II) phosphate was the main precipitate formed in 125ml batches, which is the first evidence of the ability of microbes to synthesise manganese phosphates. XPS and XANES spectra did not detect Mn(II) oxidation. Although the release of protein-like compounds by the viable bacteria increased in the presence of Mn2+ (and, by contrast, the release of carbohydrates did not change), electrochemical analyses did not indicate any aqueous complexation of Mn(II) by the organic ligands. © 2012 Elsevier Ltd.

  7. Viable Reserve Networks Arise From Individual Landholder Responses To Conservation Incentives

    Directory of Open Access Journals (Sweden)

    Kenneth M. Chomitz

    2006-12-01

    Full Text Available Conservation in densely settled biodiversity hotspots often requires setting up reserve networks that maintain sufficient contiguous habitat to support viable species populations. Because it is difficult to secure landholder compliance with a tightly constrained reserve network design, attention has shifted to voluntary incentive mechanisms, such as purchase of conservation easements by reverse auction or through a fixed-price offer. These mechanisms carry potential advantages of transparency, simplicity, and low cost. However, uncoordinated individual response to these incentives has been assumed incompatible with the conservation goal of viability, which depends on contiguous habitat and biodiversity representation. We model such incentives for southern Bahia in the Brazilian Atlantic Forest, one of the biologically richest and most threatened global biodiversity hotspots. Here, forest cover is spatially autocorrelated and associated with depressed land values, a situation that may be characteristic of long-settled areas with forests fragmented by agriculture. We find that in this situation, a voluntary incentive system can yield a reserve network characterized by large, viable patches of contiguous forest, and representation of subregions with distinct vegetation types and biotic assemblages, without explicit planning for those outcomes.

  8. Liquid biopsy in cancer patients: advances in capturing viable CTCs for functional studies using the EPISPOT assay.

    Science.gov (United States)

    Alix-Panabières, Catherine; Pantel, Klaus

    2015-01-01

    Circulating tumor cells (CTCs) in the blood of cancer patients have received increasing attention as new diagnostic tool enabling 'liquid biopsies'. In contrast to the wealth of descriptive studies demonstrating the clinical relevance of CTCs as biomarkers, the extremely low concentration of CTCs in the peripheral blood of most cancer patients challenges further functional studies. This article discusses the current possibilities to enrich and, in particular, detect viable CTCs with emphasis on the EPithelial ImmunoSPOT technology. This functional assay detects viable CTCs at the single-cell level and has been used on hundreds of patients with different tumor types including epithelial tumors (breast, prostate and colon cancer) and melanomas. Moreover, the article summarizes recent advances in the in vitro and in vivo expansion of CTCs from cancer patients. These functional analyses will contribute to identifying the biological properties of metastatic cells and reveal new therapeutic targets against disseminating cancer cells.

  9. Comparative 2D-DIGE Proteomic Analysis of Bovine Mammary Epithelial Cells during Lactation Reveals Protein Signatures for Lactation Persistency and Milk Yield

    Science.gov (United States)

    Janjanam, Jagadeesh; Singh, Surender; Jena, Manoj K.; Varshney, Nishant; Kola, Srujana; Kumar, Sudarshan; Kaushik, Jai K.; Grover, Sunita; Dang, Ajay K.; Mukesh, Manishi; Prakash, B. S.; Mohanty, Ashok K.

    2014-01-01

    Mammary gland is made up of a branching network of ducts that end with alveoli which surrounds the lumen. These alveolar mammary epithelial cells (MEC) reflect the milk producing ability of farm animals. In this study, we have used 2D-DIGE and mass spectrometry to identify the protein changes in MEC during immediate early, peak and late stages of lactation and also compared differentially expressed proteins in MEC isolated from milk of high and low milk producing cows. We have identified 41 differentially expressed proteins during lactation stages and 22 proteins in high and low milk yielding cows. Bioinformatics analysis showed that a majority of the differentially expressed proteins are associated in metabolic process, catalytic and binding activity. The differentially expressed proteins were mapped to the available biological pathways and networks involved in lactation. The proteins up-regulated during late stage of lactation are associated with NF-κB stress induced signaling pathways and whereas Akt, PI3K and p38/MAPK signaling pathways are associated with high milk production mediated through insulin hormone signaling. PMID:25111801

  10. Surface heterogeneity on hemispheres-in-cell model yields all experimentally-observed non-straining colloid retention mechanisms in porous media in the presence of energy barriers.

    Science.gov (United States)

    Ma, Huilian; Pazmino, Eddy; Johnson, William P

    2011-12-20

    Many mechanisms of colloid retention in porous media under unfavorable conditions have been identified from experiments or theory, such as attachment at surface heterogeneities, wedging at grain to grain contacts, retention via secondary energy minimum association in zones of low flow drag, and straining in pore throats too small to pass. However, no previously published model is capable of representing all of these mechanisms of colloid retention. In this work, we demonstrate that incorporation of surface heterogeneity into our hemispheres-in-cell model yields all experimentally observed non-straining retention mechanisms in porous media under unfavorable conditions. We also demonstrate that the predominance of any given retention mechanism depends on the coupled colloid-collector-flow interactions that are governed by parameters such as the size and spatial frequency of heterogeneous attractive domains, colloid size, and solution ionic strength. The force/torque balance-simulated retention is shown to decrease gradually with decreasing solution ionic strength, in agreement with experimental observations. This gradual decrease stands in sharp contrast to predictions from mean field theory that does not account for discrete surface heterogeneity. © 2011 American Chemical Society

  11. Random insertion of split-cans of the fluorescent protein venus into Shaker channels yields voltage sensitive probes with improved membrane localization in mammalian cells.

    Science.gov (United States)

    Jin, Lei; Baker, Bradley; Mealer, Robbie; Cohen, Lawrence; Pieribone, Vincent; Pralle, Arnd; Hughes, Thomas

    2011-07-15

    FlaSh-YFP, a fluorescent protein (FP) voltage sensor that is a fusion of the Shaker potassium channel with yellow fluorescent protein (YFP), is primarily expressed in the endoplasmic reticulum (ER) of mammalian cells, possibly due to misfolded monomers. In an effort to improve plasma membrane expression, the FP was split into two non-fluorescent halves. Each half was randomly inserted into Shaker monomers via a transposon reaction. Shaker subunits containing the 5' half were co-expressed with Shaker subunits containing the 3' half. Tetramerization of Shaker subunits is required for re-conjugation of the FP. The misfolded monomers trapped in ER are unlikely to tetramerize and reconstitute the beta-can structure, and thus intracellular fluorescence might be reduced. This split-can transposon approach yielded 56 fluorescent probes, 30 (54%) of which were expressed at the plasma membrane and were capable of optically reporting changes in membrane potential. The largest signal from these novel FP-sensors was a -1.4% in ΔF/F for a 100 mV depolarization, with on time constants of about 15 ms and off time constants of about 200 ms. This split-can transposon approach has the potential to improve other multimeric probes. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Effect of chosen factors on milk yield, basic composition and somatic cell count of organic milk of Brown short-haired goats

    Directory of Open Access Journals (Sweden)

    Šárka Králíčková

    2013-01-01

    Full Text Available Effect of stage of lactation (SL, parity (PA, litter size (LS and month of kidding (MK on daily milk yield, basic composition and somatic cell count of organic goat milk were evaluated using 32 goats of Brown short-haired breed which were reared on an organic farm in Olešenka. Goats were on the 1st (n = 15, 2nd (n = 10 and 3rd (n = 7 lactation; 15 of them gave birth to singles and 17 to twins. Kidding occurred in January (n = 5, February (n = 13 and March (n = 8. During the experiment all goats were reared on permanent pasture, in one flock and under the identical conditions. The PA had a significant effect on all monitored parameters; also the SL had a significant effect on all monitored parameters except SCC. Systematic factors LS and MK had a significant effect only on DMY and SCC; on the other hand, these factors had no significant effect on basic milk composition. The milk fat was the most variable component of the milk. On the other hand the lactose content was during lactation very well-balanced. SCCs were relatively low which indicates good health status of mammary gland. The highest DMY, SCC and contents of basic milk components, except lactose, were found in goats on PA2. The higher DMY was expected in goats with twins compared to goats with singles, but the opposite trend proved to be true.

  13. Modulation of the endocannabinoid system in viable and non-viable first trimester pregnancies by pregnancy-related hormones

    Directory of Open Access Journals (Sweden)

    Taylor Anthony H

    2011-11-01

    Full Text Available Abstract Background In early pregnancy, increased plasma levels of the endocannabinoid anandamide (AEA are associated with miscarriage through mechanisms that might affect the developing placenta or maternal decidua. Methods In this study, we compare AEA levels in failed and viable pregnancies with the levels of the trophoblastic hormones (beta-human chorionic gonadotrophin (beta-hCG, progesterone (P4 and (pregnancy-associated placental protein-A (PAPP-A essential for early pregnancy success and relate that to the expression of the cannabinoid receptors and enzymes that modulate AEA levels. Results The median plasma AEA level in non-viable pregnancies (1.48 nM; n = 20 was higher than in viable pregnancies (1.21 nM; n = 25; P = 0.013, as were progesterone and beta-hCG levels (41.0 vs 51.5 ng/mL; P = 0.052 for P4 and 28,650 vs 6,560 mIU/L; P = 0.144 for beta-hCG, respectively, but were not statistically significant. Serum PAPP-A levels in the viable group were approximately 6.8 times lower than those in the non-viable group (1.82 vs 12.25 mg/L; P = 0.071, but again these differences were statistically insignificant. In the spontaneous miscarriage group, significant correlations between P4 and beta-hCG, P4 and PAPP-A and AEA and PAPP-A levels were observed. Simultaneously, immunohistochemical distributions of the two main cannabinoid receptors and the AEA-modifying enzymes, fatty acid amide hydrolase (FAAH and N-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD, changed within both the decidua and trophoblast. Conclusions The association of higher AEA levels with early pregnancy failure and with beta-hCG and PAPP-A, but not with progesterone concentrations suggest that plasma AEA levels and pregnancy failure are linked via a mechanism that may involve trophoblastic beta-hCG, and PAPP-A, but not, progesterone production. Although the trophoblast, decidua and embryo contain receptors for AEA, the main AEA target in early pregnancy failure

  14. The viable but nonculturable state induction and genomic analyses of Lactobacillus casei BM-LC14617, a beer-spoilage bacterium.

    Science.gov (United States)

    Liu, Junyan; Li, Lin; Peters, Brian M; Li, Bing; Chen, Lequn; Deng, Yang; Xu, Zhenbo; Shirtliff, Mark E

    2017-10-01

    This study aimed to investigate the viable but nonculturable (VBNC) state and genomic features of a beer-spoilage strain, Lactobacillus caseiBM-LC14617. Induction on the VBNC state of L. casei strain BM-LC14617 was conducted by both low-temperature storage and continuous passage in beer, and formation of VBNC state was detected after 196 ± 3.3 days and 32 ± 1.6 subcultures, respectively. Resuscitation of VBNC cells was successfully induced by addition of catalase, and culturable, VBNC, and resuscitated cells shared similar beer-spoilage capability. Whole genome sequencing was performed, and out of a total of 3,964 predicted genes, several potential VBNC and beer-spoilage-associated genes were identified. L. casei is capable of entering into and resuscitating from the VBNC state and possesses beer-spoilage capability. The genomic characterization yield insightful elucidation of VBNC state for L. casei. This study represents the first evidence on VBNC state induction of L. casei and beer-spoilage capability of VBNC and resuscitated cells. Also, this is the first genomic characterization of L. casei as a beer-spoilage bacterium. The current study may aid in further study on L. casei and other beer-spoilage bacteria, and guide the prevention and control of beer spoilage. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  15. Removal of viable bioaerosol particles with a low-efficiency HVAC filter enhanced by continuous emission of unipolar air ions.

    Science.gov (United States)

    Huang, R; Agranovski, I; Pyankov, O; Grinshpun, S

    2008-04-01

    Continuous emission of unipolar ions has been shown to improve the performance of respirators and stationary filters challenged with non-biological particles. In this study, we investigated the ion-induced enhancement effect while challenging a low-efficiency heating, ventilation and air-conditioning (HVAC) filter with viable bacterial cells, bacterial and fungal spores, and viruses. The aerosol concentration was measured in real time. Samples were also collected with a bioaerosol sampler for viable microbial analysis. The removal efficiency of the filter was determined, respectively, with and without an ion emitter. The ionization was found to significantly enhance the filter efficiency in removing viable biological particles from the airflow. For example, when challenged with viable bacteria, the filter efficiency increased as much as four- to fivefold. For viable fungal spores, the ion-induced enhancement improved the efficiency by a factor of approximately 2. When testing with virus-carrying liquid droplets, the original removal efficiency provided by the filter was rather low: 9.09 +/- 4.84%. While the ion emission increased collection about fourfold, the efficiency did not reach 75-100% observed with bacteria and fungi. These findings, together with our previously published results for non-biological particles, demonstrate the feasibility of a new approach for reducing aerosol particles in HVAC systems used for indoor air quality control. Recirculated air in HVAC systems used for indoor air quality control in buildings often contains considerable number of viable bioaerosol particles because of limited efficiency of the filters installed in these systems. In the present study, we investigated - using aerosolized bacterial cells, bacterial and fungal spores, and virus-carrying particles - a novel idea of enhancing the performance of a low-efficiency HVAC filter utilizing continuous emission of unipolar ions in the filter vicinity. The findings described in

  16. Molar Pregnancy with a Co-Existing Viable Fetus

    Directory of Open Access Journals (Sweden)

    Ruya Deveer

    2014-03-01

    Full Text Available     The aim of this study was to report the clinical features, management, and outcome of a case of molar pregnancy with a coexisting viable fetus and to review the literature. In this article, we report a case of pregnancy with diffuse placental molar change and a normal fetus which presented with hyperemesis gravidarum and hyperthyroidism. Genetic amniocentesis showed normal fetal karyotype. A healthy full-term live male infant was delivered by cesarean section. In molar pregnancies with a normal karyotype fetus, with intensive maternal follow-up, continuation of pregnancy can be suggested.

  17. Use of sodium lauroyl sarcosinate (sarkosyl) in viable real-time PCR for enumeration of Escherichia coli.

    Science.gov (United States)

    Wang, Hui; Gill, Colin O; Yang, Xianqin

    2014-03-01

    The cell membranes of inactivated Escherichia coli are not always permeable to propidium monoazide (PMA). This limits the use of PMA real-time PCR (PMA-qPCR) for quantification of DNA from only viable cells for enumeration of E. coli. The aim of this study was to develop PMA-qPCR procedures for E. coli with improved selectivity for viable cells. E. coli inactivated by incubation at 52°C were treated with 12 detergents before PMA treatment, and DNA was quantified by real-time PCR. Treatment with each of the 12 detergents and PMA increased the cycle threshold (Ct) values for heat inactivated E. coli suspensions. The greatest increase, of 10.68 Ct was obtained with sarkosyl. Treatment with sodium deoxycholate (NaDC) increased the Ct value by 8.99 Ct. Treatment with sarkosyl or NaDC of 16 heat treated 5-strain cocktails of verotoxigenic E. coli (VTEC) increased the mean Ct values by 8.15 or 6.82 Ct, respectively. Those mean values were significantly (pnumbers of viable E. coli were 2.24 and 2.47, respectively, with regression coefficient values ≥0.85. The findings show that sarkosyl was more effective than NaDC for dissipation of PMA-barrier properties of membranes of inactivated E. coli cells. Viable E. coli in mixtures of viable E. coli and E. coli inactivated by heat, lactic acid or peroxyacetic acid could be reliably enumerated by sarkosyl PMA-qPCR. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Propidium monoazide combined with real-time quantitative PCR to quantify viable Alternaria spp. contamination in tomato products.

    Science.gov (United States)

    Crespo-Sempere, Ana; Estiarte, Núria; Marín, Sonia; Sanchis, Vicente; Ramos, Antonio J

    2013-08-01

    Alternaria is a common contaminating genus of fungi in fruits, grains, and vegetables that causes severe economic losses to farmers and the food industry. Furthermore, it is claimed that Alternaria spp. are able to produce phytotoxic metabolites, and mycotoxins that are unsafe for human and animal health. DNA amplification techniques are being increasingly applied to detect, identify, and quantify mycotoxigenic fungi in foodstuffs, but the inability of these methods to distinguish between viable and nonviable cells might lead to an overestimation of mycotoxin-producing living cells. A promising technique to overcome this problem is the pre-treatment of samples with nucleic acid intercalating dyes, such as propidium monoazide (PMA), prior to quantitative PCR (qPCR). PMA selectively penetrates cells with a damaged membrane inhibiting DNA amplification during qPCRs. In our study, a primer pair (Alt4-Alt5) to specifically amplify and quantify Alternaria spp. by qPCR was designed. Quantification data of qPCR achieved a detection limit of 10(2)conidia/g of tomato. Here, we have optimized for the first time a DNA amplification-based PMA sample pre-treatment protocol for detecting viable Alternaria spp. cells. Artificially inoculated tomato samples treated with 65μM of PMA, showed a reduction in the signal by almost 7cycles in qPCR between live and heat-killed Alternaria spp. conidia. The tomato matrix had a protective effect on the cells against PMA toxicity, reducing the efficiency to distinguish between viable and nonviable cells. The results reported here indicate that the PMA-qPCR method is a suitable tool for quantifying viable Alternaria cells, which could be useful for estimating potential risks of mycotoxin contamination. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Identification of Viable Helicobacter pylori in Drinking Water Supplies by Cultural and Molecular Techniques.

    Science.gov (United States)

    Santiago, Paula; Moreno, Yolanda; Ferrús, M Antonía

    2015-08-01

    Helicobacter pylori is one of the most common causes of chronic bacterial infection in humans, directly related to peptic ulcer and gastric cancer. It has been suggested that H. pylori can be acquired through different transmission routes, including water. In this study, culture and qPCR were used to detect and identify the presence of H. pylori in drinking water. Furthermore, the combined techniques PMA-qPCR and DVC-FISH were applied for detection of viable cells of H. pylori. Among 24 drinking water samples, 16 samples were positive for the presence of H. pylori, but viable cells were only detected in six samples. Characteristic colonies, covered by a mass of bacterial unspecific growth, were observed on selective agar plates from an only sample, after enrichment. The mixed culture was submitted to DVC-FISH and qPCR analysis, followed by sequencing of the amplicons. Molecular techniques confirmed the growth of H. pylori on the agar plate. Our results demonstrate for the first time that H. pylori can survive and be potentially infective in drinking water, showing that water distribution systems could be a potential route for H. pylori transmission. © 2015 John Wiley & Sons Ltd.

  20. Real-time quantification of viable bacteria in liquid medium using infrared thermography

    Science.gov (United States)

    Salaimeh, Ahmad A.; Campion, Jeffrey J.; Gharaibeh, Belal Y.; Evans, Martin E.; Saito, Kozo

    2011-11-01

    Quantifying viable bacteria in liquids is important in environmental, food processing, manufacturing, and medical applications. Since vegetative bacteria generate heat as a result of biochemical reactions associated with cellular functions, thermal sensing techniques, including infrared thermography (IRT), have been used to detect viable cells in biologic samples. We developed a novel method that extends the dynamic range and improves the sensitivity of bacterial quantification by IRT. The approach uses IRT video, thermodynamics laws, and heat transfer mechanisms to directly measure, in real-time, the amount of energy lost as heat from the surface of a liquid sample containing bacteria when the specimen cools to a lower temperature over 2 min. We show that the Energy Content ( EC) of liquid media containing as few as 120 colony-forming units (CFU) of Escherichia coli per ml was significantly higher than that of sterile media ( P method that provides real-time bacterial enumeration over a wide dynamic range without the need for sample concentration, modification, or destruction. The approach could be adapted to quantify other living cells in a liquid milieu and has the potential for automation and high throughput.

  1. Evaluation of selective dry cow treatment following on-farm culture: Milk yield and somatic cell count in the subsequent lactation.

    Science.gov (United States)

    Cameron, M; Keefe, G P; Roy, J-P; Stryhn, H; Dohoo, I R; McKenna, S L

    2015-04-01

    Compared with blanket dry cow therapy (DCT), the selective antimicrobial treatment of cows based upon on-farm culture results has the potential to reduce the amount of antimicrobials used in dairy production. The objective of the current study was to determine the effect of a Petrifilm (3M Canada, London, Ontario) on-farm culture-based selective DCT program on milk yield and somatic cell count (SCC) in the following lactation. A total of 729 low-SCC (milk samples collected on the day before drying off, cows in the selective DCT group were treated at drying off based on the results obtained by the Petrifilm on-farm culture system with DCT and ITS (Petrifilm culture positive) or ITS alone (Petrifilm culture negative). Milk test-day records for the following lactation were obtained from Dairy Herd Improvement for all cows enrolled in the trial. Repeated measures linear mixed models were used to assess the effect of study group (blanket or selective DCT) on test-day milk production and natural logarithm of SCC over the first 180 d of the subsequent lactation. According to the final multivariable models, when low-SCC cows were selectively treated with DCT at drying off based on results obtained using the Petrifilm on-farm culture system, no effect on milk production (least squares means for blanket DCT = 39.3 kg vs. selective DCT = 39.0 kg) or natural logarithm of SCC (least squares means for blanket DCT = 3.95 vs. selective DCT = 3.97) was observed in the subsequent lactation when compared with cows receiving blanket DCT. The results of this study indicate that selective DCT based on results obtained by the Petrifilm on-farm culture system enabled a reduction in the use of DCT without negatively affecting milk production and milk quality. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  2. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  3. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2015-12-01

    Full Text Available BACKGROUND Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. METHODS In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. RESULTS Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1- fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity CONCLUSIONS Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  4. Social Networking and Smart Technology: Viable Environmental Communication Tools…?

    Science.gov (United States)

    Montain, J.; Byrne, J. M.

    2010-12-01

    To what extent do popular social networking channels represent a viable means for disseminating information regarding environmental change to the general public? Are new forms of communication such as YouTube™, Facebook™, MySpace™ and Twitter™ and smart devices such as iPhone™ and BlackBerry™ useful and effective in terms motivating people into social action and behavioural modification; or do they simply pay ‘lip service’ to these pressing environmental issues? This project will explore the background connections between social networking and environmental communication and education; and outline why such tools might be an appropriate way to connect to a broad audience in an efficient and unconventional manner. Further, research will survey the current prevalence of reliable environmental change information on social networking Internet-based media; and finally, suggestions for improved strategies and new directions will be provided.

  5. Dissolvable tattoo sensors: from science fiction to a viable technology

    Science.gov (United States)

    Cheng, Huanyu; Yi, Ning

    2017-01-01

    Early surrealistic painting and science fiction movies have envisioned dissolvable tattoo electronic devices. In this paper, we will review the recent advances that transform that vision into a viable technology, with extended capabilities even beyond the early vision. Specifically, we focus on the discussion of a stretchable design for tattoo sensors and degradable materials for dissolvable sensors, in the form of inorganic devices with a performance comparable to modern electronics. Integration of these two technologies as well as the future developments of bio-integrated devices is also discussed. Many of the appealing ideas behind developments of these devices are drawn from nature and especially biological systems. Thus, bio-inspiration is believed to continue playing a key role in future devices for bio-integration and beyond.

  6. A viable logarithmic f(R) model for inflation

    Energy Technology Data Exchange (ETDEWEB)

    Amin, M.; Khalil, S. [Center for Fundamental Physics, Zewail City of Science and Technology,6 October City, Giza (Egypt); Salah, M. [Center for Fundamental Physics, Zewail City of Science and Technology,6 October City, Giza (Egypt); Department of Mathematics, Faculty of Science, Cairo University,Giza (Egypt)

    2016-08-18

    Inflation in the framework of f(R) modified gravity is revisited. We study the conditions that f(R) should satisfy in order to lead to a viable inflationary model in the original form and in the Einstein frame. Based on these criteria we propose a new logarithmic model as a potential candidate for f(R) theories aiming to describe inflation consistent with observations from Planck satellite (2015). The model predicts scalar spectral index 0.9615

  7. Simultaneous pyometra and viable puppies’ gestation in a bitch

    Directory of Open Access Journals (Sweden)

    A. Risso

    2014-08-01

    Full Text Available Here we describe a case of pyometra coexisting with gestation in a 4.5 year-old miniature short-haired Dachshund. The dog exhibited depression, vaginal discharge, polydipsia and dehydration. Ultrasound examination revealed the presence of low to moderate anechoic fluid collection in the left uterine horn. Blood analysis revealed mild neutrophilia with a left shift. Based on these findings a presumptive diagnosis of pyometra was made and the bitch was treated using amoxicillin-clavulanate with dopaminergic agonist (cabergoline. A second ultrasound scan revealed the presence of two gestational vesicles in the right uterine horn that were successfully carried to term. Unusually, while pyometra persisted in the left uterine horn, two viable puppies were delivered by caesarean section from the right uterine horn.

  8. The viable but non-culturable state in pathogenic Escherichia coli: A general review

    Directory of Open Access Journals (Sweden)

    Jennifer A. Pienaar

    2016-05-01

    Objectives: This review discusses various general aspects of the VBNC state, the mechanisms and possible public health impact of indicator and pathogenic E. coli entering into the VBNC state. Method: A literature review was conducted to ascertain the possibleimpact of E. coli entering into the VBNC state. Results: Escherichia coli enter into the VBNC state by means of several induction mechanisms. Various authors have found that E. coli can be resuscitated post-VBNC. Certain strains of pathogenic E. coli are still able to produce toxins in the VBNC state, whilst others are avirulent during the VBNC state but are able to regain virulence after resuscitation. Conclusion: Pathogenic and indicator E. coli entering into the VBNC state could have an adverse effect on public health if conventional detection methods are used, where the number of viable cells could be underestimated and the VBNC cells still produce toxins or could, at anytime, be resuscitated and become virulent again.

  9. Profiling Total Viable Bacteria in a Hemodialysis Water Treatment System.

    Science.gov (United States)

    Chen, Lihua; Zhu, Xuan; Zhang, Menglu; Wang, Yuxin; Lv, Tianyu; Zhang, Shenghua; Yu, Xin

    2017-05-28

    Culture-dependent methods, such as heterotrophic plate counting (HPC), are usually applied to evaluate the bacteriological quality of hemodialysis water. However, these methods cannot detect the uncultured or viable but non-culturable (VBNC) bacteria, both of which may be quantitatively predominant throughout the hemodialysis water treatment system. Therefore, propidium monoazide (PMA)-qPCR associated with HPC was used together to profile the distribution of the total viable bacteria in such a system. Moreover, high-throughput sequencing of 16S rRNA gene amplicons was utilized to analyze the microbial community structure and diversity. The HPC results indicated that the total bacterial counts conformed to the standards, yet the bacteria amounts were abruptly enhanced after carbon filter treatment. Nevertheless, the bacterial counts detected by PMA-qPCR, with the highest levels of 2.14 × 10 7 copies/100 ml in softener water, were much higher than the corresponding HPC results, which demonstrated the occurrence of numerous uncultured or VBNC bacteria among the entire system before reverse osmosis (RO). In addition, the microbial community structure was very different and the diversity was enhanced after the carbon filter. Although the diversity was minimized after RO treatment, pathogens such as Escherichia could still be detected in the RO effluent. In general, both the amounts of bacteria and the complexity of microbial community in the hemodialysis water treatment system revealed by molecular approaches were much higher than by traditional method. These results suggested the higher health risk potential for hemodialysis patients from the up-to-standard water. The treatment process could also be optimized, based on the results of this study.

  10. Yield stress fluids slowly yield to analysis

    NARCIS (Netherlands)

    Bonn, D.; Denn, M.M.

    2009-01-01

    We are surrounded in everyday life by yield stress fluids: materials that behave as solids under small stresses but flow like liquids beyond a critical stress. For example, paint must flow under the brush, but remain fixed in a vertical film despite the force of gravity. Food products (such as

  11. Yield enhancement with DFM

    Science.gov (United States)

    Paek, Seung Weon; Kang, Jae Hyun; Ha, Naya; Kim, Byung-Moo; Jang, Dae-Hyun; Jeon, Junsu; Kim, DaeWook; Chung, Kun Young; Yu, Sung-eun; Park, Joo Hyun; Bae, SangMin; Song, DongSup; Noh, WooYoung; Kim, YoungDuck; Song, HyunSeok; Choi, HungBok; Kim, Kee Sup; Choi, Kyu-Myung; Choi, Woonhyuk; Jeon, JoongWon; Lee, JinWoo; Kim, Ki-Su; Park, SeongHo; Chung, No-Young; Lee, KangDuck; Hong, YoungKi; Kim, BongSeok

    2012-03-01

    A set of design for manufacturing (DFM) techniques have been developed and applied to 45nm, 32nm and 28nm logic process technologies. A noble technology combined a number of potential confliction of DFM techniques into a comprehensive solution. These techniques work in three phases for design optimization and one phase for silicon diagnostics. In the DFM prevention phase, foundation IP such as standard cells, IO, and memory and P&R tech file are optimized. In the DFM solution phase, which happens during ECO step, auto fixing of process weak patterns and advanced RC extraction are performed. In the DFM polishing phase, post-layout tuning is done to improve manufacturability. DFM analysis enables prioritization of random and systematic failures. The DFM technique presented in this paper has been silicon-proven with three successful tape-outs in Samsung 32nm processes; about 5% improvement in yield was achieved without any notable side effects. Visual inspection of silicon also confirmed the positive effect of the DFM techniques.

  12. Yield Improvement in Steel Casting (Yield II)

    Energy Technology Data Exchange (ETDEWEB)

    Richard A. Hardin; Christoph Beckermann; Tim Hays

    2002-02-18

    This report presents work conducted on the following main projects tasks undertaken in the Yield Improvement in Steel Casting research program: Improvement of Conventional Feeding and Risering Methods, Use of Unconventional Yield Improvement Techniques, and Case Studies in Yield Improvement. Casting trials were conducted and then simulated using the precise casting conditions as recorded by the participating SFSA foundries. These results present a statistically meaningful set of experimental data on soundness versus feeding length. Comparisons between these casting trials and casting trials performed more than forty years ago by Pellini and the SFSA are quite good and appear reasonable. Comparisons between the current SFSA feeding rules and feeding rules based on the minimum Niyama criterion reveal that the Niyama-based rules are generally less conservative. The niyama-based rules also agree better with both the trials presented here, and the casting trails performed by Pellini an d the SFSA years ago. Furthermore, the use of the Niyama criterion to predict centerline shrinkage for horizontally fed plate sections has a theoretical basis according to the casting literature reviewed here. These results strongly support the use of improved feeding rules for horizontal plate sections based on the Niyama criterion, which can be tailored to the casting conditions for a given alloy and to a desired level of soundness. The reliability and repeatability of ASTM shrinkage x-ray ratings was investigated in a statistical study performed on 128 x-rays, each of which were rated seven different times. A manual ''Feeding and Risering Guidelines for Steel Castings' is given in this final report. Results of casting trials performed to test unconventional techniques for improving casting yield are presented. These use a stacked arrangement of castings and riser pressurization to increase the casting yield. Riser pressurization was demonstrated to feed a casting up to

  13. Bond yield curve construction

    Directory of Open Access Journals (Sweden)

    Kožul Nataša

    2014-01-01

    Full Text Available In the broadest sense, yield curve indicates the market's view of the evolution of interest rates over time. However, given that cost of borrowing it closely linked to creditworthiness (ability to repay, different yield curves will apply to different currencies, market sectors, or even individual issuers. As government borrowing is indicative of interest rate levels available to other market players in a particular country, and considering that bond issuance still remains the dominant form of sovereign debt, this paper describes yield curve construction using bonds. The relationship between zero-coupon yield, par yield and yield to maturity is given and their usage in determining curve discount factors is described. Their usage in deriving forward rates and pricing related derivative instruments is also discussed.

  14. The elusive minimum viable population size for white sturgeon

    Energy Technology Data Exchange (ETDEWEB)

    Jager, Yetta [ORNL; Lepla, Ken B. [Idaho Power Company; Van Winkle, Webb [Van Windle Environmental Consulting; James, Mr Brad [Washington Department of Fish and Wildlife; McAdam, Dr Steve [University of British Columbia, Vancouver

    2010-01-01

    Biological conservation of sturgeon populations is a concern for many species. Those responsible for managing the white sturgeon (Acipenser transmontanus) and similar species are interested in identifying extinction thresholds to avoid. Two thresholds that exist in theory are the minimum viable population size (MVP) and minimum amount of suitable habitat. In this paper, we present both model and empirical estimates of these thresholds. We modified a population viability analysis (PVA) model for white sturgeon to include two new Allee mechanisms. Despite this, PVA-based MVP estimates were unrealistically low compared with empirical estimates unless opportunities for spawning were assumed to be less frequent. PVA results revealed a trade-off between MVP and habitat thresholds; smaller populations persisted in longer river segments and vice versa. Our empirical analyses suggested (1) a MVP range based on population trends from 1,194 to 27,700 individuals, and (2) a MVP estimate of 4,000 individuals based on recruitment. Long-term historical population surveys are needed for more populations to pinpoint an MVP based on trends, whereas the available data were sufficient to estimate MVP based on recruitment. Beyond the MVP, we developed a hierarchical model for population status based on empirical data. Metapopulation support was the most important predictor of population health, followed by the length of free-flowing habitat, with habitat thresholds at 26 and 150 km. Together, these results suggest that habitat and connectivity are important determinants of population status that likely influence the site-specific MVP thresholds.

  15. Towards a viable and just global nursing ethics.

    Science.gov (United States)

    Crigger, Nancy J

    2008-01-01

    Globalization, an outgrowth of technology, while informing us about people throughout the world, also raises our awareness of the extreme economic and social disparities that exist among nations. As part of a global discipline, nurses are vitally interested in reducing and eliminating disparities so that better health is achieved for all people. Recent literature in nursing encourages our discipline to engage more actively with social justice issues. Justice in health care is a major commitment of nursing; thus questions in the larger sphere of globalization, justice and ethics, are our discipline's questions also. Global justice, or fairness, is not an issue for some groups or institutions, but a deeper human rights issue that is a responsibility for everyone. What can we do to help reduce or eliminate the social and economic disparities that are so evident? What kind of ethical milieu is needed to address the threat that globalization imposes on justice and fairness? This article enriches the conceptualization of globalization by investigating recent work by Schweiker and Twiss. In addition, I discuss five qualities or characteristics that will facilitate the development of a viable and just global ethic. A global ethic guides all people in their response to human rights and poverty. Technology and business, two major forces in globalization that are generally considered beneficial, are critiqued as barriers to social justice and the common good.

  16. Keeping checkpoint/restart viable for exascale systems.

    Energy Technology Data Exchange (ETDEWEB)

    Riesen, Rolf E.; Bridges, Patrick G. (IBM Research, Ireland, Mulhuddart, Dublin); Stearley, Jon R.; Laros, James H., III; Oldfield, Ron A.; Arnold, Dorian (University of New Mexico, Albuquerque, NM); Pedretti, Kevin Thomas Tauke; Ferreira, Kurt Brian; Brightwell, Ronald Brian

    2011-09-01

    Next-generation exascale systems, those capable of performing a quintillion (10{sup 18}) operations per second, are expected to be delivered in the next 8-10 years. These systems, which will be 1,000 times faster than current systems, will be of unprecedented scale. As these systems continue to grow in size, faults will become increasingly common, even over the course of small calculations. Therefore, issues such as fault tolerance and reliability will limit application scalability. Current techniques to ensure progress across faults like checkpoint/restart, the dominant fault tolerance mechanism for the last 25 years, are increasingly problematic at the scales of future systems due to their excessive overheads. In this work, we evaluate a number of techniques to decrease the overhead of checkpoint/restart and keep this method viable for future exascale systems. More specifically, this work evaluates state-machine replication to dramatically increase the checkpoint interval (the time between successive checkpoint) and hash-based, probabilistic incremental checkpointing using graphics processing units to decrease the checkpoint commit time (the time to save one checkpoint). Using a combination of empirical analysis, modeling, and simulation, we study the costs and benefits of these approaches on a wide range of parameters. These results, which cover of number of high-performance computing capability workloads, different failure distributions, hardware mean time to failures, and I/O bandwidths, show the potential benefits of these techniques for meeting the reliability demands of future exascale platforms.

  17. Is Greenberg’s “Macro-Carib” viable?

    Directory of Open Access Journals (Sweden)

    Spike Gildea

    2007-08-01

    Full Text Available In his landmark work Language in the Americas, Greenberg (1987 proposed that Macro-Carib was one of the major low-level stocks of South America, which together with Macro-Panoan and Macro-Ge-Bororo were claimed to comprise the putative Ge-Pano-Carib Phylum. His Macro-Carib includes the isolates Andoke and Kukura, and the Witotoan, Peba-Yaguan, and Cariban families. Greenberg’s primary evidence came from person-marking paradigms in individual languages, plus scattered words from individual languages collected into 79 Macro-Carib ‘etymologies’ and another 64 Amerind ‘etymologies’. The goal of this paper is to re-evaluate Greenberg’s Macro-Carib claim in the light of the much more extensive and reliable language data that has become available largely since 1987. Based on full person-marking paradigms for Proto-Cariban, Yagua, Bora and Andoke, we conclude that Greenberg’s morphological claims are unfounded. For our lexical comparison, we created lexical lists for Proto-Cariban, Proto-Witotoan, Yagua and Andoke, for both Greenberg’s 143 putative etymologies and for the Swadesh 100 list. From both lists, a total of 23 potential cognates were found, but no consonantal correspondences were repeated even once. We conclude that our greatly expanded and improved database does not provide sufficient evidence to convince the skeptic that the Macro-Carib hypothesis is viable.

  18. Case-based anatomy teaching: a viable alternative?

    Science.gov (United States)

    Eseonu, Onyedikachi; Carachi, Robert; Brindley, Nicola

    2013-08-01

    Over the last two decades, there has been a decline in the amount of time available for anatomy teaching in the medical undergraduate curriculum, and new methods of anatomy teaching have been adopted for pragmatic reasons, with little evidence base to support their proposed educational benefits. This study seeks to establish the effect of a case-based teaching method on students' confidence in anatomy. Forty-three student volunteers in the clinical phase of the Glasgow medical course were given weekly anatomy teaching sessions based on clinical case presentations over 4 weeks. The students were given an anatomy test, and were asked to rate their confidence in their anatomy knowledge before and after the teaching sessions. There was a two-point increase in students' self-rated confidence, and a 10.9 per cent increase in average test score after the case-based anatomy teaching sessions. Both of these increases were statistically significant (p teaching was also highly rated by students, which may make it a viable option for the teaching of anatomy in the modern medical curriculum. © 2013 John Wiley & Sons Ltd.

  19. Protein design algorithms predict viable resistance to an experimental antifolate.

    Science.gov (United States)

    Reeve, Stephanie M; Gainza, Pablo; Frey, Kathleen M; Georgiev, Ivelin; Donald, Bruce R; Anderson, Amy C

    2015-01-20

    Methods to accurately predict potential drug target mutations in response to early-stage leads could drive the design of more resilient first generation drug candidates. In this study, a structure-based protein design algorithm (K* in the OSPREY suite) was used to prospectively identify single-nucleotide polymorphisms that confer resistance to an experimental inhibitor effective against dihydrofolate reductase (DHFR) from Staphylococcus aureus. Four of the top-ranked mutations in DHFR were found to be catalytically competent and resistant to the inhibitor. Selection of resistant bacteria in vitro reveals that two of the predicted mutations arise in the background of a compensatory mutation. Using enzyme kinetics, microbiology, and crystal structures of the complexes, we determined the fitness of the mutant enzymes and strains, the structural basis of resistance, and the compensatory relationship of the mutations. To our knowledge, this work illustrates the first application of protein design algorithms to prospectively predict viable resistance mutations that arise in bacteria under antibiotic pressure.

  20. Towards viable cosmological models of disformal theories of gravity

    Science.gov (United States)

    Sakstein, Jeremy

    2015-01-01

    The late-time cosmological dynamics of disformal gravity are investigated using dynamical systems methods. It is shown that in the general case there are no stable attractors that screen fifth forces locally and simultaneously describe a dark energy dominated universe. Viable scenarios have late-time properties that are independent of the disformal parameters and are identical to the equivalent conformal quintessence model. Our analysis reveals that configurations where the Jordan frame metric becomes singular are only reached in the infinite future, thus explaining the natural pathology resistance observed numerically by several previous works. The viability of models where this can happen is discussed in terms of both the cosmological dynamics and local phenomena. We identify a special parameter tuning such that there is a new fixed point that can match the presently observed dark energy density and equation of state. This model is unviable when the scalar couples to the visible sector but may provide a good candidate model for theories where only dark matter is disformally coupled.

  1. SMA actuators: a viable practical technology (Presentation Video)

    Science.gov (United States)

    Browne, Alan L.; Brown, Jeffrey; Hodgson, Darel E.

    2015-04-01

    Diverse products either based solely on or incorporating Shape Memory Alloys (SMA) have and are being made in a wide range of industries, and IP is being captured. Why then compared to SE (superelastic) Nitinol, and especially conventional technology, do so few ideas reach production? This presentation delves deeply into this topic in reaching the final assessment that SMA actuators are indeed now a viable practical technology. The presentation begins with an introduction to and description of the fundamental basis of SMA actuator technology. Examples of multiple commercially available geometric forms of SMA actuators are given and the functionalities that they provide are described. This is followed by examples of multiple commercial products incorporating such SMA actuators. Given that there are literally millions of commercial products incorporating conventional actuator technologies, indications are given as to why there are their less than 1000 that utilize SMA. Experience based challenges to the commercial use of SMA actuators are described. Besides having to compete with existing non-SMA technology which is quite mature additional challenges that are unique to SM actuators are indicated these including a wider than expected set of technical engineering problems and challenges and that a broader scope of dynamics is required.

  2. Detection of quantitative trait loci in Danish Holstein cattle affecting clinical mastitis, somatic cell score, udder conformation traits, and assessment of associated effects on milk yield

    DEFF Research Database (Denmark)

    Lund, M S; Guldbrandtsen, B; Buitenhuis, A J

    2008-01-01

    either a pleiotropic QTL affecting 2 traits or 2 QTL each affecting 1 trait gave some evidence to distinguish between these models. For Bos taurus autosome 5, the most likely models were a pleiotropic QTL affecting CM2, CM3, and SCS, and a linked QTL affecting fat yield index. For Bos taurus autosome 9...

  3. Increase in viral yield in eggs and MDCK cells of reassortant H5N1 vaccine candidate viruses caused by insertion of 38 amino acids into the NA stalk.

    Science.gov (United States)

    Zhang, Wenjun; Xue, Tao; Wu, Xiaowei; Zhang, Pinghu; Zhao, Guo; Peng, Daxing; Hu, Shunlin; Wang, Xiaoquan; Liu, Xiaowen; Liu, Wenbo; Liu, Xiufan

    2011-10-19

    The H5N1 subtype of highly pathogenic avian influenza viruses has spread to over 63 countries in Asia, Europe, and Africa and has become endemic in poultry. Since 2004, vaccination against H5N1 influenza has become common in domestic poultry operations in China. Most influenza vaccines have been produced in embryonated chicken eggs. High yield is the essential feature of a good vaccine candidate virus. Therefore, the large-scale manufacture of such a vaccine requires that the viral yield of H5N1 reassortant vaccine viruses in eggs and MDCK cells be increased. We generated two sets of reassortant H5N1 viruses based on backbone viruses A/Chicken/F/98 (H9N2) and A/Puerto Rico/8/34 (H1N1) using reverse genetics. The HAs and NAs of the reassortants were derived from the three epidemic H5N1 strains found in China. We compared the replication properties of these recombinant H5N1 viruses in embryonated chicken eggs and MDCK cells after inserting either 20 or 38 amino acids into their NA stalks. In this study, we demonstrated that inserting 38 amino acids into the NA stalks can significantly increase the viral yield of H5N1 reassortant viruses in both embryonated chicken eggs and MDCK cells, while inserting only 20 amino acids into the same NA stalks does not. Hemagglutinin inhibition testing and protection assays indicated that recombinant H5N1 viruses with 38 aa inserted into their NA stalks had the same antigenicity as the viruses with wt-NA. These results suggest that the generation of an H5N1 recombinant vaccine seed by the insertion of 38 aa into the NA stalk may be a suitable and more economical strategy for the increase in viral yield in both eggs and MDCK cells for the purposes of vaccine production. Copyright © 2011 Elsevier Ltd. All rights reserved.

  4. Cell-Type Transcriptomes of the Multicellular Green Alga Volvox carteri Yield Insights into the Evolutionary Origins of Germ and Somatic Differentiation Programs.

    Science.gov (United States)

    Matt, Gavriel Y; Umen, James G

    2017-12-05

    Germ-soma differentiation is a hallmark of complex multicellular organisms, yet its origins are not well understood. Volvox carteri is a simple multicellular green alga that has recently evolved a simple germ-soma dichotomy with only two cell types: large germ cells called gonidia and small terminally differentiated somatic cells. Here, we provide a comprehensive characterization of the gonidial and somatic transcriptomes of Volvox to uncover fundamental differences between the molecular and metabolic programming of these cell types. We found extensive transcriptome differentiation between cell types, with somatic cells expressing a more specialized program overrepresented in younger, lineage-specific genes and gonidial cells expressing a more generalist program overrepresented in more ancient genes that shared striking overlap with stem-cell-specific genes from animals and land plants. Directed analyses of metabolic pathways revealed a strong dichotomy between cell types with gonidial cells expressing growth-related genes and somatic cells expressing an altruistic metabolic program geared towards the assembly of flagella, which support organismal motility, and the conversion of storage carbon to sugars, which act as donors of extracellular matrix glycoproteins whose secretion enables massive organismal expansion. Volvox orthologs of Chlamydomonas diurnally controlled genes were analyzed for cell-type distribution and found to be strongly partitioned, with expression of dark-phase genes overrepresented in somatic cells and light-phase genes overrepresented in gonidial cells, a result that is consistent with cell type programs in Volvox arising by cooption of temporal regulons in a unicellular ancestor. Together our findings reveal fundamental molecular, metabolic, and evolutionary mechanisms that underlie the origins of germ-soma differentiation in Volvox and provide a template for understanding the acquisition of germ-soma differentiation in other multicellular

  5. The viable but non-culturable state in pathogenic Escherichia coli: A general review

    Directory of Open Access Journals (Sweden)

    Jennifer A. Pienaar

    2016-02-01

    Full Text Available Background: The persistence and pathogenicity of pathogenic bacteria are dependent on the ability of the species to survive in adverse conditions. During the infectious process, the organism may need to pass through certain hostile anatomical sites, such as the stomach. Under various environmental stresses, many bacteria enter into the viable but non-culturable (VBNC state, where they are ‘alive’ or metabolically active, but will not grow on conventional media. Escherichia coli bacteria encounter several diverse stress factors during their growth, survival and infection and thus may enter into the VBNC state.Objectives: This review discusses various general aspects of the VBNC state, the mechanisms and possible public health impact of indicator and pathogenic E. coli entering into the VBNC state.Method: A literature review was conducted to ascertain the possibleimpact of E. coli entering into the VBNC state.Results: Escherichia coli enter into the VBNC state by means of several induction mechanisms. Various authors have found that E. coli can be resuscitated post-VBNC. Certain strains of pathogenic E. coli are still able to produce toxins in the VBNC state, whilst others are avirulent during the VBNC state but are able to regain virulence after resuscitation.Conclusion: Pathogenic and indicator E. coli entering into the VBNC state could have an adverse effect on public health if conventional detection methods are used, where the number of viable cells could be underestimated and the VBNC cells still produce toxins or could, at anytime, be resuscitated and become virulent again.

  6. Acupuntura un tratamiento viable para las adicciones en Colombia

    Directory of Open Access Journals (Sweden)

    Hernán López-Suescún

    2013-09-01

    Full Text Available Resumen La acupuntura es una antigua técnica terapéutica desarrollada en China, que ha evidenciado ser efectiva en síntomas como las náuseas, vómito y dolor dentario. A pesar del sustento fisiológico que posibilitaría un uso efectivo en otras patologías, incluyendo el campo de las adicciones, los estudios son contradictorios, posiblemente por la diferencias de visión entre la medicina oriental y la occidental. El consumo de psicoactivos es un problema de salud pública en Colombia y en el mundo que genera grandes costos tangibles e intangibles, los cuales, en países desarrollados, puede llegar hasta el 1,6 % del PIB. En contraste, el beneficio económico del tratamiento de las adicciones, según las Naciones Unidas Contra la Droga y el Delito (UNODC, está entre 1:3 a 1:13; por lo tanto, cualquier esfuerzo que se realice en favor de los consumidores es una ganancia. Con base en estos datos, los organismos internacionales han generado políticas que ayudan a aminorar estos efectos. Colombia, como integrante de estos organismos, ha realizado varios compromisos para llevar a cabo dichas metas. Los tratamientos con auriculoterapia, como el protocolo NADA (National Acupuncture Detoxification Association, son los métodos más usados para las adicciones en el mundo, y aunque no se ha logrado evidenciar su efectividad, por su costo, facilidad y el poco riesgo de efectos adversos se hace viable en un país con pocos recursos económicos como Colombia.

  7. A propidium monoazide–quantitative PCR method for the detection and quantification of viable Enterococcus faecalis in large-volume samples of marine waters

    KAUST Repository

    Salam, Khaled W.

    2014-08-23

    The development of rapid detection assays of cell viability is essential for monitoring the microbiological quality of water systems. Coupling propidium monoazide with quantitative PCR (PMA-qPCR) has been successfully applied in different studies for the detection and quantification of viable cells in small-volume samples (0.25-1.00 mL), but it has not been evaluated sufficiently in marine environments or in large-volume samples. In this study, we successfully integrated blue light-emitting diodes for photoactivating PMA and membrane filtration into the PMA-qPCR assay for the rapid detection and quantification of viable Enterococcus faecalis cells in 10-mL samples of marine waters. The assay was optimized in phosphate-buffered saline and seawater, reducing the qPCR signal of heat-killed E. faecalis cells by 4 log10 and 3 log10 units, respectively. Results suggest that high total dissolved solid concentration (32 g/L) in seawater can reduce PMA activity. Optimal PMA-qPCR standard curves with a 6-log dynamic range and detection limit of 102 cells/mL were generated for quantifying viable E. faecalis cells in marine waters. The developed assay was compared with the standard membrane filter (MF) method by quantifying viable E. faecalis cells in seawater samples exposed to solar radiation. The results of the developed PMA-qPCR assay did not match that of the standard MF method. This difference in the results reflects the different physiological states of E. faecalis cells in seawater. In conclusion, the developed assay is a rapid (∼5 h) method for the quantification of viable E. faecalis cells in marine recreational waters, which should be further improved and tested in different seawater settings. © 2014 Springer-Verlag Berlin Heidelberg.

  8. Particle debonding using different yield criteria

    DEFF Research Database (Denmark)

    Legarth, Brian Nyvang; Kuroda, Mitsutoshi

    2004-01-01

    Effects of plastic anisotropy in relation to debonding of rigid inclusions embedded in an elastic-viscoplastic metal are studied. Full finite strain analyses are carried out for plane cells assuming plane stress or plane strain. The overall stress strain response is calculated, when the cell...... extent and shape of the particular yield function considered. The required overall straining of the cell for debonding initiation is related to the extent of the yield surfaces, since a high yield stress promotes debonding. Additionally, the maximum overall stress level for the cell is lower for the Hill...... [Hill, R., 1948. Proc. Roy. Soc. London Ser. A 193, 281-297] and Barlat et al. [Barlat, F., Lege, D.J., Brem, J.C., 1991. Int. J. Plasticity 7, 693-712] materials than that predicted by the other three yield functions. In all cases analyzed the non-normality flow rule hastens the particle...

  9. Detection of Only Viable Bacterial Spores Using a Live/Dead Indicator in Mixed Populations

    Science.gov (United States)

    Behar, Alberto E.; Stam, Christina N.; Smiley, Ronald

    2013-01-01

    This method uses a photoaffinity label that recognizes DNA and can be used to distinguish populations of bacterial cells from bacterial spores without the use of heat shocking during conventional culture, and live from dead bacterial spores using molecular-based methods. Biological validation of commercial sterility using traditional and alternative technologies remains challenging. Recovery of viable spores is cumbersome, as the process requires substantial incubation time, and the extended time to results limits the ability to quickly evaluate the efficacy of existing technologies. Nucleic acid amplification approaches such as PCR (polymerase chain reaction) have shown promise for improving time to detection for a wide range of applications. Recent real-time PCR methods are particularly promising, as these methods can be made at least semi-quantitative by correspondence to a standard curve. Nonetheless, PCR-based methods are rarely used for process validation, largely because the DNA from dead bacterial cells is highly stable and hence, DNA-based amplification methods fail to discriminate between live and inactivated microorganisms. Currently, no published method has been shown to effectively distinguish between live and dead bacterial spores. This technology uses a DNA binding photoaffinity label that can be used to distinguish between live and dead bacterial spores with detection limits ranging from 109 to 102 spores/mL. An environmental sample suspected of containing a mixture of live and dead vegetative cells and bacterial endospores is treated with a photoaffinity label. This step will eliminate any vegetative cells (live or dead) and dead endospores present in the sample. To further determine the bacterial spore viability, DNA is extracted from the spores and total population is quantified by real-time PCR. The current NASA standard assay takes 72 hours for results. Part of this procedure requires a heat shock step at 80 degC for 15 minutes before the

  10. Coscheduling in Clusters: Is It a Viable Alternative?

    Energy Technology Data Exchange (ETDEWEB)

    Choi, G S; Kim, J H; Ersoz, D; Yoo, A B; Das, C R

    2003-11-10

    than spin-based techniques like PB on a Linux platform. Third, the proposed HYBRID scheduling provides the best performance-energy behavior and can be implemented on any cluster with little effort. All these results suggest that blocking-based coscheduling techniques are viable candidates to be used instead of batching scheme for significant performance-energy benefits.

  11. Maize Residue as a Viable Substrate for Farm Scale Cultivation of Oyster Mushroom (Pleurotus ostreatus

    Directory of Open Access Journals (Sweden)

    Abena O. Adjapong

    2015-01-01

    Full Text Available In the search for alternatives to sawdust as growing media in commercial mushroom cultivation, three organic substrates obtainable as crop residue, maize husk, maize cob, and maize stalk, with each being supplemented with rice bran, were evaluated as growth media for the oyster mushroom, Pleurotus ostreatus (Kummer. For the tested alternatives to sawdust, the harvested weight of fruiting bodies that sprouted on a kilogram maize husk media per crop (32.99 g was the highest. Sawdust media supported significantly (P<0.001 heavier fruiting bodies (42.18 than the maize residues. The peak mushroom harvests for the various substrates were obtained between the first and seventh fruiting body flushes. The biological efficiency of the substrates, which measured usable nutrients indicated that maize stalk supplemented with rice bran, was 39% compared to that of the sawdust media (60%. The maize husk media and the maize cob media had biological efficiencies of 32% and 9.5%, respectively. These results indicate that two of the tested growing media (maize stalk or husk produced mushrooms with yield characteristics that were comparable to the well-used sawdust in the cultivation of oyster mushrooms. The environmental and economic parameters involved in the use and carting of sawdust make these on-farm crop residues a viable alternative for mushroom cultivation in especially nonforest zones of Ghana.

  12. Is flapless implant surgery a viable option in posterior maxilla? A review.

    Science.gov (United States)

    Doan, N; Du, Z; Crawford, R; Reher, P; Xiao, Y

    2012-09-01

    This article reviews the literature on the outcome of flapless surgery for dental implants in the posterior maxilla. The literature search was carried out in using the keywords: flapless, dental implants and maxilla. A hand search and Medline search were carried out on studies published between 1971 and 2011. The authors included research involving a minimum of 15 dental implants with a follow-up period of 1 year, an outcome measurement of implant survival, but excluded studies involving multiple simultaneous interventions, and studies with missing data. The Cochrane approach for cohort studies and Oxford Centre for Evidence-Based Medicine were applied. Of the 56 published papers selected, 14 papers on the flapless technique showed high overall implant survival rates. The prospective studies yielded 97.01% (95% CI: 90.72-99.0) while retrospective studies or case series illustrated 95.08% (95% CI: 91.0-97.93) survival. The average of intraoperative complications was 6.55% using the flapless procedure. The limited data obtained showed that flapless surgery in posterior maxilla areas could be a viable and predictable treatment method for implant placement. Flapless surgery tends to be more applicable in this area of the mouth. Further long-term clinical controlled studies are needed. Copyright © 2012 International Association of Oral and Maxillofacial Surgeons. All rights reserved.

  13. Expression of adhesion molecules, chemokines and matrix metallo- proteinases (MMPs) in viable and degenerating stage of Taenia solium metacestode in swine neurocysticercosis.

    Science.gov (United States)

    Singh, Satyendra K; Singh, Aloukick K; Prasad, Kashi N; Singh, Amrita; Singh, Avinash; Rai, Ravi P; Tripathi, Mukesh; Gupta, Rakesh K; Husain, Nuzhat

    2015-11-30

    Neurocysticercosis (NCC) is a parasitic infection of central nervous system (CNS). Expression of adhesion molecules, chemokines and matrix metalloproteinases (MMPs) were investigated on brain tissues surrounding viable (n=15) and degenerating cysticerci (n=15) of Taenia solium in swine by real-time RT-PCR and ELISA. Gelatin gel zymography was performed for MMPs activity. ICAM-1 (intercellular adhesion molecule-1), E-selectin, MIP-1α (macrophage inflammatory protein-1α), Eotaxin-1 and RANTES (regulated on activation, normal T cell expressed and secreted) were associated with degenerating cysticerci (cysts). However, VCAM-1 (vascular cell adhesion molecule-1), MCP-1 (monocyte chemotactic protein-1), MMP-2 and MMP-9 were associated with both viable and degenerating cysts. In conclusion, viable and degenerating cysticerci have different immune molecule profiles and role of these molecules in disease pathogenesis needs to be investigated. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Impact of constitutional polymorphisms in VCAM1 and CD44 on CD34+ cell collection yield after administration of granulocyte colony-stimulating factor to healthy donors.

    Science.gov (United States)

    Martín-Antonio, Beatriz; Carmona, Magdalena; Falantes, Jose; Gil, Encarnación; Baez, Alicia; Suarez, María; Marín, Pedro; Espigado, Ildefonso; Urbano-Ispizua, Alvaro

    2011-01-01

    Background The number of CD34(+) cells mobilized from bone marrow to peripheral blood after administration of granulocyte colony-stimulating factor varies greatly among healthy donors. This fact might be explained, at least in part, by constitutional differences in genes involved in the interactions tethering CD34(+) cells to the bone marrow. We analyzed genetic characteristics associated with CD34(+) cell mobilization in 112 healthy individuals receiving granulocyte colony-stimulating factor (filgrastim; 10 μg/kg; 5 days). Genetic variants in VCAM1 and in CD44 were associated with the number of CD34(+) cells in peripheral blood after granulocyte colony-stimulating factor administration (P = 0.02 and P = 0.04, respectively), with the quantity of CD34(+) cells ×10⁶/kg of donor (4.6 versus 6.3; P donor and total CD34(+) cells ×10⁶ (5.3 versus 6.7; P = 0.02 and 399 versus 533; P = 0.01, respectively). Conclusions In conclusion, genetic variability in molecules involved in migration and homing of CD34(+) cells influences the degree of mobilization of these cells.

  15. Detection and quantification of viable Bacillus cereus group species in milk by propidium monoazide quantitative real-time PCR.

    Science.gov (United States)

    Cattani, Fernanda; Barth, Valdir C; Nasário, Jéssica S R; Ferreira, Carlos A S; Oliveira, Sílvia D

    2016-04-01

    The Bacillus cereus group includes important spore-forming bacteria that present spoilage capability and may cause foodborne diseases. These microorganisms are traditionally evaluated in food using culturing methods, which can be laborious and time-consuming, and may also fail to detect bacteria in a viable but nonculturable state. The purpose of this study was to develop a quantitative real-time PCR (qPCR) combined with a propidium monoazide (PMA) treatment to analyze the contamination of UHT milk by B. cereus group species viable cells. Thirty micrograms per milliliter of PMA was shown to be the most effective concentration for reducing the PCR amplification of extracellular DNA and DNA from dead cells. The quantification limit of the PMA-qPCR assay was 7.5 × 10(2) cfu/mL of milk. One hundred thirty-five UHT milk samples were analyzed to evaluate the association of PMA to qPCR to selectively detect viable cells. The PMA-qPCR was able to detect B. cereus group species in 44 samples (32.6%), whereas qPCR without PMA detected 78 positive samples (57.8%). Therefore, the PMA probably inhibited the amplification of DNA from cells that were killed during UHT processing, which avoided an overestimation of bacterial cells when using qPCR and, thus, did not overvalue potential health risks. A culture-based method was also used to detect and quantify B. cereus sensu stricto in the same samples and showed positive results in 15 (11.1%) samples. The culture method and PMA-qPCR allowed the detection of B. cereus sensu stricto in quantities compatible with the infective dose required to cause foodborne disease in 3 samples, indicating that, depending on the storage conditions, even after UHT treatment, infective doses may be reached in ready-to-consume products. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  16. Eradication of high viable loads of Listeria monocytogenes contaminating food-contact surfaces

    Directory of Open Access Journals (Sweden)

    Silvia ede Candia

    2015-07-01

    Full Text Available This study demonstrates the efficacy of cold gaseous ozone treatments at low concentrations in the eradication of high Listeria monocytogenes viable cell loads from glass, polypropylene, stainless steel and expanded polystyrene food-contact surfaces. Using a step by step approach, involving the selection of the most resistant strain-surface combinations, 11 Listeria spp. strains resulted inactivated by a continuous ozone flow at 1.07 mg m-3 after 24 or 48 h of cold incubation, depending on both strain and surface evaluated. Increasing the inoculum level to 9 log CFU coupon-1, the best inactivation rate was obtained after 48h of treatment at 3.21 mg m-3 ozone concentration when cells were deposited onto stainless steel and expanded polystyrene coupons, resulted the most resistant food-contact surfaces in the previous assays.The addition of naturally microbiologically contaminated meat extract to a high load of L. monocytogenes LMG 23775 cells, the most resistant strain out of the 11 assayed Listeria spp. strains, led to its complete inactivation after four days of treatment.To the best of our knowledge, this is the first report describing the survival of L. monocytogenes and the effect of ozone treatment under cold storage conditions on expanded polystyrene, a commonly-used material in food packaging. These results could be useful for reducing pathogen cross-contamination phenomena during cold food storage.

  17. Electron-deficient N-alkyloyl derivatives of thieno[3,4-c]pyrrole-4,6-dione yield efficient polymer solar cells with open-circuit voltages > 1 v

    KAUST Repository

    Warnan, Julien

    2014-05-13

    Poly(benzo[1,2-b:4,5-b′]dithiophene-thieno[3,4-c]pyrrole-4,6-dione) (PBDTTPD) polymer donors yield some of the highest open-circuit voltages (V OC, ca. 0.9 V) and fill factors (FF, ca. 70%) in conventional bulk-heterojunction (BHJ) solar cells with PCBM acceptors. Recent work has shown that the incorporation of ring substituents into the side chains of the BDT motifs in PBDTTPD can induce subtle variations in material properties, resulting in an increase of the BHJ device VOC to ∼1 V. In this contribution, we report on the synthesis of N-alkyloyl-substituted TPD motifs (TPD(CO)) and show that the electron-deficient motifs can further lower both the polymer LUMO and HOMO levels, yielding device VOC > 1 V (up to ca. 1.1 V) in BHJ solar cells with PCBM. Despite the high VOC achieved (i.e., low polymer HOMO), BHJ devices cast from TPD(CO)-based polymer donors can reach power conversion efficiencies (PCEs) of up to 6.7%, making these promising systems for use in the high-band-gap cell of tandem solar cells. © 2014 American Chemical Society.

  18. Colpoda secrete viable Listeria monocytogenes within faecal pellets.

    Science.gov (United States)

    Raghu Nadhanan, Rethish; Thomas, Connor J

    2014-02-01

    Transmission electron microscopy was used to demonstrate that co-cultures of the ciliate Colpoda RR (an environmental isolate) and Colpoda MLS-5 (a food processing environment isolate) with the pathogenic Listeria monocytogenes DRDC8 resulted in secretion of faecal pellets containing intact DRDC8 cells. A green fluorescent protein expressing variant of DRDC8 was used in co-cultures to confirm that the pellet-associated bacterial cells were L. monocytogenes. Viability was confirmed by plate counts, and assay of microbial respiratory activity-proved DRDC8 cells present within faecal pellets was metabolically active. Following treatment of faecal pellets secreted by Colpoda RR and MLS-5 with gentamycin and sodium hypochlorite (NaOCl), no loss of viability of the pellet-located DRDC8 cells was observed, indicating that faecal pellet encapsulated DRDC8 cells are resistant to biocidal agents. This work suggests that Colpoda-derived faecal pellets may provide a mechanism for transmission of L. monocytogenes and other pathogenic bacteria. Furthermore, bacteria encapsulated by faecal pellets may be resistant to disinfectants and cleaning agents used in food manufacturing and preparation facilities. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

  19. Effect of myocardial revascularisation on left ventricular systolic function in patients with and without viable myocardium: should non-viable segments be revascularised?

    Science.gov (United States)

    Stipac, Alja Vlahovic; Stankovic, Ivan; Vidakovic, Radosav; Putnikovic, Biljana; Ilic, Ivan; Milicic, Biljana; Neskovic, Aleksandar N

    2013-12-01

    To assess the effect of surgical revascularisation on left ventricular (LV) systolic function in patients with viable and non-viable dysfunctional LV segments determined by low dose dobutamine stress echocardiography (DSE). Prospective observational cohort study. Single tertiary care centre. Consecutive patients referred to surgical revascularisation (n=115). DSE and surgical revascularisation. Functional recovery defined as increase in ejection fraction ≥ 5% 1 year after revascularisation in patients with and without viable myocardium (viability defined as improvement of contractility in ≥ 4 LV segments on DSE). The mean age, ejection fraction and wall motion score index (WMSi) of patients were 59 ± 9 years, 44 ± 9% and 1.82 ± 0.31, respectively. There was no difference between DSE positive and DSE negative patients for any of those parameters at baseline study (p>0.05 for all). After 12 months, the ejection fraction increased 11 ± 1% in patients with viable myocardium vs 7 ± 1% in patients without viable myocardium (p=0.002). Moreover, in patients with viable myocardium, the greatest increase of ejection fraction occurred 1 month after surgery (9 ± 1%), whereas in those patients with negative DSE the ejection fraction increased more gradually (2±1% after 1 month, p=0.002 between groups for 1 month vs preoperative value), but still improved after 12 months follow-up (pmyocardial revascularisation. Functional recovery continuously occurs throughout the first year after surgical treatment.

  20. Genome-wide association mapping in winter barley for grain yield and culm cell wall polymer content using the high-throughput CoMPP technique

    DEFF Research Database (Denmark)

    Bellucci, Andrea; Tondelli, Alessandro; Fangel, Jonatan Ulrik

    2017-01-01

    A collection of 112 winter barley varieties (Hordeum vulgare L.) was grown in the field for two years (2008/09 and 2009/10) in northern Italy and grain and straw yields recorded. In the first year of the trial, a severe attack of barley yellow mosaic virus (BaYMV) strongly influenced final...... gene, which is known to regulate resistance to BaYMV. Subsequently the QTL was shown to be tightly linked to rym4, a locus for resistance to the virus. GWAs on arabinans quantified by LM6 resulted in the identification of major QTLs closely located on 3H and hypotheses regarding putative candidate...

  1. Increased mobilization and yield of stem cells using plerixafor in combination with granulocyte-colony stimulating factor for the treatment of non-Hodgkin’s lymphoma and multiple myeloma

    Science.gov (United States)

    Pelus, Louis M; Farag, Sherif S

    2011-01-01

    Multiple myeloma and non-Hodgkin’s lymphoma remain the most common indications for high-dose chemotherapy and autologous peripheral blood stem cell rescue. While a CD34+ cell dose of 1 × 106/kg is considered the minimum required for engraftment, higher CD34+ doses correlate with improved outcome. Numerous studies, however, support targeting a minimum CD34+ cell dose of 2.0 × 106/kg, and an “optimal” dose of 4 to 6 × 106/kg for a single transplant. Unfortunately, up to 40% of patients fail to mobilize an optimal CD34+ cell dose using myeloid growth factors alone. Plerixafor is a novel reversible inhibitor of CXCR4 that significantly increases the mobilization and collection of higher numbers of hematopoietic progenitor cells. Two randomized multi-center clinical trials in patients with non-Hodgkin’s lymphoma and multiple myeloma have demonstrated that the addition of plerixafor to granulocyte-colony stimulating factor increases the mobilization and yield of CD34+ cells in fewer apheresis days, which results in durable engraftment. This review summarizes the pharmacology and evidence for the clinical efficacy of plerixafor in mobilizing hematopoietic stem and progenitor cells, and discusses potential ways to utilize plerixafor in a cost-effective manner in patients with these diseases. PMID:24198526

  2. Predictors of mother and child DNA yields in buccal cell samples collected in pediatric cancer epidemiologic studies: a report from the Children's Oncology group

    National Research Council Canada - National Science Library

    Poynter, Jenny N; Ross, Julie A; Hooten, Anthony J; Langer, Erica; Blommer, Crystal; Spector, Logan G

    2013-01-01

    ...; however, DNA collection in young children poses additional challenges. Here, we have evaluated predictors of DNA quantity in buccal cells collected for population-based studies of infant leukemia (N...

  3. Experimental investigation of temperature effect on yield of a pool of oil possessing viscoelastic properties

    Energy Technology Data Exchange (ETDEWEB)

    Ametov, I.M.; Baidikov, Yu.N.; Berezhnoi, N.I.; Bereryuk, I.M.; Ruzin, L.M.

    1982-02-01

    The effect of temperature on petroleum yield of a pool of viscoelastic oil when displaced by water from a viable porous medium model is considered. Based on experiments, a nonmonotonous character of the dependence of the oil yield on temperature is established. A decrease of oil yield following a rise in temperature is noted. This is explained by an anomaly of viscoelastic properties within that temperature interval.

  4. Lengthening of high-yield production levels of monoclonal antibody-producing Chinese hamster ovary cells by downregulation of breast cancer 1.

    Science.gov (United States)

    Matsuyama, Rima; Yamano, Noriko; Kawamura, Namiko; Omasa, Takeshi

    2017-03-01

    The establishment process of high-producing Chinese hamster ovary (CHO) cells for therapeutic protein production is usually laborious and time consuming because of the low probability of obtaining stable, high-producing clones over a long term. Thus, development of an efficient approach is required to establish stable, high-producing cells. This study presents a novel method that can efficiently establish sustainably high-producing cell lines by acceleration of transgene amplification and suppression of transgene silencing. The effects of breast cancer 1 (BRCA1) downregulation on gene amplification efficiency and long-term productivity were investigated in CHO cells. Small interfering RNA expression vectors against BRCA1 were transfected into the CHO DG44-derived antibody-producing cell clone. Individual cell clones were obtained after induction of gene amplification in the presence of 400 nM methotrexate, which were cultured until passage 20. BRCA1-downregulated cell clones CHO B1Sa and B1Sb displayed 2.2- and 1.6-fold higher specific production rates than the S-Mock clone. Fluorescence in situ hybridization showed that transgene amplification occurred at a high frequency in B1Sa and B1Sb clones. Moreover, B1Sa and B1Sb clones at 20 passages had approximately 3.5- and 5.3-fold higher productivity than the S-Mock clone. Histone modification analysis revealed a decrease in an active mark for transcription, trimethylation of histone H3 at lysine 4 (H3K4), in the transgene locus of the S-Mock clone. However, H3K4 trimethylation levels were not decreased in B1Sa and B1Sb clones during long term culture. Our results suggest that high-producing cells, which maintain their productivity long-term, were efficiently established by BRCA1 downregulation. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  5. A technique for determining viable military logistics support alternatives

    Science.gov (United States)

    Hester, Jesse Stuart

    A look at today's US military will see them operating much beyond the scope of protecting and defending the United States. These operations now consist of, but are not limited to humanitarian aid, disaster relief, peace keeping, and conflict resolution. This broad spectrum of operational environments has necessitated a transformation of the individual military services to a hybrid force that is attempting to leverage the inherent and emerging capabilities and strengths of all those under the umbrella of the Department of Defense (DOD), this concept has been coined Joint Operations. Supporting Joint Operations requires a new approach to determining a viable military logistics support system. The logistics architecture for these operations has to accommodate scale, time, varied mission objectives, and imperfect information. Compounding the problem is the human in the loop (HITL) decision maker (DM) who is a necessary component for quickly assessing and planning logistics support activities. Past outcomes are not necessarily good indicators of future results, but they can provide a reasonable starting point for planning and prediction of specific needs for future requirements. Adequately forecasting the necessary logistical support structure and commodities needed for any resource intensive environment has progressed well beyond stable demand assumptions to one in which dynamic and nonlinear environments can be captured with some degree of fidelity and accuracy. While these advances are important, a holistic approach that allows exploration of the operational environment or design space does not exist to guide the military logistician in a methodical way to support military forecasting activities. To bridge this capability gap, a method called Adaptive Technique for Logistics Architecture Solutions (ATLAS) has been developed. This method provides a process that facilitates the use of techniques and tools that filter and provide relevant information to the DM. By doing

  6. Lipogenic metabolism: a viable target for prostate cancer treatment?

    Directory of Open Access Journals (Sweden)

    Mengmeng Liang

    2014-10-01

    Full Text Available Cancer cells often depend on altered metabolism compared with their normal counterparts. [1],[2],[3],[4] As observed in 1924 by Otto Warburg, cancer cells show preferential glucose consumption by way of aerobic glycolysis while normal cells generally assume mitochondrial oxidative phosphorylation. [4] Another metabolic hallmark of carcinogenesis is altered lipid metabolism, whereby cancer cells may adopt enhanced de novo lipid production (lipogenesis. [1],[2],[3] Enhanced lipid metabolism is also observed in individuals with metabolic syndromes potentially a consequence of increasing popularity of the Standard American Diet, composed of high levels of saturated fats and carbohydrates. [5] A growing body of epidemiological data indicates a positive correlation between the occurrence of metabolic syndromes, such as cardiovascular disease, obesity, type-2 diabetes and associated hyperinsulemia, with the aggressiveness of cancer. [6],[7],[8],[9] Remarkably, it is estimated that for every 1% reduction in saturated fats, replaced by polyunsaturated, there would be a 2%-3% reduction in cardiovascular disease. [10] Thus, it is conceivable that an equally remarkable attenuation in cancer progression might be achieved with such a reduction in lipid accumulation.

  7. An In-vivo Prospective Study of the Diagnostic Yield and Accuracy of Optical Biopsy Compared with Conventional Renal Mass Biopsy for the Diagnosis of Renal Cell Carcinoma: The Interim Analysis.

    Science.gov (United States)

    Buijs, Mara; Wagstaff, Peter G K; de Bruin, Daniel M; Zondervan, Patricia J; Savci-Heijink, Cemile Dilara; van Delden, Otto M; van Leeuwen, Ton G; van Moorselaar, R Jeroen A; de la Rosette, Jean J M C H; Laguna Pes, Maria Pilar

    2017-10-24

    Lack of accuracy in preoperative imaging leads to overtreatment of benign renal masses (RMs) or indolent renal cell carcinomas (RCCs). Optical coherence tomography (OCT) is real time and high resolution, enabling quantitative analysis through attenuation coefficient (μOCT, mm(-1)). To determine the accuracy and diagnostic yield of OCT and renal mass biopsy (RMB) for the differentiation of benign RMs versus RCC and oncocytoma versus RCC. From October 2013 to June 2016, 95 patients with solid enhancing RMs on cross-sectional imaging were prospectively included. All patients underwent subsequent excision or ablation. Percutaneous, image-guided, needle-based OCT followed by RMB in an outpatient setting under local anaesthesia. Accuracy and diagnostic yield, μOCT correlated to resection pathology or second biopsy during ablation. Tables (2×2) for RMB, receiver operating characteristic curve for OCT. Mann-Whitney test to differentiate μOCT of RMs. RMB diagnostic yield was 79% with sensitivity, specificity, positive predictive value, and negative predictive value (NPV) of 100%, 89%, 99%, and 100%, respectively. Diagnostic yield and added value of OCT to differentiate RCC from benign was 99% and 15%, respectively. Significant difference was observed in median μOCT between benign RMs (3.2mm(-1), interquartile range [IQR]: 2.65-4.35) and RCCs (4.3mm(-1), IQR: 3.70-5.00), p=0.0171, and oncocytomas (3.38mm(-1), IQR: 2.68-3.95) and RCCs (4.3mm(-1), IQR: 3.70-5.00), p=0.0031. OCT showed sensitivity, specificity, positive predictive value. and NPV of 91%, 56%, 91%, and 56%, respectively, to differentiate benign RMs from RCCs and 92%, 67%, 95%, and 55%, respectively, to differentiate oncocytoma from RCC. Limitations include two reference standards and heterogeneity benign RMs. Compared with RMB, OCT has a higher diagnostic yield. OCT accurately distinguishes benign RMs from RCCs, and oncocytoma from RCCs, although specificity and NPV are lower. Optical coherence tomography, a

  8. Articular Cartilage Repair Using Marrow Stimulation Augmented with a Viable Chondral Allograft: 9-Month Postoperative Histological Evaluation

    Directory of Open Access Journals (Sweden)

    James K. Hoffman

    2015-01-01

    Full Text Available Marrow stimulation is frequently employed to treat focal chondral defects of the knee. However, marrow stimulation typically results in fibrocartilage repair tissue rather than healthy hyaline cartilage, which, over time, predisposes the repair to failure. Recently, a cryopreserved viable chondral allograft was developed to augment marrow stimulation. The chondral allograft is comprised of native viable chondrocytes, chondrogenic growth factors, and extracellular matrix proteins within the superficial, transitional, and radial zones of hyaline cartilage. Therefore, host mesenchymal stem cells that infiltrate the graft from the underlying bone marrow following marrow stimulation are provided with the optimal microenvironment to undergo chondrogenesis. The present report describes treatment of a trochlear defect with marrow stimulation augmented with this novel chondral allograft, along with nine month postoperative histological results. At nine months, the patient demonstrated complete resolution of pain and improvement in function, and the repair tissue consisted of 85% hyaline cartilage. For comparison, a biopsy obtained from a patient 8.2 months after treatment with marrow stimulation alone contained only 5% hyaline cartilage. These outcomes suggest that augmenting marrow stimulation with the viable chondral allograft can eliminate pain and improve outcomes, compared with marrow stimulation alone.

  9. Real-time detection of viable microorganisms by intracellular phototautomerism

    Directory of Open Access Journals (Sweden)

    Schuren Frank

    2010-06-01

    Full Text Available Abstract Background To date, the detection of live microorganisms present in the environment or involved in infections is carried out by enumeration of colony forming units on agar plates, which is time consuming, laborious and limited to readily cultivable microorganisms. Although cultivation-independent methods are available, they involve multiple incubation steps and do mostly not discriminate between dead or live microorganisms. We present a novel generic method that is able to specifically monitor living microorganisms in a real-time manner. Results The developed method includes exposure of cells to a weak acid probe at low pH. The neutral probe rapidly permeates the membrane and enters the cytosol. In dead cells no signal is obtained, as the cytosolic pH reflects that of the acidic extracellular environment. In live cells with a neutral internal pH, the probe dissociates into a fluorescent phototautomeric anion. After reaching peak fluorescence, the population of live cells decays. This decay can be followed real-time as cell death coincides with intracellular acidification and return of the probe to its uncharged non-fluorescent state. The rise and decay of the fluorescence signal depends on the probe structure and appears discriminative for bacteria, fungi, and spores. We identified 13 unique probes, which can be applied in the real-time viability method described here. Under the experimental conditions used in a microplate reader, the reported method shows a detection limit of 106 bacteria ml-1, while the frequently used LIVE/DEAD BacLight™ Syto9 and propidium iodide stains show detection down to 106 and 107 bacteria ml-1, respectively. Conclusions We present a novel fluorescence-based method for viability assessment, which is applicable to all bacteria and eukaryotic cell types tested so far. The RTV method will have a significant impact in many areas of applied microbiology including research on biocidal activity, improvement of

  10. State-transitions facilitate robust quantum yields and cause an over-estimation of electron transport in Dunaliella tertiolecta cells held at the CO₂ compensation point and re-supplied with DIC.

    Science.gov (United States)

    Ihnken, Sven; Kromkamp, Jacco C; Beardall, John; Silsbe, Greg M

    2014-03-01

    Photosynthetic energy consumption and non-photosynthetic energy quenching processes are inherently linked. Both processes must be controlled by the cell to allow cell maintenance and growth, but also to avoid photodamage. We used the chlorophyte algae Dunaliella tertiolecta to investigate how the interactive regulation of photosynthetic and non-photosynthetic pathways varies along dissolved inorganic carbon (DIC) and photon flux gradients. Specifically, cells were transferred to DIC-deplete media to reach a CO₂ compensation before being re-supplied with DIC at various concentrations and different photon flux levels. Throughout these experiments we monitored and characterized the photophysiological responses using pulse amplitude modulated fluorescence, oxygen evolution, 77 K fluorescence emission spectra, and fast-repetition rate fluorometry. O₂ uptake was not significantly stimulated at DIC depletion, which suggests that O₂ production rates correspond to assimilatory photosynthesis. Fluorescence-based measures of relative electron transport rates (rETRs) over-estimated oxygen-based photosynthetic measures due to a strong state-transitional response that facilitated high effective quantum yields. Adoption of an alternative fluorescence-based rETR calculation that accounts for state-transitions resulted in improved linear oxygen versus rETR correlation. This study shows the extraordinary capacity of D. tertiolecta to maintain stable effective quantum yields by flexible regulation of state-transitions. Uncertainties about the control mechanisms of state-transitions are presented.

  11. Birth of viable puppies derived from breeding cloned female dogs with a cloned male.

    Science.gov (United States)

    Park, J E; Hong, S G; Kang, J T; Oh, H J; Kim, M K; Kim, M J; Kim, H J; Kim, D Y; Jang, G; Lee, B C

    2009-09-15

    Since the establishment of production of viable cloned dogs by somatic cell nucleus transfer, great concern has been given to the reproductive abilities of these animals (Canis familiaris). Therefore, we investigated reproductive activity of cloned dogs by (1) performing sperm analysis using computer-assisted sperm analysis and early embryonic development, (2) assessing reproductive cycling by measuring serum progesterone (P4) levels and performing vaginal cytology, and (3) breeding cloned dogs using artificial insemination. Results showed that most parameters of sperm motility in a cloned male dog were within the reference range, and in vivo-matured oocytes from a noncloned female were successfully fertilized by spermatozoa from a cloned male dog and develop normally to the 8-cell stage. Three cloned female dogs displayed normal patterns of P4 levels and morphologic changes of the vaginal epithelium. Two cloned female dogs became pregnant using semen from a cloned male dog and successfully delivered 10 puppies by natural labor. In conclusion, these data demonstrated that both cloned male and female dogs are fertile, and their puppies are currently alive and healthy with normal growth patterns.

  12. The Role of Mitochondria from Mature Oocyte to Viable Blastocyst

    Directory of Open Access Journals (Sweden)

    Scott Chappel

    2013-01-01

    Full Text Available The oocyte requires a vast supply of energy after fertilization to support critical events such as spindle formation, chromatid separation, and cell division. Until blastocyst implantation, the developing zygote is dependent on the existing pool of mitochondria. That pool size within each cell decreases with each cell division. Mitochondria obtained from oocytes of women of advanced reproductive age harbor DNA deletions and nucleotide variations that impair function. The combination of lower number and increased frequency of mutations and deletions may result in inadequate mitochondrial activity necessary for continued embryo development and cause pregnancy failure. Previous reports suggested that mitochondrial activity within oocytes may be supplemented by donor cytoplasmic transfer at the time of intracytoplasmic sperm injection (ICSI. Those reports showed success; however, safety concerns arose due to the potential of two distinct populations of mitochondrial genomes in the offspring. Mitochondrial augmentation of oocytes is now reconsidered in light of our current understanding of mitochondrial function and the publication of a number of animal studies. With a better understanding of the role of this organelle in oocytes immediately after fertilization, blastocyst and offspring, mitochondrial augmentation may be reconsidered as a method to improve oocyte quality.

  13. Molecular design of materials for cell separation.

    Science.gov (United States)

    Kataoka, K

    1988-12-01

    There has been a strong demand in biomedical sciences to isolate viable cell populations with high yield and purity. An important facet of this work was to develop new polymeric adsorbent for the separation of lymphocyte subpopulations. Based on our strategy of separating cells through their differential ionic affinity toward multiphase-structured adsorbent with ionically derivatized microdomains, a series of poly(2-hydroxyethyl methacrylate)/polyamine graft copolymers (HA copolymers) was prepared. HA copolymer columns were found to show specific adsorption affinity toward B lymphocytes, and allows for separation of B and T lymphocytes in high yield and purity with a short operating time. Separation mechanism involved in the resolution of B and T lymphocytes by HA copolymer column is discussed in this paper. Further, photo-induced desorption of cells from the adsorbent derivatized with photo-responsive functional group (azobenzene group) was demonstrated to emphasize the feasibility of photo-regulated chromatography as a novel tool in cell separation technology.

  14. Cybernetically sound organizational structures II: Relating de Sitter's design theory to Beer's viable system model

    NARCIS (Netherlands)

    Achterbergh, J.M.I.M.; Vriens, D.J.

    2011-01-01

    - Purpose – The purpose of this paper is to show how the viable system model (VSM) and de Sitter's design theory can complement each other in the context of the diagnosis and design of viable organizations. - Design/methodology/approach – Key concepts from Beer's model and de Sitter's design

  15. Transcription of mouse Sp2 yields alternatively spliced and sub-genomic mRNAs in a tissue- and cell type-specific fashion

    Science.gov (United States)

    Yin, Haifeng; Nichols, Teresa D.; Horowitz, Jonathan M.

    2010-01-01

    The Sp-family of transcription factors is comprised by nine members, Sp1-9, that share a highly-conserved DNA-binding domain. Sp2 is a poorly characterized member of this transcription factor family that is widely expressed in murine and human cell lines yet exhibits little DNA-binding or trans-activation activity in these settings. As a prelude to the generation of a “knock-out” mouse strain, we isolated a mouse Sp2 cDNA and performed a detailed analysis of Sp2 transcription in embryonic and adult mouse tissues. We report that (1) the 5′ untranslated region of Sp2 is subject to alternative splicing, (2) Sp2 transcription is regulated by at least two promoters that differ in their cell-type specificity, (3) one Sp2 promoter is highly active in nine mammalian cell lines and strains and is regulated by at least five discrete stimulatory and inhibitory elements, (4) a variety of sub-genomic messages are synthesized from the Sp2 locus in a tissue- and cell type-specific fashion and these transcripts have the capacity to encode a novel partial-Sp2 protein, and (5) RNA in situ hybridization assays indicate that Sp2 is widely expressed during mouse embryogenesis, particularly in the embryonic brain, and robust Sp2 expression occurs in neurogenic regions of the post-natal and adult brain. PMID:20353838

  16. Hymenolepis nana: immunity against oncosphere challenge in mice previously given viable or non-viable oncospheres of H. nana, H. diminuta, H. microstoma and Taenia taeniaeformis.

    Science.gov (United States)

    Ito, A; Onitake, K; Sasaki, J; Takami, T

    1991-04-01

    When mice, previously given oral inoculation with viable oncospheres of the heterologous cestode species (Hymenolepis diminuta, H. microstoma, Taenia taeniaeformis) and the homologous one (H. nana), were challenged with oncospheres of H. nana 4 days after the primary inoculation, they showed strong and complete resistance to H. nana challenge, respectively. However, the resistance was not evoked in mice given either infective eggs of Toxocara canis or non-viable oncospheres of all cestode species examined. Congenitally athymic nude mice given viable oncospheres did not show any resistance to H. nana either. Eosinophil infiltration around cysticercoids of H. nana in the intestinal villi appeared to be more prominent in mice previously given viable oncospheres of H. diminuta than in mice given non-viable oncospheres or PBS only. Some of the eosinophils in the villus harboring cysticercoid(s) of H. nana invaded the epithelia in the former, whereas all eosinophils remained in the lamina propria in the latter. There was almost no eosinophil infiltration in nude mice. Microscopic observations revealed that oncospheres of H. diminuta, which require beetles as the intermediate host like H. microstoma, could invade the mouse intestinal tissue. Therefore, it is strongly suggested that the strong cross resistance to H. nana in mice, induced by oncospheres of all heterologous cestode species, is thymus-dependent and due to oncospheral invasion into the intestinal tissue of mice.

  17. A Rapid Method for Quantifying Viable Mycobacterium avium subsp. paratuberculosis in Cellular Infection Assays

    Science.gov (United States)

    Pooley, Hannah B.; de Silva, Kumudika; Purdie, Auriol C.; Begg, Douglas J.; Whittington, Richard J.

    2016-01-01

    ABSTRACT Determining the viability of bacteria is a key outcome of in vitro cellular infection assays. Currently, this is done by culture, which is problematic for fastidious slow-growing bacteria such as Mycobacterium avium subsp. paratuberculosis, where it can take up to 4 months to confirm growth. This study aimed to identify an assay that can rapidly quantify the number of viable M. avium subsp. paratuberculosis cells in a cellular sample. Three commercially available bacterial viability assays along with a modified liquid culture method coupled with high-throughput quantitative PCR growth detection were assessed. Criteria for assessment included the ability of each assay to differentiate live and dead M. avium subsp. paratuberculosis organisms and their accuracy at low bacterial concentrations. Using the culture-based method, M. avium subsp. paratuberculosis growth was reliably detected and quantified within 2 weeks. There was a strong linear association between the 2-week growth rate and the initial inoculum concentration. The number of viable M. avium subsp. paratuberculosis cells in an unknown sample was quantified based on the growth rate, by using growth standards. In contrast, none of the commercially available viability assays were suitable for use with samples from in vitro cellular infection assays. IMPORTANCE Rapid quantification of the viability of Mycobacterium avium subsp. paratuberculosis in samples from in vitro cellular infection assays is important, as it allows these assays to be carried out on a large scale. In vitro cellular infection assays can function as a preliminary screening tool, for vaccine development or antimicrobial screening, and also to extend findings derived from experimental animal trials. Currently, by using culture, it takes up to 4 months to obtain quantifiable results regarding M. avium subsp. paratuberculosis viability after an in vitro infection assay; however, with the quantitative PCR and liquid culture method

  18. Nut crop yield records show that budbreak-based chilling requirements may not reflect yield decline chill thresholds

    Science.gov (United States)

    Pope, Katherine S.; Dose, Volker; Da Silva, David; Brown, Patrick H.; DeJong, Theodore M.

    2015-06-01

    Warming winters due to climate change may critically affect temperate tree species. Insufficiently cold winters are thought to result in fewer viable flower buds and the subsequent development of fewer fruits or nuts, decreasing the yield of an orchard or fecundity of a species. The best existing approximation for a threshold of sufficient cold accumulation, the "chilling requirement" of a species or variety, has been quantified by manipulating or modeling the conditions that result in dormant bud breaking. However, the physiological processes that affect budbreak are not the same as those that determine yield. This study sought to test whether budbreak-based chilling thresholds can reasonably approximate the thresholds that affect yield, particularly regarding the potential impacts of climate change on temperate tree crop yields. County-wide yield records for almond ( Prunus dulcis), pistachio ( Pistacia vera), and walnut ( Juglans regia) in the Central Valley of California were compared with 50 years of weather records. Bayesian nonparametric function estimation was used to model yield potentials at varying amounts of chill accumulation. In almonds, average yields occurred when chill accumulation was close to the budbreak-based chilling requirement. However, in the other two crops, pistachios and walnuts, the best previous estimate of the budbreak-based chilling requirements was 19-32 % higher than the chilling accumulations associated with average or above average yields. This research indicates that physiological processes beyond requirements for budbreak should be considered when estimating chill accumulation thresholds of yield decline and potential impacts of climate change.

  19. Fusion of antigen to a dendritic cell targeting chemokine combined with adjuvant yields a malaria DNA vaccine with enhanced protective capabilities.

    Science.gov (United States)

    Luo, Kun; Zhang, Hong; Zavala, Fidel; Biragyn, Arya; Espinosa, Diego A; Markham, Richard B

    2014-01-01

    Although sterilizing immunity to malaria can be elicited by irradiated sporozoite vaccination, no clinically practical subunit vaccine has been shown to be capable of preventing the approximately 600,000 annual deaths attributed to this infection. DNA vaccines offer several potential advantages for a disease that primarily affects the developing world, but new approaches are needed to improve the immunogenicity of these vaccines. By using a novel, lipid-based adjuvant, Vaxfectin, to attract immune cells to the immunization site, in combination with an antigen-chemokine DNA construct designed to target antigen to immature dendritic cells, we elicited a humoral immune response that provided sterilizing immunity to malaria challenge in a mouse model system. The chemokine, MIP3αCCL20, did not significantly enhance the cellular infiltrate or levels of cytokine or chemokine expression at the immunization site but acted with Vaxfectin to reduce liver stage malaria infection by orders of magnitude compared to vaccine constructs lacking the chemokine component. The levels of protection achieved were equivalent to those observed with irradiated sporozoites, a candidate vaccine undergoing development for further large scale clinical trial. Only vaccination with the combined regimen of adjuvant and chemokine provided 80-100% protection against the development of bloodstream infection. Treating the immunization process as requiring the independent steps of 1) attracting antigen-presenting cells to the site of immunization and 2) specifically directing vaccine antigen to the immature dendritic cells that initiate the adaptive immune response may provide a rational strategy for the development of a clinically applicable malaria DNA vaccine.

  20. 3D rotating wall vessel and 2D cell culture of four veterinary virus pathogens: A comparison of virus yields, portions of infectious particles and virus growth curves.

    Science.gov (United States)

    Malenovská, Hana

    2016-02-01

    Only very few comparative studies have been performed that evaluate general trends of virus growth under 3D in comparison with 2D cell culture conditions. The aim of this study was to investigate differences when four animal viruses are cultured in 2D and 3D. Suid herpesvirus 1 (SuHV-1), Vesicular stomatitis virus (VSIV), Bovine adenovirus (BAdV) and Bovine parainfluenza 3 virus (BPIV-3) were cultivated in 3D rotating wall vessels (RWVs) and conventional 2D cultures. The production of virus particles, the portion of infectious particles, and the infectious growth curves were compared. For all viruses, the production of virus particles (related to cell density), including the non-infectious ones, was lower in 3D than in 2D culture. The production of only infectious particles was significantly lower in BAdV and BPIV-3 in 3D cultures in relation to cell density. The two cultivation approaches resulted in significantly different virus particle-to-TCID50 ratios in three of the four viruses: lower in SuHV-1 and BPIV-3 and higher in BAdV in 3D culture. The infectious virus growth rates were not significantly different in all viruses. Although 3D RWV culture resulted in lower production of virus particles compared to 2D systems, the portion of infectious particles was higher for some viruses. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. In Vivo Selection of a Computationally Designed SCHEMA AAV Library Yields a Novel Variant for Infection of Adult Neural Stem Cells in the SVZ.

    Science.gov (United States)

    Ojala, David S; Sun, Sabrina; Santiago-Ortiz, Jorge L; Shapiro, Mikhail G; Romero, Philip A; Schaffer, David V

    2017-09-08

    Directed evolution continues to expand the capabilities of complex biomolecules for a range of applications, such as adeno-associated virus vectors for gene therapy; however, advances in library design and selection strategies are key to develop variants that overcome barriers to clinical translation. To address this need, we applied structure-guided SCHEMA recombination of the multimeric adeno-associated virus (AAV) capsid to generate a highly diversified chimeric library with minimal structural disruption. A stringent in vivo Cre-dependent selection strategy was implemented to identify variants that transduce adult neural stem cells (NSCs) in the subventricular zone. A novel variant, SCH9, infected 60% of NSCs and mediated 24-fold higher GFP expression and a 12-fold greater transduction volume than AAV9. SCH9 utilizes both galactose and heparan sulfate as cell surface receptors and exhibits increased resistance to neutralizing antibodies. These results establish the SCHEMA library as a valuable tool for directed evolution and SCH9 as an effective gene delivery vector to investigate subventricular NSCs. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

  2. Predicting yields for autotrophic and cometabolic processes

    Energy Technology Data Exchange (ETDEWEB)

    Andrews, G. [Idaho National Engineering Lab., Idaho Falls, ID (United States)

    1995-12-31

    The goal of bioprocess engineering is to state how the optimum design and control strategy for a bioprocess follow from the metabolism of the particular microorganism. A necessary step toward this goal is to show how the parameters used in quantitative descriptions of a process (e.g., yield and maintenance coefficients) are related to those describing the metabolism [e.g., Y{sub ATP}, (P/O)]. The {open_quotes}yield equation{close_quotes} approach to this problem involves dividing metabolism into the separate pathways for catabolism, anabolism, respiration, and product formation and balancing the production and consumption of reducing equivalents and ATP. The general approach, demonstrated previously for heterotrophic cell growth and products of fermentation, is illustrated by three new examples: the cell yield for chemoautotrophic iron-oxidizing bacteria, the cometabolic degradation of chloroform by methanotrophic bacteria, and the theoretical yield of succinic acid from glucose.

  3. Vibrio cholerae classical biotype is converted to the viable non-culturable state when cultured with the El Tor biotype.

    Science.gov (United States)

    Pradhan, Subhra; Mallick, Sanjaya K; Chowdhury, Rukhsana

    2013-01-01

    A unique event in bacterial epidemiology was the emergence of the El Tor biotype of Vibrio cholerae O1 and the subsequent rapid displacement of the existing classical biotype as the predominant cause of epidemic cholera. We demonstrate that when the El Tor and classical biotypes were cocultured in standard laboratory medium a precipitous decline in colony forming units (CFU) of the classical biotype occurred in a contact dependent manner. Several lines of evidence including DNA release, microscopy and flow cytometric analysis indicated that the drastic reduction in CFU of the classical biotype in cocultures was not accompanied by lysis, although when the classical biotype was grown individually in monocultures, lysis of the cells occurred concomitant with decrease in CFU starting from late stationary phase. Furthermore, uptake of a membrane potential sensitive dye and protection of genomic DNA from extracellular DNase strongly suggested that the classical biotype cells in cocultures retained viability in spite of loss of culturability. These results suggest that coculturing the classical biotype with the El Tor biotype protects the former from lysis allowing the cells to remain viable in spite of the loss of culturability. The stationary phase sigma factor RpoS may have a role in the loss of culturability of the classical biotype in cocultures. Although competitive exclusion of closely related strains has been reported for several bacterial species, conversion of the target bacterial population to the viable non-culturable state has not been demonstrated previously and may have important implications in the evolution of bacterial strains.

  4. Phylogenetic analysis of pectin-related gene families in Physcomitrella patens and nine other plant species yields evolutionary insights into cell walls.

    Science.gov (United States)

    McCarthy, Thomas W; Der, Joshua P; Honaas, Loren A; dePamphilis, Claude W; Anderson, Charles T

    2014-03-26

    Pectins are acidic sugar-containing polysaccharides that are universally conserved components of the primary cell walls of plants and modulate both tip and diffuse cell growth. However, many of their specific functions and the evolution of the genes responsible for producing and modifying them are incompletely understood. The moss Physcomitrella patens is emerging as a powerful model system for the study of plant cell walls. To identify deeply conserved pectin-related genes in Physcomitrella, we generated phylogenetic trees for 16 pectin-related gene families using sequences from ten plant genomes and analyzed the evolutionary relationships within these families. Contrary to our initial hypothesis that a single ancestral gene was present for each pectin-related gene family in the common ancestor of land plants, five of the 16 gene families, including homogalacturonan galacturonosyltransferases, polygalacturonases, pectin methylesterases, homogalacturonan methyltransferases, and pectate lyase-like proteins, show evidence of multiple members in the early land plant that gave rise to the mosses and vascular plants. Seven of the gene families, the UDP-rhamnose synthases, UDP-glucuronic acid epimerases, homogalacturonan galacturonosyltransferase-like proteins, β-1,4-galactan β-1,4-galactosyltransferases, rhamnogalacturonan II xylosyltransferases, and pectin acetylesterases appear to have had a single member in the common ancestor of land plants. We detected no Physcomitrella members in the xylogalacturonan xylosyltransferase, rhamnogalacturonan I arabinosyltransferase, pectin methylesterase inhibitor, or polygalacturonase inhibitor protein families. Several gene families related to the production and modification of pectins in plants appear to have multiple members that are conserved as far back as the common ancestor of mosses and vascular plants. The presence of multiple members of these families even before the divergence of other important cell wall-related genes

  5. The effect of formulation on the penetration of coated and uncoated zinc oxide nanoparticles into the viable epidermis of human skin in vivo.

    Science.gov (United States)

    Leite-Silva, Vânia R; Le Lamer, Marina; Sanchez, Washington Y; Liu, David C; Sanchez, Washington H; Morrow, Isabel; Martin, Darren; Silva, Heron D T; Prow, Tarl W; Grice, Jeffrey E; Roberts, Michael S

    2013-06-01

    The use of nanoparticulate zinc oxide (ZnO-NP) in sunscreens and other cosmetic products has raised public health concerns. The two key issues are the extent of exposure to ZnO-NP and the likely hazard after the application of ZnO-NP in sunscreen and cosmetic products to humans in vivo. Our aims were to assess exposure by the extent of ZnO-NP penetration into the viable epidermis and hazard by changes in the viable epidermal redox state for a number of topical products. Of particular interest is the role of the particle coating, formulation used, and the presence of any enhancers. Multiphoton tomography with fluorescence lifetime imaging microscopy (MPT-FLIM) was used to simultaneously observe ZnO-NP penetration and potential metabolic changes within the viable epidermis of human volunteers after topical application of various ZnO-NP products. Coated and uncoated ZnO-NP remained in the superficial layers of the SC and in the skin furrows. We observed limited penetration of coated ZnO-NP dispersed in a water-in-oil emulsion formulation, which was predominantly localized adjacent to the skin furrow. However, the presence of ZnO-NP in the viable epidermis did not alter the metabolic state or morphology of the cells. In summary, our data suggest that some limited penetration of coated and uncoated ZnO-NP may occur into viable stratum granulosum epidermis adjacent to furrows, but that the extent is not sufficient to affect the redox state of those viable cells. Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.

  6. Growth Phase, Oxygen, Temperature and Starvation Affect the Development of Viable but Non-Culturable State of Vibrio cholerae

    Directory of Open Access Journals (Sweden)

    Bin eWu

    2016-03-01

    Full Text Available AbstractVibrio cholerae can enter into a viable but non-culturable (VBNC state in order to survive in unfavourable environments. In this study, we studied the roles of five physicochemical and microbiological factors or states, namely, different strains, growth phases, oxygen, temperature, and starvation, on the development of VBNC of V. cholerae in artificial sea water (ASW. Different strains of the organism, the growth phase, and oxygen levels affected the progress of VBNC development. It was found that the VBNC state was induced faster in V. cholerae serogroup O1 classical biotype strain O395 than in O1 El Tor biotype strains C6706 and N16961. When cells in different growth phases were used for VBNC induction, stationary-phase cells lost their culturability more quickly than exponential-phase cells, while induction of a totally non-culturable state took longer to achieve for stationary-phase cells in all three strains, suggesting that heterogeneity of cells should be considered. Aeration strongly accelerated the loss of culturability. During the development of the VBNC state, the culturable cell count under aeration conditions was almost 106-fold lower than under oxygen-limited conditions for all three strains. The other two factors, temperature and nutrients-rich environment, may prevent the induction of VBNC cells. At 22°C or 37°C in ASW, most of the cells rapidly died and the culturable cell count reduced from about 108 CFU/mL to 106–105 CFU/mL. The total cell counts showed that cells that lost viability were decomposed, and the viable cell counts were the same as culturable cell counts, indicating that the cells did not reach the VBNC state. VBNC state development was blocked when ASW was supplied with Luria-Bertani broth (LB, but it was not affected in ASW with M9, suggesting that specific nutrients in LB may prevent the development of VBNC state. These results revealed that the five factors evaluated in this study had different

  7. Detection and Quantification of Viable and Nonviable Trypanosoma cruzi Parasites by a Propidium Monoazide Real-Time Polymerase Chain Reaction Assay

    Science.gov (United States)

    Cancino-Faure, Beatriz; Fisa, Roser; Alcover, M. Magdalena; Jimenez-Marco, Teresa; Riera, Cristina

    2016-01-01

    Molecular techniques based on real-time polymerase chain reaction (qPCR) allow the detection and quantification of DNA but are unable to distinguish between signals from dead or live cells. Because of the lack of simple techniques to differentiate between viable and nonviable cells, the aim of this study was to optimize and evaluate a straightforward test based on propidium monoazide (PMA) dye action combined with a qPCR assay (PMA-qPCR) for the selective quantification of viable/nonviable epimastigotes of Trypanosoma cruzi. PMA has the ability to penetrate the plasma membrane of dead cells and covalently cross-link to the DNA during exposure to bright visible light, thereby inhibiting PCR amplification. Different concentrations of PMA (50–200 μM) and epimastigotes of the Maracay strain of T. cruzi (1 × 105–10 parasites/mL) were assayed; viable and nonviable parasites were tested and quantified by qPCR with a TaqMan probe specific for T. cruzi. In the PMA-qPCR assay optimized at 100 μM PMA, a significant qPCR signal reduction was observed in the nonviable versus viable epimastigotes treated with PMA, with a mean signal reduction of 2.5 logarithm units and a percentage of signal reduction > 98%, in all concentrations of parasites assayed. This signal reduction was also observed when PMA-qPCR was applied to a mixture of live/dead parasites, which allowed the detection of live cells, except when the concentration of live parasites was low (10 parasites/mL). The PMA-qPCR developed allows differentiation between viable and nonviable epimastigotes of T. cruzi and could thus be a potential method of parasite viability assessment and quantification. PMID:27139452

  8. Overexpressing Ferredoxins in Chlamydomonas reinhardtii Increase Starch and Oil Yields and Enhance Electric Power Production in a Photo Microbial Fuel Cell

    Directory of Open Access Journals (Sweden)

    Li-Fen Huang

    2015-08-01

    Full Text Available Ferredoxins (FDX are final electron carrier proteins in the plant photosynthetic pathway, and function as major electron donors in diverse redox-driven metabolic pathways. We previously showed that overexpression of a major constitutively expressed ferredoxin gene PETF in Chlamydomonas decreased the reactive oxygen species (ROS level and enhanced tolerance to heat stress. In addition to PETF, an endogenous anaerobic induced FDX5 was overexpressed in transgenic Chlamydomonas lines here to address the possible functions of FDX5. All the independent FDX transgenic lines showed decreased cellular ROS levels and enhanced tolerance to heat and salt stresses. The transgenic Chlamydomonas lines accumulated more starch than the wild-type line and this effect increased almost three-fold in conditions of nitrogen depletion. Furthermore, the lipid content was higher in the transgenic lines than in the wild-type line, both with and without nitrogen depletion. Two FDX-overexpressing Chlamydomonas lines were assessed in a photo microbial fuel cell (PMFC; power density production by the transgenic lines was higher than that of the wild-type cells. These findings suggest that overexpression of either PETF or FDX5 can confer tolerance against heat and salt stresses, increase starch and oil production, and raise electric power density in a PMFC.

  9. Survival of Erwinia amylovora in mature apple fruit calyces through the viable but nonculturable (VBNC) state.

    Science.gov (United States)

    Ordax, M; Biosca, E G; Wimalajeewa, S C; López, M M; Marco-Noales, E

    2009-07-01

    Survival of Erwinia amylovora, causal agent of fire blight in pome fruits and other rosaceous plants, was monitored inside mature apples calyces under some storage conditions utilized in fruit. Apple fruit calyces inoculated with two E. amylovora strains and their respective GFP-marked strains were maintained at 26 degrees and 5 degrees C, and the effect of copper treatment was assayed at 0.01 and 0.1 mmol l(-1) CuSO4. In nontreated apples at 26 degrees C, part of the population of E. amylovora survived in the 'viable but nonculturable' (VBNC) state, whereas at 5 degrees C the majority of the population retained culturability. In copper-treated apples, the whole population adopted the VBNC state irrespective of temperature. Regardless of temperature, copper and inoculum dose, VBNC cells recovered culturability and pathogenicity in King's B broth or by host plant passage. Erwinia amylovora survived for at least 35 days in mature apple calyces. Besides, the ability of the pathogen in the VBNC state to regain culturability and pathogenicity suggests that the apple fruit could be a potential carrier of E. amylovora contributing to the spreading of fire blight disease. The risk of E. amylovora dissemination through mature fruit transport, although low, has been demonstrated, and should be considered in pest risk assessments.

  10. High-yield production of scutellaria radix flavonoids (baicalein, baicalin and wogonin by liquid-culture of Scutellaria baicalensis root-derived cells

    Directory of Open Access Journals (Sweden)

    Takashi Ohtsuki

    2009-04-01

    Full Text Available Production of baicalein, baicalin and wogonin by liquid culture of Scutellaria baicalensis cells derived from the plant root was studied. The maximum production obtained were 119 mg/L of baicalein at two week, 1372 mg/L of baicalin at eight week, and 14 mg/L of wogonin at two week. In addition, the production of baicalin was drastically increased to 1000 mg/L level at 3-week culture, and the extremely high production rate (339 mg/L•week was obtained. In the comparison of total antioxidative activities among baicalein, baicalin and wogonin, evaluated by thiocyanate method, it was suggested that the location of hydroxyl groups both at 5- and 6-position contributed to enhancement of radical scavenging activity, and/or methoxylation at 8-position diminished the activity. The possibility of utilizing these flavonoids for natural antioxidants and medicine is also discussed.

  11. Origin of apparent viscosity in yield stress fluids below yielding

    NARCIS (Netherlands)

    Møller, P.C.F.; Fall, A.; Bonn, D.

    2009-01-01

    For more than 20 years it has been debated if yield stress fluids are solid below the yield stress or actually flow; whether true yield stress fluids exist or not. Advocates of the true yield stress picture have demonstrated that the effective viscosity increases very rapidly as the stress is

  12. Detection of viable Helicobacter pylori inside free-living amoebae in wastewater and drinking water samples from Eastern Spain.

    Science.gov (United States)

    Moreno-Mesonero, Laura; Moreno, Yolanda; Alonso, José Luis; Ferrús, M Antonia

    2017-10-01

    Helicobacter pylori is one of the most concerning emerging waterborne pathogens. It has been suggested that it could survive in water inside free-living amoebae (FLA), but nobody has studied this relationship in the environment yet. Thus, we aimed to detect viable H. pylori cells from inside FLA in water samples. Sixty-nine wastewater and 31 drinking water samples were collected. FLA were purified and identified by PCR and sequencing. For exclusively detecting H. pylori inside FLA, samples were exposed to sodium hypochlorite and assayed by specific PMA-qPCR, DVC-FISH and culture. FLA were detected in 38.7% of drinking water and 79.7% of wastewater samples, even after disinfection. In wastewater, Acanthamoeba spp. and members of the family Vahlkampfiidae were identified. In drinking water, Acanthamoeba spp. and Echinamoeba and/or Vermamoeba were present. In 39 (58.2%) FLA-positive samples, H. pylori was detected by PMA-qPCR. After DVC-FISH, 21 (31.3%) samples harboured viable H. pylori internalized cells. H. pylori was cultured from 10 wastewater samples. To our knowledge, this is the first report that demonstrates that H. pylori can survive inside FLA in drinking water and wastewater, strongly supporting the hypothesis that FLA could play an important role in the transmission of H. pylori to humans. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  13. Escherichia coli detection using mTEC agar and fluorescent antibody direct viable counting on coastal recreational water samples.

    Science.gov (United States)

    Zimmerman, A M; Rebarchik, D M; Flowers, A R; Williams, J L; Grimes, D J

    2009-10-01

    Escherichia coli is the faecal indicator species recommended by the US Environmental Protection Agency (USEPA) for monitoring fresh recreational water. Viable but nonculturable (VBNC) E. coli are living cells that are dormant and not culturable using standard microbiological cultivation methods. This study reports a comparison between the mTEC culture method recommended by USEPA for E. coli enumeration and a fluorescent antibody-direct viable count (FA-DVC) method to visualize living E. coli cells with a microscope. Escherichia coli, faecal coliforms and Enterococcus were detected using standard methods recommended by the USEPA. VBNC E. coli was visualized with FA-DVC. Results were analysed with standard statistical methods (Pearson correlation; paired-sample t-test). Significantly higher numbers of E. coli were detected using the FA-DVC method than using the mTEC method. Escherichia coli results were also compared with faecal coliform (mFC broth) and Enterococcus (mEI agar) counts in the same samples. The results of this comparative study demonstrate that E. coli can be present in higher numbers than what are detected with standard culture methods. This study re-emphasizes the need for a rapid, accurate and precise method for detecting health risks to humans who use recreational waters.

  14. Triaxial testing beyond yielding

    DEFF Research Database (Denmark)

    Sabaliauskas, Tomas; Ibsen, Lars Bo

    2017-01-01

    This paper is continuation of work published at ISOPE 2015, where capabilities of undrained triaxial testing were presented. Now, drained loading is emphasized, recovery of disturbed sand properties is observed. After liquefying or yielding, sand becomes disturbed: stiffness and resistance...... to liquefaction become compromised. However, sand can "heal" itself. It can recover during drained deformation cycles. The recovery mechanism can be observed using a triaxial apparatus. Such tests are relevant for offshore, seismic, and other fields of engineering, where disturbed soil states are encountered....

  15. Scanning number and brightness yields absolute protein concentrations in live cells: a crucial parameter controlling functional bio-molecular interaction networks.

    Science.gov (United States)

    Papini, Christina; Royer, Catherine A

    2018-02-01

    Biological function results from properly timed bio-molecular interactions that transduce external or internal signals, resulting in any number of cellular fates, including triggering of cell-state transitions (division, differentiation, transformation, apoptosis), metabolic homeostasis and adjustment to changing physical or nutritional environments, amongst many more. These bio-molecular interactions can be modulated by chemical modifications of proteins, nucleic acids, lipids and other small molecules. They can result in bio-molecular transport from one cellular compartment to the other and often trigger specific enzyme activities involved in bio-molecular synthesis, modification or degradation. Clearly, a mechanistic understanding of any given high level biological function requires a quantitative characterization of the principal bio-molecular interactions involved and how these may change dynamically. Such information can be obtained using fluctation analysis, in particular scanning number and brightness, and used to build and test mechanistic models of the functional network to define which characteristics are the most important for its regulation.

  16. Clinically Viable Gene Expression Assays with Potential for Predicting Benefit from MEK Inhibitors.

    Science.gov (United States)

    Brant, Roz; Sharpe, Alan; Liptrot, Tom; Dry, Jonathan R; Harrington, Elizabeth A; Barrett, J Carl; Whalley, Nicky; Womack, Christopher; Smith, Paul; Hodgson, Darren R

    2017-03-15

    Purpose: To develop a clinically viable gene expression assay to measure RAS/RAF/MEK/ERK (RAS-ERK) pathway output suitable for hypothesis testing in non-small cell lung cancer (NSCLC) clinical studies. Experimental Design: A published MEK functional activation signature (MEK signature) that measures RAS-ERK functional output was optimized for NSCLC in silico NanoString assays were developed for the NSCLC optimized MEK signature and the 147-gene RAS signature. First, platform transfer from Affymetrix to NanoString, and signature modulation following treatment with KRAS siRNA and MEK inhibitor, were investigated in cell lines. Second, the association of the signatures with KRAS mutation status, dynamic range, technical reproducibility, and spatial and temporal variation was investigated in NSCLC formalin-fixed paraffin-embedded tissue (FFPET) samples. Results: We observed a strong cross-platform correlation and modulation of signatures in vitro Technical and biological replicates showed consistent signature scores that were robust to variation in input total RNA; conservation of scores between primary and metastatic tumor was statistically significant. There were statistically significant associations between high MEK ( P = 0.028) and RAS ( P = 0.003) signature scores and KRAS mutation in 50 NSCLC samples. The signatures identify overlapping but distinct candidate patient populations from each other and from KRAS mutation testing. Conclusions: We developed a technically and biologically robust NanoString gene expression assay of MEK pathway output, compatible with the quantities of FFPET routinely available. The gene signatures identified a different patient population for MEK inhibitor treatment compared with KRAS mutation testing. The predictive power of the MEK signature should be studied further in clinical trials. Clin Cancer Res; 23(6); 1471-80. ©2016 AACR See related commentary by Xue and Lito, p. 1365 . ©2016 American Association for Cancer Research.

  17. Immediate natural tooth pontic: A viable yet temporary prosthetic solution: A patient reported outcome

    Directory of Open Access Journals (Sweden)

    Sudhir Bhandari

    2012-01-01

    Conclusion: The concept of immediate pontic placement is surely a viable treatment option and promises an excellent transient esthetic solution for a lost tooth as well as enables good preparation of the extraction site for future prosthetic replacement.

  18. Improved identification of viable myocardium using second harmonic imaging during dobutamine stress echocardiography

    NARCIS (Netherlands)

    F. Sozzi (Fabiola); D. Poldermans (Don); J.J. Bax (Jeroen); A. Elhendy (Abdou); E.C. Vourvouri (Eleni); R. Valkema (Roelf); J. de Sutter; A.F.L. Schinkel (Arend); A. Borghetti; J.R.T.C. Roelandt (Jos)

    2001-01-01

    textabstractOBJECTIVE: To determine whether, compared with fundamental imaging, second harmonic imaging can improve the accuracy of dobutamine stress echocardiography for identifying viable myocardium, using nuclear imaging as a reference. PATIENTS: 30 patients with chronic left

  19. Marine environmental pollution stress detection through direct viable counts of bacteria

    Digital Repository Service at National Institute of Oceanography (India)

    Ramaiah, N.; Kenkre, V.D.; Verlecar, X.N.

    Direct viable counts (DVC) of bacteria were quantified from polluted and relatively less/non-polluted coastal locations during different seasons to assess whether they can be routinely monitored for an understanding of environmental stress(es...

  20. Optimization of culture conditions to produce high yields of active Acetobacter sp. CCTCC M209061 cells for anti-Prelog reduction of prochiral ketones

    Directory of Open Access Journals (Sweden)

    Chen Xiao-Hong

    2011-11-01

    Full Text Available Abstract Background Chiral alcohols are widely used in the synthesis of chiral pharmaceuticals, flavors and functional materials and appropriate whole-cell biocatalysts offer a highly enantioselective, minimally polluting route to these valuable compounds. The recently isolated strain Acetobacter sp. CCTCC M209061 showed exclusive anti-Prelog stereoselectivity for the reduction of prochiral ketones, but the low biomass has limited its commercialization and industrial applications. To tackle this problem, the effects of medium components and culture conditions on the strain's growth and reduction activity were explored. Results By using a one-at-a-time method and a central composite rotatable design (CCRD, the optimal medium and culture conditions were found to be as follows: glucose 8.26 g/L, fructose 2.50 g/L, soy peptone 83.92 g/L, MnSO4·H2O 0.088 g/L, pH 5.70, 30°C and 10% (v/v inoculum. Under the above-mentioned conditions, the biomass after 30 h cultivation reached 1.10 ± 0.03 g/L, which was 9.5-fold higher than that obtained with basic medium. Also, the reduction activity towards 4'-chloroacetophenone was markedly enhanced to 39.49 ± 0.96 μmol/min/g from 29.34 ± 0.65 μmol/min/g, with the product e.e. being above 99%. Comparable improvements were also seen with the enantioselective bioreduction of 4-(trimethylsilyl-3-butyn-2-one to the key pharmaceutical precursor (R - 4-(trimethylsilyl-3-butyn-2-ol. Conclusions The biomass and reduction activity of Acetobacter sp. CCTCC M209061 can be greatly enhanced through the optimization strategy. This facilitates use of the strain in the anti-Prelog stereoselective reduction of prochiral ketones to enantiopure chiral alcohols as building blocks for many industries.

  1. Validation of the NCCN-IPI for diffuse large B-cell lymphoma (DLBCL): the addition of β2-microglobulin yields a more accurate GELTAMO-IPI.

    Science.gov (United States)

    Montalbán, Carlos; Díaz-López, Antonio; Dlouhy, Ivan; Rovira, Jordina; Lopez-Guillermo, Armando; Alonso, Sara; Martín, Alejandro; Sancho, Juan M; García, Olga; Sánchez, Jose M; Rodríguez, Mario; Novelli, Silvana; Salar, Antonio; Gutiérrez, Antonio; Rodríguez-Salazar, Maria J; Bastos, Mariana; Domínguez, Juan F; Fernández, Rubén; Gonzalez de Villambrosia, Sonia; Queizan, José A; Córdoba, Raul; de Oña, Raquel; López-Hernandez, Andrés; Freue, Julian M; Garrote, Heidys; López, Lourdes; Martin-Moreno, Ana M; Rodriguez, Jose; Abraira, Víctor; García, Juan F

    2017-03-01

    The study included 1848 diffuse large B-cell lymphoma (DLBCL)patients treated with chemotherapy/rituximab. The aims were to validate the National Comprehensive Cancer Network International Prognostic Index (NCCN-IPI) and explore the effect of adding high Beta-2 microglobulin (β2M), primary extranodal presentation and intense treatment to the NCCN-IPI variables in order to develop an improved index. Comparing survival curves, NCCN-IPI discriminated better than IPI, separating four risk groups with 5-year overall survival rates of 93%, 83%, 67% and 49%, but failing to identify a true high-risk population. For the second aim the series was split into training and validation cohorts: in the former the multivariate model identified age, lactate dehydrogenase, Eastern Cooperative Oncology Group performance status, Stage III-IV, and β2M as independently significant, whereas the NCCN-IPI-selected extranodal sites, primary extranodal presentation and intense treatments were not. These results were confirmed in the validation cohort. The Grupo Español de Linfomas/Trasplante de Médula ósea (GELTAMO)-IPI developed here, with 7 points, significantly separated four risk groups (0, 1-3, 4 or ≥5 points) with 11%, 58%, 17% and 14% of patients, and 5-year overall survival rates of 93%, 79%, 66% and 39%, respectively. In the comparison GELTAMO IPI discriminated better than the NCCN-IPI. In conclusion, GELTAMO-IPI is more accurate than the NCCN-IPI and has statistical and practical advantages in that the better discrimination identifies an authentic high-risk group and is not influenced by primary extranodal presentation or treatments of different intensity. © 2017 John Wiley & Sons Ltd.

  2. Immunization of rodents against Hymenolepis infections using non-viable homologous oncospheres.

    Science.gov (United States)

    Fan, Ping-Chin; Chung, Wen-Cheng; Ito, Akira

    2004-12-01

    Immunity to Taiwan Taenia infection in pigs can be stimulated using homologous or heterologous non-viable Taenia oncospheres. This study was designed to determine whether homologous non-viable oncospheres could stimulate immunity to Hymenolepis infection in rodents. Hatched oncospheres were prepared from eggs of Hymenolepis diminuta, Hymenolepis nana, and Hymenolepis microstoma and kept at -70 degrees C for more than 1 month. A mixture of 500 non-viable oncospheres of each tapeworm and complete Freund's adjuvant was injected subcutaneously in four groups of Sprague-Dawley rats or ICR mice one to four times at an interval of 1 week; controls were not immunized. After immunization, each rodent was orally inoculated with three fresh active cysticercoids of H. diminuta or H. microstoma or 500 fresh eggs of H. nana. The animals were then necropsied for adult tapeworms. No rats or mice immunized with non-viable oncospheres of H. diminuta or H. nana were infected by the challenge inoculation. However, 28 of 34 mice immunized with non-viable H. microstoma oncospheres were infected after inoculation with cysticercoids. This study demonstrated complete protection against infection by homologous parasites in rats or mice immunized with non-viable oncospheres of H. diminuta and H. nana, respectively. Repeated immunization may not be required if resistance is stimulated in rodent hosts.

  3. [Viable myocardium detecting by CARTO voltage mapping in swine model of acute myocardial infarction].

    Science.gov (United States)

    Lin, Tao; Ma, Yi-Tong; Yang, Yi-Ning; Mu, Hu-Yati; He, Peng-Yi; Yang, Yu-Chun; Chou, Ping; Liu, Fen; Zhang, Yan-Yi

    2010-08-01

    To evaluate the accuracy and practicability of detecting viable myocardium by CARTO voltage mapping in swine model of acute myocardial infarction (MI). MI was induced in 13 anesthetized swines via occluding the distal of left anterior descending coronary arteries by angioplasty balloon for 60-90 minutes. The viable myocardium detection by CARTO voltage mapping was made after reconstruction of the left ventricle using CARTO and the results were compared with TTC staining. The standard of CARTO voltage to detect viable myocardium was 0.5 - 1.5 mV while viable myocardium showed pink color by TTC staining. Eleven out of 13 swines survived the operation and 2 swines died of ventricular fibrillation at 45 and 65 minutes post ischemia. Left ventricle was divided into 16 segments and 176 segments from 11 swines were analyzed. Viable myocardium detected by CARTO voltage mapping was identical as identified by TTC staining (Kappa = 0.816, P < 0.001). Taken the TTC result as standard, the sensitivity, specificity and accuracy rate of CARTO voltage mapping are 71.8%, 96.5% and 90.9% respectively. CARTO voltage mapping could be used as a reliable tool to detect viable myocardium in this model.

  4. “Limits of Control” – Crucial Parameters for a Reliable Quantification of Viable Campylobacter by Real-Time PCR

    Science.gov (United States)

    Krüger, Nora-Johanna; Buhler, Christiane; Iwobi, Azuka N.; Huber, Ingrid; Ellerbroek, Lüppo; Appel, Bernd; Stingl, Kerstin

    2014-01-01

    The unsuitability of the “CFU” parameter and the usefulness of cultivation-independent quantification of Campylobacter on chicken products, reflecting the actual risk for infection, is increasingly becoming obvious. Recently, real-time PCR methods in combination with the use of DNA intercalators, which block DNA amplification from dead bacteria, have seen wide application. However, much confusion exists in the correct interpretation of such assays. Campylobacter is confronted by oxidative and cold stress outside the intestine. Hence, damage caused by oxidative stress probably represents the most frequent natural death of Campylobacter on food products. Treatment of Campylobacter with peroxide led to complete loss of CFU and to significant entry of any tested DNA intercalator, indicating disruption of membrane integrity. When we transiently altered the metabolic state of Campylobacter by abolishing the proton-motive force or by inhibiting active efflux, CFU was constant but enhanced entry of ethidium bromide (EtBr) was observed. Consistently, ethidium monoazide (EMA) also entered viable Campylobacter, in particular when nutrients for bacterial energization were lacking (in PBS) or when the cells were less metabolically active (in stationary phase). In contrast, propidium iodide (PI) and propidium monoazide (PMA) were excluded from viable bacterial cells, irrespective of their metabolic state. As expected for a diffusion-limited process, the extent of signal reduction from dead cells depended on the temperature, incubation time and concentration of the dyes during staining, prior to crosslinking. Consistently, free protein and/or DNA present in varying amounts in the heterogeneous matrix lowered the concentration of the DNA dyes at the bacterial membrane and led to considerable variation of the residual signal from dead cells. In conclusion, we propose an improved approach, taking into account principles of method variability and recommend the implementation of

  5. "Limits of control"--crucial parameters for a reliable quantification of viable campylobacter by real-time PCR.

    Science.gov (United States)

    Krüger, Nora-Johanna; Buhler, Christiane; Iwobi, Azuka N; Huber, Ingrid; Ellerbroek, Lüppo; Appel, Bernd; Stingl, Kerstin

    2014-01-01

    The unsuitability of the "CFU" parameter and the usefulness of cultivation-independent quantification of Campylobacter on chicken products, reflecting the actual risk for infection, is increasingly becoming obvious. Recently, real-time PCR methods in combination with the use of DNA intercalators, which block DNA amplification from dead bacteria, have seen wide application. However, much confusion exists in the correct interpretation of such assays. Campylobacter is confronted by oxidative and cold stress outside the intestine. Hence, damage caused by oxidative stress probably represents the most frequent natural death of Campylobacter on food products. Treatment of Campylobacter with peroxide led to complete loss of CFU and to significant entry of any tested DNA intercalator, indicating disruption of membrane integrity. When we transiently altered the metabolic state of Campylobacter by abolishing the proton-motive force or by inhibiting active efflux, CFU was constant but enhanced entry of ethidium bromide (EtBr) was observed. Consistently, ethidium monoazide (EMA) also entered viable Campylobacter, in particular when nutrients for bacterial energization were lacking (in PBS) or when the cells were less metabolically active (in stationary phase). In contrast, propidium iodide (PI) and propidium monoazide (PMA) were excluded from viable bacterial cells, irrespective of their metabolic state. As expected for a diffusion-limited process, the extent of signal reduction from dead cells depended on the temperature, incubation time and concentration of the dyes during staining, prior to crosslinking. Consistently, free protein and/or DNA present in varying amounts in the heterogeneous matrix lowered the concentration of the DNA dyes at the bacterial membrane and led to considerable variation of the residual signal from dead cells. In conclusion, we propose an improved approach, taking into account principles of method variability and recommend the implementation of

  6. "Limits of control"--crucial parameters for a reliable quantification of viable campylobacter by real-time PCR.

    Directory of Open Access Journals (Sweden)

    Nora-Johanna Krüger

    Full Text Available The unsuitability of the "CFU" parameter and the usefulness of cultivation-independent quantification of Campylobacter on chicken products, reflecting the actual risk for infection, is increasingly becoming obvious. Recently, real-time PCR methods in combination with the use of DNA intercalators, which block DNA amplification from dead bacteria, have seen wide application. However, much confusion exists in the correct interpretation of such assays. Campylobacter is confronted by oxidative and cold stress outside the intestine. Hence, damage caused by oxidative stress probably represents the most frequent natural death of Campylobacter on food products. Treatment of Campylobacter with peroxide led to complete loss of CFU and to significant entry of any tested DNA intercalator, indicating disruption of membrane integrity. When we transiently altered the metabolic state of Campylobacter by abolishing the proton-motive force or by inhibiting active efflux, CFU was constant but enhanced entry of ethidium bromide (EtBr was observed. Consistently, ethidium monoazide (EMA also entered viable Campylobacter, in particular when nutrients for bacterial energization were lacking (in PBS or when the cells were less metabolically active (in stationary phase. In contrast, propidium iodide (PI and propidium monoazide (PMA were excluded from viable bacterial cells, irrespective of their metabolic state. As expected for a diffusion-limited process, the extent of signal reduction from dead cells depended on the temperature, incubation time and concentration of the dyes during staining, prior to crosslinking. Consistently, free protein and/or DNA present in varying amounts in the heterogeneous matrix lowered the concentration of the DNA dyes at the bacterial membrane and led to considerable variation of the residual signal from dead cells. In conclusion, we propose an improved approach, taking into account principles of method variability and recommend the

  7. Evaluation of propidium monoazide real-time PCR for early detection of viable Mycobacterium tuberculosis in clinical respiratory specimens.

    Science.gov (United States)

    Kim, Young Jin; Lee, Sun Min; Park, Byung Kyu; Kim, Sung Soo; Yi, Jongyoun; Kim, Hyung Hoi; Lee, Eun Yup; Chang, Chulhun Ludgerus

    2014-05-01

    Conventional acid-fast bacilli (AFB) staining cannot differentiate viable from dead cells. Propidium monoazide (PMA) is a photoreactive DNA-binding dye that inhibits PCR amplification by DNA modification. We evaluated whether PMA real-time PCR is suitable for the early detection of viable Mycobacterium tuberculosis (MTB) in clinical respiratory specimens. A total of 15 diluted suspensions from 5 clinical MTB isolates were quadruplicated and subjected to PMA treatment and/or heat inactivation. Eighty-three AFB-positive sputum samples were also tested to compare the ΔCT values (CT value in PMA-treated sputum samples-CT value in non-PMA-treated sputum samples) between culture-positive and culture-negative specimens. Real-time PCR was performed using Anyplex MTB/NTM Real-Time Detection (Seegene, Korea), and the CT value changes after PMA treatment were compared between culture-positive and culture-negative groups. In MTB suspensions, the increase in the CT value after PMA treatment was significant in dead cells (P=0.0001) but not in live cells (P=0.1070). In 14 culture-negative sputum samples, the median ΔCT value was 5.3 (95% confidence interval [CI], 4.1-8.2; P<0.0001), whereas that in 69 culture-positive sputum samples was 1.1 (95% CI, 0.7-2.0). In the ROC curve analysis, the cutoff ΔCT value for maximum sensitivity (89.9%) and specificity (85.7%) for differentiating dead from live cells was 3.4. PMA real-time PCR is a useful approach for differentiating dead from live bacilli in AFB smear-positive sputum samples.

  8. Detection of viable Salmonella in ice cream by TaqMan real-time polymerase chain reaction assay combining propidium monoazide.

    Science.gov (United States)

    Wang, Yuexia; Yang, Ming; Liu, Shuchun; Chen, Wanyi; Suo, Biao

    2015-09-01

    Real-time polymerase chain reaction (PCR) allows rapid detection of Salmonella in frozen dairy products, but it might cause a false positive detection result because it might amplify DNA from dead target cells as well. In this study, Salmonella-free frozen ice cream was initially inoculated with heat-killed Salmonella Typhimurium cells and stored at -18°C. Bacterial DNA extracted from the sample was amplified using TaqMan probe-based real-time PCR targeting the invA gene. Our results indicated that DNA from the dead cells remained stable in frozen ice cream for at least 20 days, and could produce fluorescence signal for real-time PCR as well. To overcome this limitation, propidium monoazide (PMA) was combined with real-time PCR. PMA treatment can effectively prevent PCR amplification from heat-killed Salmonella cells in frozen ice cream. The PMA real-time PCR assay can selectively detect viable Salmonella at as low as 103 CFU/mL. Combining 18 hours of pre-enrichment with the assay allows for the detection of viable Salmonella at 100 CFU/mL and avoiding the false-positive result of dead cells. The PMA real-time PCR assay provides an alternative specifically for detection of viable Salmonella in ice cream. However, when the PMA real-time PCR assay was evaluated in ice cream subjected to frozen storage, it obviously underestimated the contamination situation of viable Salmonella, which might lead to a false negative result. According to this result, the use of enrichment prior to PMA real-time PCR analysis remains as the more appropriate approach. Copyright © 2015. Published by Elsevier B.V.

  9. Detection of viable Salmonella in ice cream by TaqMan real-time polymerase chain reaction assay combining propidium monoazide

    Directory of Open Access Journals (Sweden)

    Yuexia Wang

    2015-09-01

    Full Text Available Real-time polymerase chain reaction (PCR allows rapid detection of Salmonella in frozen dairy products, but it might cause a false positive detection result because it might amplify DNA from dead target cells as well. In this study, Salmonella-free frozen ice cream was initially inoculated with heat-killed Salmonella Typhimurium cells and stored at −18°C. Bacterial DNA extracted from the sample was amplified using TaqMan probe-based real-time PCR targeting the invA gene. Our results indicated that DNA from the dead cells remained stable in frozen ice cream for at least 20 days, and could produce fluorescence signal for real-time PCR as well. To overcome this limitation, propidium monoazide (PMA was combined with real-time PCR. PMA treatment can effectively prevent PCR amplification from heat-killed Salmonella cells in frozen ice cream. The PMA real-time PCR assay can selectively detect viable Salmonella at as low as 103 CFU/mL. Combining 18 hours of pre-enrichment with the assay allows for the detection of viable Salmonella at 100 CFU/mL and avoiding the false-positive result of dead cells. The PMA real-time PCR assay provides an alternative specifically for detection of viable Salmonella in ice cream. However, when the PMA real-time PCR assay was evaluated in ice cream subjected to frozen storage, it obviously underestimated the contamination situation of viable Salmonella, which might lead to a false negative result. According to this result, the use of enrichment prior to PMA real-time PCR analysis remains as the more appropriate approach.

  10. Assessments of total and viable Escherichia coli O157:H7 on field and laboratory grown lettuce.

    Directory of Open Access Journals (Sweden)

    Anne-Laure Moyne

    Full Text Available Leafy green produce has been associated with numerous outbreaks of foodborne illness caused by strains of Escherichia coli O157:H7. While the amounts of culturable E. coli O157:H7 rapidly decline after introduction onto lettuce in the field, it remains to be determined whether the reduction in cell numbers is due to losses in cell viability, cell injury and a subsequent inability to be detected by standard laboratory culturing methods, or a lack of adherence and hence rapid removal of the organism from the plants during application. To assess which of these options is most relevant for E. coli O157:H7 on leafy green produce, we developed and applied a propidium monoazide (PMA real-time PCR assay to quantify viable (with PMA and total (without PMA E. coli O157:H7 cells on growth chamber and field-grown lettuce. E. coli O157:H7, suspended in 0.1% peptone, was inoculated onto 4-week-old lettuce plants at a level of approximately 10(6 CFU/plant. In the growth chamber at low relative humidity (30%, culturable amounts of the nontoxigenic E. coli O157:H7 strain ATCC 700728 and the virulent strain EC4045 declined 100 to 1000-fold in 24 h. Fewer E. coli O157:H7 cells survived when applied onto plants in droplets with a pipette compared with a fine spray inoculation. Total cells for both strains were equivalent to inoculum levels for 7 days after application, and viable cell quantities determined by PMA real-time PCR were approximately 10(4 greater than found by colony enumeration. Within 2 h after application onto plants in the field, the number of culturable E. coli ATCC 700728 was reduced by up to 1000-fold, whereas PCR-based assessments showed that total cell amounts were equivalent to inoculum levels. These findings show that shortly after inoculation onto plants, the majority of E. coli O157:H7 cells either die or are no longer culturable.

  11. Assessments of Total and Viable Escherichia coli O157:H7 on Field and Laboratory Grown Lettuce

    Science.gov (United States)

    Moyne, Anne-Laure; Harris, Linda J.; Marco, Maria L.

    2013-01-01

    Leafy green produce has been associated with numerous outbreaks of foodborne illness caused by strains of Escherichia coli O157:H7. While the amounts of culturable E. coli O157:H7 rapidly decline after introduction onto lettuce in the field, it remains to be determined whether the reduction in cell numbers is due to losses in cell viability, cell injury and a subsequent inability to be detected by standard laboratory culturing methods, or a lack of adherence and hence rapid removal of the organism from the plants during application. To assess which of these options is most relevant for E. coli O157:H7 on leafy green produce, we developed and applied a propidium monoazide (PMA) real-time PCR assay to quantify viable (with PMA) and total (without PMA) E. coli O157:H7 cells on growth chamber and field-grown lettuce. E. coli O157:H7, suspended in 0.1% peptone, was inoculated onto 4-week-old lettuce plants at a level of approximately 106 CFU/plant. In the growth chamber at low relative humidity (30%), culturable amounts of the nontoxigenic E. coli O157:H7 strain ATCC 700728 and the virulent strain EC4045 declined 100 to 1000-fold in 24 h. Fewer E. coli O157:H7 cells survived when applied onto plants in droplets with a pipette compared with a fine spray inoculation. Total cells for both strains were equivalent to inoculum levels for 7 days after application, and viable cell quantities determined by PMA real-time PCR were approximately 104 greater than found by colony enumeration. Within 2 h after application onto plants in the field, the number of culturable E. coli ATCC 700728 was reduced by up to 1000-fold, whereas PCR-based assessments showed that total cell amounts were equivalent to inoculum levels. These findings show that shortly after inoculation onto plants, the majority of E. coli O157:H7 cells either die or are no longer culturable. PMID:23936235

  12. {sup 177}Lu-DOTMP: a viable agent for palliative radiotherapy of painful bone metastasis

    Energy Technology Data Exchange (ETDEWEB)

    Das, T.; Chakraborty, S.; Banerjee, S. [Radiopharmaceuticals Div., Bhabha Atomic Research Centre, Mumbai (India); Sarma, H.D. [Radiation Biology and Health Sciences Div., Bhabha Atomic Research Centre, Mumbai (India)

    2008-07-01

    The suitable nuclear decay characteristics [T{sub 1/2} = 6.73 d, E{sub {beta}}{sub (max)} = 497 keV, E{sub {gamma}} = 113 keV (6.4%), 208 keV (11%)] as well as the feasibility of large-scale production with adequate specific activity and radionuclidic purity using a moderate flux reactor are important attributes towards {sup 177}Lu to be considered as a promising radionuclide for palliative care in painful bone metastasis. The present study describes the preparation of {sup 177}Lu complex of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetramethylene phosphonic acid (DOTMP) and its preliminary biological evaluation in animal models with an aim to proposing it as a viable radiopharmaceutical for bone pain palliation. The choice DOTMP as the polyaminophosphonic acid carrier ligand is based on the enhanced thermodynamic stability and kinetic inertness of the metal-ligand complexes with macrocyclic chelators. {sup 177}Lu was produced with a specific activity of {proportional_to} 12 GBq/mg ({proportional_to} 324 mCi/mg) and radionuclidic purity of 99.98% by irradiation of natural Lu{sub 2}O{sub 3} target at a thermal neutron flux of {proportional_to} 6 x 10{sup 13} n/cm{sup 2} s for 21 d. {sup 177}Lu-DOTMP complex was prepared in high yield and excellent radiochemical purity (> 99%) using DOTMP synthesized and characterized in-house. The complex exhibited excellent in-vitro stability at room temperature. Biodistribution studies in Wistar rats showed rapid skeletal accumulation of the injected activity [(1.60{+-}0.19)% per gram in femur at 3 h post-injection] with fast clearance from blood and minimal uptake in any of the major organs. Scintigraphic studies carried out in normal Wistar rats and New Zealand white rabbits also demonstrated significant accumulation of the agent in skeleton and almost no retention in any other vital organs. (orig.)

  13. Correlation Analysis of some Growth, Yield, Yield Components and ...

    African Journals Online (AJOL)

    3Department of Soil Science, Faculty of Agriculture, Usmanu Danfodiyo University, Sokoto, Nigeria ... The study was aimed at investigating the correlation between some growth, yield, yield components and ... Simple correlation coefficient ® of different crop parameters and grain yield indicated that most of the agro-.

  14. Heterosis and combining ability for grain yield and yield component ...

    African Journals Online (AJOL)

    Combining ability analysis for grain yield and yield component traits in maize were carried out in 8×8 diallel cross. The analysis of variance showed there is highly significant variation between the genotypes for all the traits considered. Year of testing was significant only for days to maturity and grain yield per hectare.

  15. Yield and yield components of six canola ( Brassica napus L ...

    African Journals Online (AJOL)

    The experiments were conducted in randomized complete block design arrangement in split factorial with four replications The results demonstrated that late planting date and interrupting of irrigation at flowering stage significantly decreased growth, yield and yield components the of canola cultivars. In addition, oil yield ...

  16. Relationships between yield and some yield components in Pea ...

    African Journals Online (AJOL)

    The experiment was designed randomized complete blocks design with four replications. At the end of the study, positive and significant relationship were found among seed yield and pods per plant and biological yield in both years. The strongest and direct positive effects were the biological yield (p = 0.6500), numbers of ...

  17. Melanocortin agonism as a viable strategy to control alveolar bone loss induced by oral infection.

    Science.gov (United States)

    Madeira, Mila F M; Queiroz-Junior, Celso M; Montero-Melendez, Trinidad; Werneck, Silvia M C; Corrêa, Jôice D; Soriani, Frederico M; Garlet, Gustavo P; Souza, Daniele G; Teixeira, Mauro M; Silva, Tarcilia A; Perretti, Mauro

    2016-12-01

    Alveolar bone loss is a result of an aggressive form of periodontal disease (PD) associated with Aggregatibacter actinomycetemcomitans (Aa) infection. PD is often observed with other systemic inflammatory conditions, including arthritis. Melanocortin peptides activate specific receptors to exert antiarthritic properties, avoiding excessing inflammation and modulating macrophage function. Recent work has indicated that melanocortin can control osteoclast development and function, but whether such protection takes place in infection-induced alveolar bone loss has not been investigated. The purpose of this study was to evaluate the role of melanocortin in Aa-induced PD. Mice were orally infected with Aa and treated with the melanocortin analog DTrp8-γMSH or vehicle daily for 30 d. Then, periodontal tissue was collected and analyzed. Aa-infected mice treated with DTrp8-γMSH presented decreased alveolar bone loss and a lower degree of neutrophil infiltration in the periodontium than vehicle-treated animals; these actions were associated with reduced periodontal levels of TNF-α, IFN-γ, and IL-17A. In vitro experiments with cells differentiated into osteoclasts showed that osteoclast formation and resorptive activity were attenuated after treatment with DTrp8-γMSH. Thus, melanocortin agonism could represent an innovative way to tame overexuberant inflammation and, at the same time, preserve bone physiology, as seen after Aa infection.-Madeira, M. F. M., Queiroz-Junior, C. M., Montero-Melendez, T., Werneck, S. M. C., Corrêa, J. D., Soriani, F. M., Garlet, G. P., Souza, D. G., Teixeira, M. M., Silva, T. A., Perretti, M. Melanocortin agonism as a viable strategy to control alveolar bone loss induced by oral infection. © FASEB.

  18. Experimental human-like model to assess the part of viable Legionella reaching the thoracic region after nebulization.

    Directory of Open Access Journals (Sweden)

    Jérémie Pourchez

    Full Text Available The incidence of Legionnaires' disease (LD in European countries and the USA has been constantly increasing since 1998. Infection of humans occurs through aerosol inhalation. To bridge the existing gap between the concentration of Legionella in a water network and the deposition of bacteria within the thoracic region (assessment of the number of viable Legionella, we validated a model mimicking realistic exposure through the use of (i recent technology for aerosol generation and (ii a 3D replicate of the human upper respiratory tract. The model's sensitivity was determined by monitoring the deposition of (i aerosolized water and Tc99m radio-aerosol as controls, and (ii bioaerosols generated from both Escherichia coli and Legionella pneumophila sg 1 suspensions. The numbers of viable Legionella prior to and after nebulization were provided by culture, flow cytometry and qPCR. This study was designed to obtain more realistic data on aerosol inhalation (vs. animal experimentation and deposition at the thoracic region in the context of LD. Upon nebulization, 40% and 48% of the initial Legionella inoculum was made of cultivable and non-cultivable cells, respectively; 0.7% of both populations reached the filter holder mimicking the thoracic region in this setup. These results are in agreement with experimental data based on quantitative microbial risk assessment methods and bring new methods that may be useful for preventing LD.

  19. On the use of the serial dilution culture method to enumerate viable phytoplankton in natural communities of plankton subjected to ballast water treatment

    OpenAIRE

    Cullen, John J.; MacIntyre, Hugh L.

    2015-01-01

    Discharge standards for ballast water treatment (BWT) systems are based on concentrations of living cells, for example, as determined with vital stains. Ultraviolet radiation (UV) stops the reproduction of microorganisms without killing them outright; they are living, but not viable, and ecologically as good as dead. Consequently, UV-treated discharge can be compliant with the intent of regulation while failing a live/dead test. An alternative evaluation of BWT can be proposed based on the as...

  20. Allogeneic cardiospheres delivered via percutaneous transendocardial injection increase viable myocardium, decrease scar size, and attenuate cardiac dilatation in porcine ischemic cardiomyopathy.

    Directory of Open Access Journals (Sweden)

    Kristine Yee

    Full Text Available Epicardial injection of heart-derived cell products is safe and effective post-myocardial infarction (MI, but clinically-translatable transendocardial injection has never been evaluated. We sought to assess the feasibility, safety and efficacy of percutaneous transendocardial injection of heart-derived cells in porcine chronic ischemic cardiomyopathy.We studied a total of 89 minipigs; 63 completed the specified protocols. After NOGA-guided transendocardial injection, we quantified engraftment of escalating doses of allogeneic cardiospheres or cardiosphere-derived cells in minipigs (n = 22 post-MI. Next, a dose-ranging, blinded, randomized, placebo-controlled ("dose optimization" study of transendocardial injection of the better-engrafting product was performed in infarcted minipigs (n = 16. Finally, the superior product and dose (150 million cardiospheres were tested in a blinded, randomized, placebo-controlled ("pivotal" study (n = 22. Contrast-enhanced cardiac MRI revealed that all cardiosphere doses preserved systolic function and attenuated remodeling. The maximum feasible dose (150 million cells was most effective in reducing scar size, increasing viable myocardium and improving ejection fraction. In the pivotal study, eight weeks post-injection, histopathology demonstrated no excess inflammation, and no myocyte hypertrophy, in treated minipigs versus controls. No alloreactive donor-specific antibodies developed over time. MRI showed reduced scar size, increased viable mass, and attenuation of cardiac dilatation with no effect on ejection fraction in the treated group compared to placebo.Dose-optimized injection of allogeneic cardiospheres is safe, decreases scar size, increases viable myocardium, and attenuates cardiac dilatation in porcine chronic ischemic cardiomyopathy. The decreases in scar size, mirrored by increases in viable myocardium, are consistent with therapeutic regeneration.

  1. Immunization of Rodents Against Hymenolepis Infections using Non-Viable Homologous Oncospheres

    Directory of Open Access Journals (Sweden)

    Ping-Chin Fan

    2004-12-01

    Full Text Available Immunity to Taiwan Taenia infection in pigs can be stimulated using homologous or heterologous nonviable Taenia oncospheres. This study was designed to determine whether homologous non-viable oncospheres could stimulate immunity to Hymenolepis infection in rodents. Hatched oncospheres were prepared from eggs of Hymenolepis diminuta, Hymenolepis nana, and Hymenolepis microstoma and kept at −70°C for more than 1 month. A mixture of 500 non-viable oncospheres of each tapeworm and complete Freund's adjuvant was injected subcutaneously in four groups of Sprague-Dawley rats or ICR mice one to four times at an interval of 1 week; controls were not immunized. After immunization, each rodent was orally inoculated with three fresh active cysticercoids of H. diminuta or H. microstoma or 500 fresh eggs of H. nana. The animals were then necropsied for adult tapeworms. No rats or mice immunized with non-viable oncospheres of H. diminuta or H. nana were infected by the challenge inoculation. However, 28 of 34 mice immunized with non-viable H. microstoma oncospheres were infected after inoculation with cysticercoids. This study demonstrated complete protection against infection by homologous parasites in rats or mice immunized with non-viable oncospheres of H. diminuta and H. nana, respectively. Repeated immunization may not be required if resistance is stimulated in rodent hosts.

  2. Airborne viable fungi in school environments in different climatic regions - A review

    Science.gov (United States)

    Salonen, Heidi; Duchaine, Caroline; Mazaheri, Mandana; Clifford, Sam; Lappalainen, Sanna; Reijula, Kari; Morawska, Lidia

    2015-03-01

    Elevated levels of fungi in indoor environments have been linked with mould/moisture damage in building structures. However, there is a lack of information about "normal" concentrations and flora as well as guidelines of viable fungi in the school environment in different climatic conditions. We have reviewed existing guidelines for indoor fungi and the current knowledge of the concentrations and flora of viable fungi in different climatic areas, the impact of the local factors on concentrations and flora of viable fungi in school environments. Meta-regression was performed to estimate the average behaviour for each analysis of interest, showing wide variation in the mean concentrations in outdoor and indoor school environments (range: 101-103 cfu/m3). These concentrations were significantly higher for both outdoors and indoors in the moderate than in the continental climatic area, showing that the climatic condition was a determinant for the concentrations of airborne viable fungi. The most common fungal species both in the moderate and continental area were Cladosporium spp. and Penicillium spp. The suggested few quantitative guidelines for indoor air viable fungi for school buildings are much lower than for residential areas. This review provides a synthesis, which can be used to guide the interpretation of the fungi measurements results and help to find indications of mould/moisture in school building structures.

  3. First study on the formation and resuscitation of viable but nonculturable state and beer spoilage capability of Lactobacillus lindneri.

    Science.gov (United States)

    Liu, Junyan; Li, Lin; Li, Bing; Peters, Brian M; Deng, Yang; Xu, Zhenbo; Shirtliff, Mark E

    2017-06-01

    This study aimed to investigate the spoilage capability of Lactobacillus lindneri during the induction and resuscitation of viable but nonculturable (VBNC) state. L. lindneri strain was identified by sequencing the PCR product (amplifying 16S rRNA gene) using ABI Prism 377 DNA Sequencer. During the VBNC state induction by low temperature storage and beer adaption, total, culturable, and viable cells were assessed by acridine orange direct counting, plate counting, and Live/Dead BacLight bacterial viability kit, respectively. Organic acids and diacetyl concentration were measured by reversed-phase high performance liquid chromatography and head dpace gas chromatography, respectively. VBNC state of L. lindneri was successfully induced by both beer adaption and low temperature storage, and glycerol frozen stock was the optimal way to maintain the VBNC state. Addition of catalase was found to be an effective method for the resuscitation of VBNC L. lindneri cells. Furthermore, spoilage capability remained similar during the induction and resuscitation of VBNC L. lindneri. This is the first report of induction by low temperature storage and resuscitation of VBNC L. lindneri strain, as well as the first identification of spoilage capability of VBNC and resuscitated L. lindneri cells. This study indicated that the potential colonization of L. lindneri strain in brewery environment, formation and resuscitation of VBNC state, as well as maintenance in beer spoilage capability, may be an important risk factor for brewery environment. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Yield Stress Effects on Mucus Plug Rupture

    Science.gov (United States)

    Hu, Yingying; Bian, Shiyao; Grotberg, John C.; Takayama, Shuichi; Grotberg, James B.

    2012-11-01

    Mucus plugs can obstruct airways, resulting in lost gas exchange and inflammation. Yield stress, one of the significant rheological properties of mucus, plays a significant role in plug rupture. We use carbopol 940 gels as mucus simulants to study dynamics of mucus plug rupture in experiments. Yield stress increases with gel concentration increasing (0.1% ~0.3%). The yield stress of the 0.2% gel is about 530 dyn/cm2, which can simulate normal mucus. A 2D PDMS channel is used to simulate a collapsed airway of the 12th generation in a human lung. Plug rupture is driven by a pressure drop of 1.6 ×104 ~ 2.0 ×104 dyn/cm2. Initial plug length varies from half to two times the half channel width. A micro-PIV technique is used to acquire velocity fields during rupture, from which wall shear stress is derived. Plug shortening velocity increases with the pressure drop, but decreases with yield stress or the initial plug length. Wall shear stress increases with yield stress, which indicates more potential damage may occur to epithelial cells when pathologic mucus has a high yield stress. Near the rupture moment, a wall shear stress peak appears at the front of the film deposited by the plug during rupture. This work is supported by NIH: HL84370 and HL85156.

  5. A Transient Cell-shielding Method for Viable MSC Delivery Within Hydrophobic Scaffolds Polymerized in situ

    Science.gov (United States)

    2015-03-27

    reporting burden for the collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching...existing data sources, gathering and maintaining the data needed, and completing and reviewing the collection of information. Send comments regarding this...Celzard A. Effect of composition and pro- cessing parameters on the characteristics of tannin -based rigid foams. Part II: physical properties. Mater

  6. A Novel Method for Determining the Level of Viable Disseminated Prostate Cancer Cells

    Science.gov (United States)

    2012-10-01

    Finally, in this final vector, we have introduced an integrin-targeting peptide ( RGD ) into the Fiber protein to broaden the tropism of the virus...3077. 22. Lupold SE, Kudrolli TA, Chowdhury WH, Wu P, Rodriguez R (2007) A novel method for generating and screening peptides and libraries displayed

  7. Cryopreserved ovarian cortex from patients with leukemia in complete remission contains no apparent viable malignant cells

    DEFF Research Database (Denmark)

    Greve, Tine; Clasen-Linde, Erik; Andersen, Morten T

    2012-01-01

    Some women suffering from leukemia require bone marrow transplantation to be cured. Bone marrow transplantation is associated with a high risk of sterility and some patients are offered fertility preservation by cryopreservation of ovarian cortex. Transplantation of ovarian cortex to women cured...

  8. A multicenter study of viable PCR using propidium monoazide to detect Legionella in water samples.

    Science.gov (United States)

    Scaturro, Maria; Fontana, Stefano; Dell'eva, Italo; Helfer, Fabrizia; Marchio, Michele; Stefanetti, Maria Vittoria; Cavallaro, Mario; Miglietta, Marilena; Montagna, Maria Teresa; De Giglio, Osvalda; Cuna, Teresa; Chetti, Leonarda; Sabattini, Maria Antonietta Bucci; Carlotti, Michela; Viggiani, Mariagabriella; Stenico, Alberta; Romanin, Elisa; Bonanni, Emma; Ottaviano, Claudio; Franzin, Laura; Avanzini, Claudio; Demarie, Valerio; Corbella, Marta; Cambieri, Patrizia; Marone, Piero; Rota, Maria Cristina; Bella, Antonino; Ricci, Maria Luisa

    2016-07-01

    Legionella quantification in environmental samples is overestimated by qPCR. Combination with a viable dye, such as Propidium monoazide (PMA), could make qPCR (named then vPCR) very reliable. In this multicentre study 717 artificial water samples, spiked with fixed concentrations of Legionella and interfering bacterial flora, were analysed by qPCR, vPCR and culture and data were compared by statistical analysis. A heat-treatment at 55 °C for 10 minutes was also performed to obtain viable and not-viable bacteria. When data of vPCR were compared with those of culture and qPCR, statistical analysis showed significant differences (P 0.05). Overall this study provided a good experimental reproducibility of vPCR but also highlighted limits of PMA in the discriminating capability of dead and live bacteria, making vPCR not completely reliable. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Issues of organizational cybernetics and viability beyond Beer's viable systems model

    Science.gov (United States)

    Nechansky, Helmut

    2013-11-01

    The paper starts summarizing the claims of Beer's viable systems model to identify five issues any viable organizations has to deal with in an unequivocal hierarchical structure of five interrelated systems. Then the evidence is introduced for additional issues and related viable structures of organizations, which deviate from Beer's model. These issues are: (1) the establishment and (2) evolution of an organization; (3) systems for independent top-down control (like "Six Sigma"); (4) systems for independent bottom-up correction of performance problems (like "Kaizen"), both working outside a hierarchical structure; (5) pull production systems ("Just in Time") and (6) systems for checks and balances of top-level power (like boards and shareholder meetings). Based on that an evolutionary approach to organizational cybernetics is outlined, addressing the establishment of organizations and possible courses of developments, including recent developments in quality and production engineering, as well as problems of setting and changing goal values determining organizational policies.

  10. Sorting of cells of the same size, shape, and cell cycle stage for a single cell level assay without staining

    Directory of Open Access Journals (Sweden)

    Yomo Tetsuya

    2006-06-01

    cell viability but decreases the proliferative ability of the PC12 cells. Conclusion We demonstrated a pretreatment method to collect well-characterized, viable, single cells without using fluorescent labels and without significant damage to the cells. This method is quantitative, rapid, single-step, and yields cells of high purity, making it applicable for a variety of single-cell level analyses.

  11. El modelo de sistema viable: un instrumento para la organización efectiva

    Directory of Open Access Journals (Sweden)

    Norlando Sánchez Rueda

    2015-05-01

    Full Text Available RESUMEN En este ensayo se presenta una interpretación teórica del denominado Modelo de Sistema Viable (MSV, de Stafford Beer y su Potencial Aplicación en Tareas de Diagnóstico  y diseño empresarial, al igual que para Mejorar las capacidades Organizacionales de Auto- Regulación  y Auto- Organización. Se explica como el Modelo del Sistema Viable permite conocer e interpretar  los mecanismos de estabilidad y adaptabilidad de las organizaciones, pilares para el crecimiento de una verdadera organización Efectiva.

  12. The amount of viable and dyssynchronous myocardium is associated with response to cardiac resynchronization therapy: initial clinical results using multiparametric ECG-gated [18F]FDG PET.

    Science.gov (United States)

    Lehner, Sebastian; Uebleis, Christopher; Schüßler, Franziska; Haug, Alexander; Kääb, Stefan; Bartenstein, Peter; Van Kriekinge, Serge D; Germano, Guido; Estner, Heidi; Hacker, Marcus

    2013-12-01

    There is still a significant amount of patients who do not sufficiently respond to cardiac resynchronization therapy (CRT). Previous studies demonstrated that the amount of dyssynchronous myocardium was predictive of response to CRT. Otherwise, non-response is frequently associated with high amounts of scar tissue. The combination of these parameters might yield a more accurate prediction of response. We hypothesized that the probability of a CRT response increases with the presence of high amounts of "viable and dyssynchronous" myocardium. A total of 19 patients (17 male, 61 ± 10 years) underwent ECG-gated [(18)F]fluorodeoxyglucose (FDG) myocardial positron emission tomography (PET) before CRT device implantation and were followed for 6 months. Response to CRT was defined as clinical improvement of at least one New York Heart Association (NYHA) class in combination with left ventricular (LV) ejection fraction (EF) improvement of >5%. Twelve responders (71%) and seven non-responders (29%) were identified. For each patient bullseye maps of FDG uptake and phase analysis were calculated (QPS/QGS 2012, Cedars-Sinai, Los Angeles, CA, USA) and fused. Amounts of myocardium representing "viable and synchronous", "scar and synchronous", viable and dyssynchronous or "scar and dyssynchronous" myocardium were quantified by planimetric measurements of the fused bullseye maps. Responders by definition showed significant decrease in NYHA class and significant increase of LVEF. Furthermore, a significantly higher amount of viable and dyssynchronous myocardium was found as compared to non-responders (21 ± 13% vs 6 ± 5%; p < 0.05). Combined assessment of myocardial viability and LV dyssynchrony is feasible using multiparametric [(18)F]FDG PET and could improve conventional response prediction criteria for CRT.

  13. DNA shuffling of adeno-associated virus yields functionally diverse viral progeny.

    Science.gov (United States)

    Koerber, James T; Jang, Jae-Hyung; Schaffer, David V

    2008-10-01

    Adeno-associated virus (AAV) vectors are extremely effective gene-delivery vehicles for a broad range of applications. However, the therapeutic efficacy of these and other vectors is currently limited by barriers to safe, efficient gene delivery, including pre-existing antiviral immunity, and infection of off-target cells. Recently, we have implemented directed evolution of AAV, involving the generation of randomly mutagenized viral libraries based on serotype 2 and high-throughput selection, to engineer enhanced viral vectors. Here, we significantly extend this capability by performing high-efficiency in vitro recombination to create a large (10(7)), diverse library of random chimeras of numerous parent AAV serotypes (AAV1, 2, 4-6, 8, and 9). In order to analyze the extent to which such highly chimeric viruses can be viable, we selected the library for efficient viral packaging and infection, and successfully recovered numerous novel chimeras. These new viruses exhibited a broad range of cell tropism both in vitro and in vivo and enhanced resistance to human intravenous immunoglobulin (IVIG), highlighting numerous functional differences between these chimeras and their parent serotypes. Thus, directed evolution can potentially yield unlimited numbers of new AAV variants with novel gene-delivery properties, and subsequent analysis of these variants can further extend basic knowledge of AAV biology.

  14. Optimization of Culture Medium Enhances Viable Biomass Production and Biocontrol Efficacy of the Antagonistic Yeast, Candida diversa

    Directory of Open Access Journals (Sweden)

    Jia Liu

    2017-10-01

    Full Text Available Viable biomass production is a key determinant of suitability of antagonistic yeasts as potential biocontrol agents. This study investigated the effects of three metal ions (magnesium, ferrous, and zinc on biomass production and viability of the antagonistic yeast, Candida diversa. Using response surface methodology to optimize medium components, a maximum biomass was obtained, when the collective Mg2+, Fe2+, and Zn2+ concentrations were adjusted in a minimal mineral (MM medium. Compared with the unmodified MM, and three ion-deficient MM media, yeast cells cultured in the three ion-modified MM medium exhibited a lower level of cellular oxidative damage, and a higher level of antioxidant enzyme activity. A biocontrol assay indicated that C. diversa grown in the ion-modified MM exhibited the greatest level of control of gray mold on apple fruit. These results provide new information on culture medium optimization to grow yeast antagonists in order to improve biomass production and biocontrol efficacy.

  15. Optimization of Culture Medium Enhances Viable Biomass Production and Biocontrol Efficacy of the Antagonistic Yeast, Candida diversa.

    Science.gov (United States)

    Liu, Jia; Li, Guangkun; Sui, Yuan

    2017-01-01

    Viable biomass production is a key determinant of suitability of antagonistic yeasts as potential biocontrol agents. This study investigated the effects of three metal ions (magnesium, ferrous, and zinc) on biomass production and viability of the antagonistic yeast, Candida diversa. Using response surface methodology to optimize medium components, a maximum biomass was obtained, when the collective Mg(2+), Fe(2+), and Zn(2+) concentrations were adjusted in a minimal mineral (MM) medium. Compared with the unmodified MM, and three ion-deficient MM media, yeast cells cultured in the three ion-modified MM medium exhibited a lower level of cellular oxidative damage, and a higher level of antioxidant enzyme activity. A biocontrol assay indicated that C. diversa grown in the ion-modified MM exhibited the greatest level of control of gray mold on apple fruit. These results provide new information on culture medium optimization to grow yeast antagonists in order to improve biomass production and biocontrol efficacy.

  16. On the use of the serial dilution culture method to enumerate viable phytoplankton in natural communities of plankton subjected to ballast water treatment.

    Science.gov (United States)

    Cullen, John J; MacIntyre, Hugh L

    2016-01-01

    Discharge standards for ballast water treatment (BWT) systems are based on concentrations of living cells, for example, as determined with vital stains. Ultraviolet radiation (UV) stops the reproduction of microorganisms without killing them outright; they are living, but not viable, and ecologically as good as dead. Consequently, UV-treated discharge can be compliant with the intent of regulation while failing a live/dead test. An alternative evaluation of BWT can be proposed based on the assessment of viable, rather than living, cells in discharge water. In principle, the serial dilution culture-most probable number (SDC-MPN) method provides the appropriate measure for phytoplankton. But, the method has been criticized, particularly because it is thought that many phytoplankton species cannot be cultured. A review of the literature shows that although SDC-MPN has been used for more than 50 years-generally to identify and count phytoplankton species that cannot be preserved-its application to enumerate total viable phytoplankton seems to be new, putting past criticisms of the method in a different light. Importantly, viable cells need to grow only enough to be detected, not to be brought into sustained culture, and competition between species in a dilution tube is irrelevant as long as the winner is detectable. Thorough consideration of sources of error leads to recommendations for minimizing and quantifying uncertainties by optimizing growth conditions and conducting systematic comparisons. We conclude that with careful evaluation, SDC-MPN is potentially an effective method for assessing the viability of phytoplankton after BWT.

  17. Rapid quantification of viable Campylobacter bacteria on chicken carcasses, using real-time PCR and propidium monoazide treatment, as a tool for quantitative risk assessment.

    Science.gov (United States)

    Josefsen, M H; Löfström, C; Hansen, T B; Christensen, L S; Olsen, J E; Hoorfar, J

    2010-08-01

    A number of intervention strategies against Campylobacter-contaminated poultry focus on postslaughter reduction of the number of cells, emphasizing the need for rapid and reliable quantitative detection of only viable Campylobacter bacteria. We present a new and rapid quantitative approach to the enumeration of food-borne Campylobacter bacteria that combines real-time quantitative PCR (Q-PCR) with simple propidium monoazide (PMA) sample treatment. In less than 3 h, this method generates a signal from only viable and viable but nonculturable (VBNC) Campylobacter bacteria with an intact membrane. The method's performance was evaluated by assessing the contributions to variability by individual chicken carcass rinse matrices, species of Campylobacter, and differences in efficiency of DNA extraction with differing cell inputs. The method was compared with culture-based enumeration on 50 naturally infected chickens. The cell contents correlated with cycle threshold (C(T)) values (R(2) = 0.993), with a quantification range of 1 x 10(2) to 1 x 10(7) CFU/ml. The correlation between the Campylobacter counts obtained by PMA-PCR and culture on naturally contaminated chickens was high (R(2) = 0.844). The amplification efficiency of the Q-PCR method was not affected by the chicken rinse matrix or by the species of Campylobacter. No Q-PCR signals were obtained from artificially inoculated chicken rinse when PMA sample treatment was applied. In conclusion, this study presents a rapid tool for producing reliable quantitative data on viable Campylobacter bacteria in chicken carcass rinse. The proposed method does not detect DNA from dead Campylobacter bacteria but recognizes the infectious potential of the VBNC state and is thereby able to assess the effect of control strategies and provide trustworthy data for risk assessment.

  18. Seed priming and sulfur effects on soybean cell membrane stability and yield in saline soil Condicionamento osmótico das sementes e disponibilidade de enxofre na estabilidade da membrana celular e produtividade de soja em solo salino

    Directory of Open Access Journals (Sweden)

    Teymur Khandan Bejandi

    2009-09-01

    Full Text Available The objective of this work was to determine the effects of seed priming and sulfur application on cell membrane characteristics, seedling emergence, chlorophyll content and grain yield of soybean (Glycine max in saline soil. A complete-block design in 4x3 factorial arrangement with three replicates was used to test four types of seed priming (water, auxin, gibberellin and non-priming and three levels of sulfate availability (0, 70 and 140 kg ha-1 K2SO4. The soil had a silty loam texture with an electrical conductivity of 3.61 ds m-1, a pH of 8.2 and a saturation percentage of about 46%. Seed priming had significant effects on mean emergence rate (MER, emergence percentage, relative water content (RWC of leaves, relative chlorophyll content, time of maturity, shoot length and grain yield. The highest values for these variables were observed in the priming treatments, except for the time of maturity. Sulfur application had significant effects on MER, shoot length, RWC, membrane injury index and grain yield. Priming treatments provide greater emergence rates and grain yields and interact sinergicaly with sulfur rates.O objetivo deste trabalho foi determinar os efeitos do condicionamento osmótico das sementes e da disponibilidade de enxofre sobre características da membrana celular, emergência de plântulas, conteúdo relativo de clorofila e produtividade de soja (Glycine max em solo salino. O experimento foi conduzido em delineamento de blocos ao acaso, em arranjo fatorial 4x3, com três repetições, para avaliar quatro tipos de condicionamento osmótico das sementes (água, auxina, giberelina e ausência de condicionamento e três níveis de disponibilidade de enxofre para as plantas (aplicação ao solo de 0, 70 e 140 kg ha-1 de K2SO4. O solo utilizado apresentava textura média, com condutividade elétrica de aproximadamente 3,61 ds m-1, pH de 8,2 e percentagem de saturação em torno de 46%. O condicionamento osmótico das sementes teve

  19. Induction of viable 2n pollen in sterile Oriental × Trumpet Lilium hybrids

    NARCIS (Netherlands)

    Luo, J.R.; Arens, P.; Niu, L.X.; Tuyl, van J.M.

    2016-01-01

    In order to induce viable 2n pollen from highly sterile diploid Oriental × Trumpet (OT) (Lilium), N2O was used to treat flower buds of four sterile diploid OT cultivars (‘Nymph’, ‘Gluhwein’, ‘Yelloween’, and ‘Shocking’) at different stages of meiosis. There was no pollen germination in

  20. 9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Detection of viable bacteria and fungi except in live vaccine. 113.26 Section 113.26 Animals and Animal Products ANIMAL AND PLANT HEALTH... in live vaccine. Each serial and subserial of biological product except live vaccines shall be tested...

  1. Oligonucleotide microarrays for the detection and identification of viable beer spoilage bacteria.

    Science.gov (United States)

    Weber, D G; Sahm, K; Polen, T; Wendisch, V F; Antranikian, G

    2008-10-01

    The design and evaluation of an oligonucleotide microarray in order to detect and identify viable bacterial species that play a significant role in beer spoilage. These belong to the species of the genera Lactobacillus, Megasphaera, Pediococcus and Pectinatus. Oligonucleotide probes specific to beer spoilage bacteria were designed. In order to detect viable bacteria, the probes were designed to target the intergenic spacer regions (ISR) between 16S and 23S rRNA. Prior to hybridization the ISR were amplified by combining reverse transcriptase and polymerase chain reactions using a designed consenus primer. The developed oligonucleotide microarrays allows the detection of viable beer spoilage bacteria. This method allows the detection and discrimination of single bacterial species in a sample containing complex microbial community. Furthermore, microarrays using oligonucleotide probes targeting the ISR allow the distinction between viable bacteria with the potential to grow and non growing bacteria. The results demonstrate the feasibility of oligonucleotide microarrays as a contamination control in food industry for the detection and identification of spoilage micro-organisms within a mixed population.

  2. Viable Techniques, Leontief’s Closed Model, and Sraffa’s Subsistence Economies

    Directory of Open Access Journals (Sweden)

    Alberto Benítez

    2014-11-01

    Full Text Available This paper studies the production techniques employed in economies that reproduce themselves. Special attention is paid to the distinction usually made between those that do not produce a surplus and those that do, which are referred to as first and second class economies, respectively. Based on this, we present a new definition of viable economies and show that every viable economy of the second class can be represented as a viable economy of the first class under two different forms, Leontief‘s closed model and Sraffa’s subsistence economies. This allows us to present some remarks concerning the economic interpretation of the two models. On the one hand, we argue that the participation of each good in the production of every good can be considered as a normal characteristic of the first model and, on the other hand, we provide a justification for the same condition to be considered a characteristic of the second model. Furthermore, we discuss three definitions of viable techniques advanced by other authors and show that they differ from ours because they admit economies that do not reproduce themselves completely.

  3. 9 CFR 113.27 - Detection of extraneous viable bacteria and fungi in live vaccines.

    Science.gov (United States)

    2010-01-01

    ... bacteria and fungi in live vaccines. 113.27 Section 113.27 Animals and Animal Products ANIMAL AND PLANT... bacteria and fungi in live vaccines. Unless otherwise specified by the Administrator or elsewhere exempted... Seed Bacteria shall be tested for extraneous viable bacteria and fungi as prescribed in this section. A...

  4. Food web (bio-)manipulation of South African reservoirs – viable ...

    African Journals Online (AJOL)

    2006-10-04

    Oct 4, 2006 ... Food web (bio-)manipulation of South African reservoirs – viable eutrophication ... Microcystis in local eutrophic waters is perceived as a primary major constraint in implementing 'classical' food-web manip- ulation. Intrinsic ... cially in 'shallow' lakes) to dismal failure (often in deep lakes) and a range of ...

  5. Contagem de células somáticas e produção de leite em vacas holandesas confinadas Somatic cells count and milk yield in confined holstein cows

    Directory of Open Access Journals (Sweden)

    Arlei Coldebella

    2004-06-01

    Full Text Available A relação entre contagem de células somáticas do leite (CCS e produção de leite é objeto de estudo de inúmeras pesquisas em diversos países, porém, no Brasil a literatura a esse respeito é escassa. Desse modo, o objetivo do presente trabalho é verificar se as perdas na produção de leite devidas ao aumento da CCS são proporcionais à produção, ou absolutas, independendo desta, e a partir de que valor de CCS elas começam a ser evidentes. Para o estudo foram utilizadas 13.725 observações, colhidas mensalmente de seis rebanhos, no período de janeiro de 2001 a junho de 2002. O modelo de análise, para curva de lactação, foi baseado na função gama incompleta, considerando os efeitos de ordem de lactação, rebanho e época do parto. A CCS foi incluída nesse modelo de duas formas: como fator multiplicativo (representando perdas relativas e como fator aditivo (representando perdas absolutas. A escolha do melhor modelo foi baseada no critério de informação de Schwarz (BIC. Concluiu-se que as perdas são absolutas, evidentes a partir de 17.000 células/mL e, para cada aumento de uma unidade na escala do logaritmo natural a partir desse valor, estimam-se perdas de 238 e 868 mL/dia para primíparas e multíparas, respectivamente.The relationship between milk somatic cells count (SCC and milk yield has been the aim of countless researches in several countries; however, in Brazil, the literature related to this subject is scarce. Therefore, the objective of this study is to verify if the losses in milk yield due to SCC increase are proportional to the yield, or absolute (independent of it and starting from which SCC, the losses become evident. For this study, 13.725 observations were used, collected monthly from 6 different herds, from January/2001 up to June/2002. The analysis model, for lactation curve, was based upon the incomplete gama function, considering the effects of lactation order, herd and calving season. The SCC

  6. How Close We Are to Achieving Commercially Viable Large-Scale Photobiological Hydrogen Production by Cyanobacteria: A Review of the Biological Aspects

    Science.gov (United States)

    Sakurai, Hidehiro; Masukawa, Hajime; Kitashima, Masaharu; Inoue, Kazuhito

    2015-01-01

    Photobiological production of H2 by cyanobacteria is considered to be an ideal source of renewable energy because the inputs, water and sunlight, are abundant. The products of photobiological systems are H2 and O2; the H2 can be used as the energy source of fuel cells, etc., which generate electricity at high efficiencies and minimal pollution, as the waste product is H2O. Overall, production of commercially viable algal fuels in any form, including biomass and biodiesel, is challenging, and the very few systems that are operational have yet to be evaluated. In this paper we will: briefly review some of the necessary conditions for economical production, summarize the reports of photobiological H2 production by cyanobacteria, present our schemes for future production, and discuss the necessity for further progress in the research needed to achieve commercially viable large-scale H2 production. PMID:25793279

  7. Systematics in delayed neutron yields

    Energy Technology Data Exchange (ETDEWEB)

    Ohsawa, Takaaki [Kinki Univ., Higashi-Osaka, Osaka (Japan). Atomic Energy Research Inst.

    1998-03-01

    An attempt was made to reproduce the systematic trend observed in the delayed neutron yields for actinides on the basis of the five-Gaussian representation of the fission yield together with available data sets for delayed neutron emission probability. It was found that systematic decrease in DNY for heavier actinides is mainly due to decrease of fission yields of precursors in the lighter side of the light fragment region. (author)

  8. Isolation of viable Toxoplasma gondii, molecular characterization, and seroprevalence in elk (Cervus canadensis) in Pennsylvania, USA.

    Science.gov (United States)

    Dubey, J P; Brown, J; Verma, S K; Cerqueira-Cézar, C K; Banfield, J; Kwok, O C H; Ying, Y; Murata, F H A; Pradhan, A K; Su, C

    2017-08-30

    Toxoplasmosis is a worldwide zoonosis. The ingestion of uncooked/undercooked meat and consumption of water contaminated with Toxoplasma gondii oocysts excreted by felids are the main modes of transmission of this parasite. T. gondii has been reported in multiple cervid species; however, little is known of the parasite in North American elk (Cervus canadensis). In the present study, antibodies to T. gondii were detected in serum of wild elk from Pennsylvania collected during 2013-2016 by the modified agglutination test (MAT, cut-off 1:25); 221 of 317 (69.7%) had MAT titers of 1:25 in 19, 1:50 in 28, 1:100 in 34, and 1:200 or higher in 140. Thus most (44.1%) elk had relatively high titers. Seroprevalence was slightly higher in males (76.9%) than females (67.5%, not statistically significant, Chi-square tests, P<0.0001) and was higher in adults (76.5%) than yearlings (46.4%, Odds ratio 3.82; 95% CL 1.72-8.47; P=0.001) or calves (21.7%, Odds ratio 12.58; 95% CL 4.51-35.10; P<0.0001). Annual seroprevalence was relatively stable throughout the period tested and ranged from 66.6% to 72.2%. Of the 101 elk harvested in 2016, hearts were bioassayed from 20 elk and tongues were bioassayed from 56; all tongue samples were negative. Viable T. gondii was isolated from hearts of two female elk, one of these was a seronegative adult and the other was a calf with no serum available for testing. Both T. gondii isolates were cultivated in cell culture and DNA derived from tachyzoites was characterized using the PCR-RFLP markers including SAG1, SAG2 (5'- 3'SAG2 and altSAG2), SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico. One isolate belongs to ToxoDB PCR-RFLP genotype #2 and the other is genotype #5. Both genotypes are frequently identified in animals in North America. Copyright © 2017. Published by Elsevier B.V.

  9. Predicting mask yields through the use of a yield model

    Science.gov (United States)

    Pond, Andrew D.

    1994-12-01

    Yield models have been successfully employed in wafer fabricators to provide data on yield learning, design for manufacturability, and the estimation of production cost for semiconductors. This paper describes how a yield model can be developed to provide important technical information for mask-making in the semiconductor industry. This information, focusing on manufacturing line loading strategy and customer charges, is different from that provided by wafer fabricators' yield models, however the underlying goal is the same: to estimate accurately the expected yield for a part produced on the manufacturing line. If this estimation is not done accurately, there can be serious cost and serviceability implications. The premise here is to categorize parts based on their expected yield (derived from the yield model) which is, itself, a function of how difficult they are to build. This model was developed using logistic regression analysis on historical data. Logistic regression has been used most commonly and successfully in epidemiological research where, for instance, the risk of an individual developing a certain type of cancer is modeled as a function of personal characteristics. methodological details of yield model development and performance monitoring are presented as well as a specific example.

  10. 7755 EFFECT OF NPK FERTILIZER ON FRUIT YIELD AND YIELD ...

    African Journals Online (AJOL)

    Win7Ent

    2013-06-03

    23] who reported maximum fruit yield in watermelon with an application of maleic hydrazide at 100 ppm. The yield increased from 7.22 tons/ha in the control to 13.1 tons/ha in the treatment with 100 ppm maleic hydrazide and ...

  11. Nitrogen rate and plant population effects on yield and yield ...

    African Journals Online (AJOL)

    This research was carried out to determine the effect of plant population and nitrogen rates on yield and yield components in soybean under Bursa, Mustafakemalpaa ecological conditions. Trials were conducted during 2000 and 2001 at the experimental fields of the Mustafakemalpaa Vocational High School, University of ...

  12. Relationship between yield and some yield components in cowpea ...

    African Journals Online (AJOL)

    Bean common mosaic virus–blackeye cowpea strain (BCMV-BlC) and cowpea aphid-borne mosaic virus (CABMV) belonging to the genus potyvirus are cosmopolitan and economically important pathogens of cowpea. This study analysed the linear relationship between yield and some yield related characters (numbers of ...

  13. Nitrogen rate and plant population effects on yield and yield ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-12-17

    Dec 17, 2008 ... This research was carried out to determine the effect of plant population and nitrogen rates on yield and yield components in soybean under Bursa, Mustafakemalpaşa ecological conditions. Trials were conducted during 2000 and 2001 at the experimental fields of the Mustafakemalpaşa Vocational High.

  14. Relationships between yield and some yield components in Pea ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-12-03

    Dec 3, 2008 ... components in Pea (Pisum sativum ssp arvense L.) genotypes by using correlation and ... end of the study, positive and significant relationship were found among seed yield and pods per plant and biological yield in both ... values of dry pea grains (%) are higher than green pea grains. Pea is a cool climate ...

  15. VARIABILITY OF YIELD AND YIELD COMPONENTS IN “EGUSI ...

    African Journals Online (AJOL)

    journal

    the population mean in seed yield plant-1. Lowest. Gs among all attributes was recorded in 100- seed weight in all crosses. DISCUSSION. High proportion of genetic variation (Table 1) implies that genetic variation plays an important role in the inheritance of yield attributes in. “egusi” melon. Raje and Rao (2000) noted that.

  16. Circular permutation prediction reveals a viable backbone disconnection for split proteins: an approach in identifying a new functional split intein.

    Directory of Open Access Journals (Sweden)

    Yun-Tzai Lee

    Full Text Available Split-protein systems have emerged as a powerful tool for detecting biomolecular interactions and reporting biological reactions. However, reliable methods for identifying viable split sites are still unavailable. In this study, we demonstrated the feasibility that valid circular permutation (CP sites in proteins have the potential to act as split sites and that CP prediction can be used to search for internal permissive sites for creating new split proteins. Using a protein ligase, intein, as a model, CP predictor facilitated the creation of circular permutants in which backbone opening imposes the least detrimental effects on intein folding. We screened a series of predicted intein CPs and identified stable and native-fold CPs. When the valid CP sites were introduced as split sites, there was a reduction in folding enthalpy caused by the new backbone opening; however, the coincident loss in entropy was sufficient to be compensated, yielding a favorable free energy for self-association. Since split intein is exploited in protein semi-synthesis, we tested the related protein trans-splicing (PTS activities of the corresponding split inteins. Notably, a novel functional split intein composed of the N-terminal 36 residues combined with the remaining C-terminal fragment was identified. Its PTS activity was shown to be better than current reported two-piece intein with a short N-terminal segment. Thus, the incorporation of in silico CP prediction facilitated the design of split intein as well as circular permutants.

  17. Comparison of spike and aerosol challenge tests for the recovery of viable influenza virus from non-woven fabrics.

    Science.gov (United States)

    Zuo, Zhili; de Abin, Martha; Chander, Yogesh; Kuehn, Thomas H; Goyal, Sagar M; Pui, David Y H

    2013-09-01

    To experimentally determine the survival kinetics of influenza virus on personal protective equipment (PPE) and to evaluate the risk of virus transfer from PPE, it is important to compare the effects on virus recovery of the method used to contaminate the PPE with virus and the type of eluent used to recover it. Avian influenza virus (AIV) was applied as a liquid suspension (spike test) and as an aerosol to three types of non-woven fabrics [polypropylene (PP), polyester (PET), and polyamide (Nylon)] that are commonly used in the manufacture of PPE. This was followed by virus recovery using eight different eluents (phosphate-buffered saline, minimum essential medium, and 1.5% or 3.0% beef extract at pH 7, 8, or 9). For spike tests, no statistically significant difference was found in virus recovery using any of the eluents tested. Hydrophobic surfaces (PP and PET) yielded higher spiked virus recovery than hydrophilic Nylon. From all materials, the virus recovery was much lower in aerosol challenge tests than in spike tests. Significant differences were found in the recovery of viable AIV from non-woven fabrics between spike and aerosol challenge tests. The findings of this study demonstrate the need for realistic aerosol challenge tests rather than liquid spike tests in studies of virus survival on surfaces where airborne transmission of influenza virus may get involved. © 2013 John Wiley & Sons Ltd.

  18. Yield gaps in oil palm

    NARCIS (Netherlands)

    Woittiez, Lotte S.; Wijk, van Mark T.; Slingerland, Maja; Noordwijk, van Meine; Giller, Ken E.

    2017-01-01

    Oil palm, currently the world's main vegetable oil crop, is characterised by a large productivity and a long life span (≥25 years). Peak oil yields of 12 t ha−1 yr−1 have been achieved in small plantations, and maximum theoretical yields as calculated with simulation models are 18.5 t oil ha−1 yr−1,

  19. Incorporating phenology into yield models

    Science.gov (United States)

    Gray, J. M.; Friedl, M. A.

    2015-12-01

    Because the yields of many crops are sensitive to meteorological forcing during specific growth stages, phenological information has potential utility in yield mapping and forecasting exercises. However, most attempts to explain the spatiotemporal variability in crop yields with weather data have relied on growth stage definitions that do not change from year-to-year, even though planting, maturity, and harvesting dates show significant interannual variability. We tested the hypothesis that quantifying temperature exposures over dynamically determined growth stages would better explain observed spatiotemporal variability in crop yields than statically defined time periods. Specifically, we used National Agricultural and Statistics Service (NASS) crop progress data to identify the timing of the start of the maize reproductive growth stage ("silking"), and examined the correlation between county-scale yield anomalies and temperature exposures during either the annual or long-term average silking period. Consistent with our hypothesis and physical understanding, yield anomalies were more correlated with temperature exposures during the actual, rather than the long-term average, silking period. Nevertheless, temperature exposures alone explained a relatively low proportion of the yield variability, indicating that other factors and/or time periods are also important. We next investigated the potential of using remotely sensed land surface phenology instead of NASS progress data to retrieve crop growth stages, but encountered challenges related to crop type mapping and subpixel crop heterogeneity. Here, we discuss the potential of overcoming these challenges and the general utility of remotely sensed land surface phenology in crop yield mapping.

  20. OF SEED YIELD IN SOYBEANS

    African Journals Online (AJOL)

    ( 1983) noted that tall plants often tended to produce high yield, while Gonzales et al. ( 1984) concluded that pod weight was the most appropriate character for indirect selection. The objective of this study was to determine the component characters whose selection would lead to improvement in seed yield of some Nigerian.

  1. Development of a specific immunomagnetic capture-PCR for rapid detection of viable Mycoplasma agalactiae in sheep milk samples.

    Science.gov (United States)

    Sanna, G; Lecca, V; Foddai, A; Tola, S

    2014-12-01

    To develop an immunomagnetic capture (IMC) to detect viable Mycoplasma agalactiae in routine ovine milk samples. Polyclonal antibodies against two M. agalactiae membrane surface proteins (P80 and P55) were covalently conjugated to magnetic beads (MBs) to form MB-Ab80 and MB-Ab55. Mycoplasma agalactiae cells were captured by a specific antigen-antibody reaction and magnetic separation. Immunomagnetic capture (IMC) was used to isolate and concentrate M. agalactiae in serial decimal dilutions and in artificially contaminated milk to facilitate subsequent detection by PCR. A 375-bp fragment of M. agalactiae was amplified using a pair of M. agalactiae-specific primers in PCR. The limit of detection of IMC-PCR method ranged from 10 to 10(2)  CCU ml(-1) when mycoplasmas were resuspended in PBS and from 10(2) to 10(3)  CCU ml(-1) when mycoplasmas were resuspended in uncontaminated ovine milk. This study also describes the application of IMC-PCR method to test for M. agalactiae in 516 milk samples collected from sheep with suspected contagious agalactia. Its performance was evaluated relative to culture. This report has demonstrated for the first time, the effective use of rapid and reliable IMC combined with PCR assay for the detection of viable M. agalactiae. The method IMC-PCR provides an alternative to conventional microbiological detection, method and it could be applied to quick detection of M. agalactiae in routine sheep milk samples. © 2014 The Authors published by John Wiley & Sons Ltd on behalf of Society for Applied Microbiology.

  2. Non-Breeding Eusocial Mole-Rats Produce Viable Sperm--Spermiogram and Functional Testicular Morphology of Fukomys anselli.

    Directory of Open Access Journals (Sweden)

    Angelica Garcia Montero

    Full Text Available Ansell's mole-rats (Fukomys anselli are subterranean rodents living in families composed of about 20 members with a single breeding pair and their non-breeding offspring. Most of them remain with their parents for their lifetime and help to maintain and defend the natal burrow system, forage, and care for younger siblings. Since incest avoidance is based on individual recognition (and not on social suppression we expect that non-breeders produce viable sperm spontaneously. We compared the sperm of breeding and non-breeding males, obtained by electroejaculation and found no significant differences in sperm parameters between both groups. Here, we used electroejaculation to obtain semen for the first time in a subterranean mammal. Spermiogram analysis revealed no significant differences in sperm parameters between breeders and non-breeders. We found significantly larger testes (measured on autopsies and on living animals per ultrasonography of breeders compared to non-breeders (with body mass having a significant effect. There were no marked histological differences between breeding and non-breeding males, and the relative area occupied by Leydig cells and seminiferous tubules on histological sections, respectively, was not significantly different between both groups. The seminiferous epithelium and to a lesser degree the interstitial testicular tissue are characterized by lesions (vacuolar degenerations, however, this feature does not hinder fertilization even in advanced stages of life. The continuous production of viable sperm also in sexually abstinent non-breeders might be best understood in light of the mating and social system of Fukomys anselli, and the potential to found a new family following an unpredictable and rare encounter with an unfamiliar female ("provoked or induced dispersal". Apparently, the non-breeders do not reproduce because they do not copulate but not because they would be physiologically infertile. The significantly

  3. Non-Breeding Eusocial Mole-Rats Produce Viable Sperm—Spermiogram and Functional Testicular Morphology of Fukomys anselli

    Science.gov (United States)

    Garcia Montero, Angelica; Vole, Christiane; Burda, Hynek; Malkemper, Erich Pascal; Holtze, Susanne; Morhart, Michaela; Saragusty, Joseph; Hildebrandt, Thomas B.; Begall, Sabine

    2016-01-01

    Ansell’s mole-rats (Fukomys anselli) are subterranean rodents living in families composed of about 20 members with a single breeding pair and their non-breeding offspring. Most of them remain with their parents for their lifetime and help to maintain and defend the natal burrow system, forage, and care for younger siblings. Since incest avoidance is based on individual recognition (and not on social suppression) we expect that non-breeders produce viable sperm spontaneously. We compared the sperm of breeding and non-breeding males, obtained by electroejaculation and found no significant differences in sperm parameters between both groups. Here, we used electroejaculation to obtain semen for the first time in a subterranean mammal. Spermiogram analysis revealed no significant differences in sperm parameters between breeders and non-breeders. We found significantly larger testes (measured on autopsies and on living animals per ultrasonography) of breeders compared to non-breeders (with body mass having a significant effect). There were no marked histological differences between breeding and non-breeding males, and the relative area occupied by Leydig cells and seminiferous tubules on histological sections, respectively, was not significantly different between both groups. The seminiferous epithelium and to a lesser degree the interstitial testicular tissue are characterized by lesions (vacuolar degenerations), however, this feature does not hinder fertilization even in advanced stages of life. The continuous production of viable sperm also in sexually abstinent non-breeders might be best understood in light of the mating and social system of Fukomys anselli, and the potential to found a new family following an unpredictable and rare encounter with an unfamiliar female (“provoked or induced dispersal”). Apparently, the non-breeders do not reproduce because they do not copulate but not because they would be physiologically infertile. The significantly increased

  4. Breeding for Grass Seed Yield

    DEFF Research Database (Denmark)

    Boelt, Birte; Studer, Bruno

    2010-01-01

    Seed yield is a trait of major interest for many fodder and amenity grass species and has received increasing attention since seed multiplication is economically relevant for novel grass cultivars to compete in the commercial market. Although seed yield is a complex trait and affected...... by agricultural practices as well as environmental factors, traits related to seed production reveal considerable genetic variation, prerequisite for improvement by direct or indirect selection. This chapter first reports on the biological and physiological basics of the grass reproduction system, then highlights...... important aspects and components affecting the seed yield potential and the agronomic and environmental aspects affecting the utilization and realization of the seed yield potential. Finally, it discusses the potential of plant breeding to sustainably improve total seed yield in fodder and amenity grasses....

  5. Closing Yield Gaps: How Sustainable Can We Be?

    Science.gov (United States)

    Pradhan, Prajal; Fischer, Günther; van Velthuizen, Harrij; Reusser, Dominik E; Kropp, Juergen P

    2015-01-01

    Global food production needs to be increased by 60-110% between 2005 and 2050 to meet growing food and feed demand. Intensification and/or expansion of agriculture are the two main options available to meet the growing crop demands. Land conversion to expand cultivated land increases GHG emissions and impacts biodiversity and ecosystem services. Closing yield gaps to attain potential yields may be a viable option to increase the global crop production. Traditional methods of agricultural intensification often have negative externalities. Therefore, there is a need to explore location-specific methods of sustainable agricultural intensification. We identified regions where the achievement of potential crop calorie production on currently cultivated land will meet the present and future food demand based on scenario analyses considering population growth and changes in dietary habits. By closing yield gaps in the current irrigated and rain-fed cultivated land, about 24% and 80% more crop calories can respectively be produced compared to 2000. Most countries will reach food self-sufficiency or improve their current food self-sufficiency levels if potential crop production levels are achieved. As a novel approach, we defined specific input and agricultural management strategies required to achieve the potential production by overcoming biophysical and socioeconomic constraints causing yield gaps. The management strategies include: fertilizers, pesticides, advanced soil management, land improvement, management strategies coping with weather induced yield variability, and improving market accessibility. Finally, we estimated the required fertilizers (N, P2O5, and K2O) to attain the potential yields. Globally, N-fertilizer application needs to increase by 45-73%, P2O5-fertilizer by 22-46%, and K2O-fertilizer by 2-3 times compared to the year 2010 to attain potential crop production. The sustainability of such agricultural intensification largely depends on the way

  6. Closing Yield Gaps: How Sustainable Can We Be?

    Directory of Open Access Journals (Sweden)

    Prajal Pradhan

    Full Text Available Global food production needs to be increased by 60-110% between 2005 and 2050 to meet growing food and feed demand. Intensification and/or expansion of agriculture are the two main options available to meet the growing crop demands. Land conversion to expand cultivated land increases GHG emissions and impacts biodiversity and ecosystem services. Closing yield gaps to attain potential yields may be a viable option to increase the global crop production. Traditional methods of agricultural intensification often have negative externalities. Therefore, there is a need to explore location-specific methods of sustainable agricultural intensification. We identified regions where the achievement of potential crop calorie production on currently cultivated land will meet the present and future food demand based on scenario analyses considering population growth and changes in dietary habits. By closing yield gaps in the current irrigated and rain-fed cultivated land, about 24% and 80% more crop calories can respectively be produced compared to 2000. Most countries will reach food self-sufficiency or improve their current food self-sufficiency levels if potential crop production levels are achieved. As a novel approach, we defined specific input and agricultural management strategies required to achieve the potential production by overcoming biophysical and socioeconomic constraints causing yield gaps. The management strategies include: fertilizers, pesticides, advanced soil management, land improvement, management strategies coping with weather induced yield variability, and improving market accessibility. Finally, we estimated the required fertilizers (N, P2O5, and K2O to attain the potential yields. Globally, N-fertilizer application needs to increase by 45-73%, P2O5-fertilizer by 22-46%, and K2O-fertilizer by 2-3 times compared to the year 2010 to attain potential crop production. The sustainability of such agricultural intensification largely depends

  7. Cytokinin: a key driver of seed yield.

    Science.gov (United States)

    Jameson, Paula Elizabeth; Song, Jiancheng

    2016-02-01

    The cytokinins have been implicated in many facets of plant growth and development including cell division and differentiation, shoot and root growth, apical dominance, senescence, fruit and seed development, and the response to biotic and abiotic stressors. Cytokinin levels are regulated by a balance between biosynthesis [isopentenyl transferase (IPT)], activation [Lonely Guy (LOG)], inactivation (O-glucosyl transferase), re-activation (β-glucosidase), and degradation [cytokinin oxidase/dehydrogenase (CKX)]. During senescence, the levels of active cytokinins decrease, with premature senescence leading to a decrease in yield. During the early stages of fruit and seed development, cytokinin levels are transiently elevated, and coincide with nuclear and cell divisions which are a determinant of final seed size. Exogenous application of cytokinin, ectopic expression of IPT, or down-regulation of CKX have, on occasions, led to increased seed yield, leading to the suggestion that cytokinin may be limiting yield. However, manipulation of cytokinins is complex, not only because of their pleiotropic nature but also because the genes coding for biosynthesis and metabolism belong to multigene families, the members of which are themselves spatially and temporally differentiated. Previous research on yield of rice showed that plant breeders could directly target the cytokinins. Modern genome editing tools could be employed to target and manipulate cytokinin levels to increase seed yield with the concurrent aim of maintaining quality. However, how the cytokinin level is modified and whether IPT or CKX is targeted may depend on whether the plant is considered to be in a source-limiting environment or to be sink limited. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  8. Effects of orally administered viable Lactobacillus rhamnosus GG and Propionibacterium freudenreichii subsp. shermanii JS on mouse lymphocyte proliferation.

    Science.gov (United States)

    Kirjavainen, P V; ElNezami, H S; Salminen, S J; Ahokas, J T; Wright, P F

    1999-11-01

    Immunomodulation by probiotics is a subject of growing interest, but the knowledge of dose response and time profile relationships is minimal. In this study we examined the effects of Lactobacillus rhamnosus GG (LGG) and Propionibacterium freudenreichii subsp. shermanii JS (PJS) on the proliferative activity of murine lymphocytes ex vivo. Dose dependency was assessed by treating animals perorally with a low or a high dose (i.e., 10(9) or 10(12) viable bacteria/kg of body weight) for 7 days. The lower dose levels of each strain appeared to enhance T-cell proliferation at the optimal concanavalin A (ConA) concentration (by 69 to 84%) and B-cell proliferation at the optimal and supraoptimal concentrations of lipopolysaccharide (by 57 to 82%). B-cell proliferation was also enhanced by the high LGG dose (by 32 to 39%) but was accompanied by a marginal decrease in T-cell proliferation (by 8%) at the optimal ConA concentration. The time profiles of the immune responses were assessed after daily treatment with the higher dose for 3, 7, and 14 days. A significant decrease in basal lymphoproliferation (by 32 to 42%) was observed with PJS treatment after the 3- and 7-day periods; however, this activity returned to control levels after 14 days of treatment, which also resulted in significantly enhanced T-cell proliferation at optimal and supraoptimal ConA concentrations (by 24 to 80%). The 14-day LGG treatment also enhanced the latter activity (by 119%). In conclusion, LGG and PJS have specific dose- and duration-dependent immunomodulatory effects on the proliferative activity of B and T lymphocytes and may also reduce lymphocyte sensitivity to the cytotoxic effects of lectin mitogens.

  9. Use of propidium monoazide and quantitative PCR for differentiation of viable Escherichia coli from E. coli killed by mild or pasteurizing heat treatments.

    Science.gov (United States)

    Yang, Xianqin; Badoni, Madhu; Gill, Colin O

    2011-12-01

    Suspensions of Escherichia coli in peptone water were heated at temperatures between 52 and 90 °C, inclusive. Samples withdrawn at suitable times were not or were treated with propidium monoazide (PMA) or deoxycholate then PMA before extraction of DNA. DNA was quantified by real-time PCR for estimation of the numbers of E. coli from which template DNA for the PCR was obtained. Numbers of viable E. coli in suspensions at the times of sampling were determined from plate counts. For samples from suspensions heated at temperatures ≥ 52 ≤ 72 °C, PCR cycle threshold (Ct) values were little or no different for DNA from corresponding samples that were or were not treated with PMA. PMA treatment of samples heated to ≥ 80 °C largely inactivated E. coli DNA for PCR. When samples heated to ≤ 72 °C were treated with deoxycholate before treatment with PMA, Ct values for treated samples were greater than the Ct values for the corresponding untreated samples. Similar results were obtained with E. coli suspended in milk or fluid from ground beef pummeled with diluent. The results indicate that cells killed by heating to ≥ 80 °C are permeable to PMA, but most cells killed by heating to ≤ 72 °C are not. However, treatment with deoxycholate renders a substantial fraction of the latter cells permeable to PMA. Numbers of viable or dead E. coli can then be estimated from Ct values for samples not treated or treated with deoxycholate and PMA, provided viable cells are ≥ 1% of the total. Copyright © 2011 Elsevier Ltd. All rights reserved.

  10. Isolation of viable Neospora caninum from brains of wild gray wolves (Canis lupus).

    Science.gov (United States)

    Dubey, J P; Jenkins, M C; Ferreira, L R; Choudhary, S; Verma, S K; Kwok, O C H; Fetterer, R; Butler, E; Carstensen, M

    2014-03-17

    Neospora caninum is a common cause of abortion in cattle worldwide. Canids, including the dog and the dingo (Canis familiaris), the coyote (Canis latrans), and the gray wolf (Canis lupus) are its definitive hosts that can excrete environmentally resistant oocysts in the environment, but also can act as intermediate hosts, harboring tissue stages of the parasite. In an attempt to isolate viable N. caninum from tissues of naturally infected wolves, brain and heart tissue from 109 wolves from Minnesota were bioassayed in mice. Viable N. caninum (NcWolfMn1, NcWolfMn2) was isolated from the brains of two wolves by bioassays in interferon gamma gene knockout mice. DNA obtained from culture-derived N. caninum tachyzoites of the two isolates were analyzed by N. caninum-specific Nc5 polymerase chain reaction and confirmed diagnosis. This is the first report of isolation of N. caninum from tissues of any wild canid host. Published by Elsevier B.V.

  11. Viable Intrauterine Pregnancy and Coexisting Molar Pregnancy in a Bicornuate Uterus: A Rare Presentation

    Directory of Open Access Journals (Sweden)

    Kavitha Krishnamoorthy

    2016-01-01

    Full Text Available A complete hydatidiform mole with a viable coexisting fetus (CMCF is a rare occurrence. Similarly, Mullerian anomalies such as a bicornuate uterus are uncommon variants of normal anatomy. We report a case of a 40-year-old female with a known bicornuate uterus presenting at 13 weeks gestation with vaginal bleeding. Ultrasound findings showed a healthy viable pregnancy in the right horn with complete molar pregnancy in the left horn. After extensive counseling, the patient desired conservative management, however, was unable to continue due to profuse vaginal bleeding. The patient underwent suction dilation and curettage under general anesthesia and evacuation of the uterine horns. Postoperatively, the patient was followed until serum beta-human chorionic gonadotropin (β-hCG level dropped to <5 mU. This is the first case of a CMCF reported in a bicornuate uterus, diagnosed with the use of ultrasound imaging.

  12. Effects of application boron on yields, yield component and oil ...

    African Journals Online (AJOL)

    The study was conducted to investigate the effects of five boron (B) doses; 0, 2.5, 5.0, 7.5 and 10.0 kg B ha-1 in B-deficient calcareous soils on yield and some yield components of four sunflower genotypes. Genotypes have shown variations with respect to their responses to B applications. AS-615 and Coban had the ...

  13. GENETIC ANALYSIS OF YIELD AND YIELD COMPONENTS IN ...

    African Journals Online (AJOL)

    ACSS

    2017-11-16

    Nov 16, 2017 ... yield and yield components and to estimate the heritabilities of important quantitative traits in rice (Oryza sativa. L.). Six generations viz., P1, P2, F1, F2, BCP1 and BCP2 of a cross between IET6279 and IR70445-146-3-3 were used for the study. Generation mean analysis suggested that additive effects had ...

  14. application of yield line the ion of yield line theory in pre ion of yield

    African Journals Online (AJOL)

    User

    Analysis of precast waffle slabs have ysis of precast waffle slabs have unanalyzed. T unanalyzed. This has led to cracks or his has led to cracks or. This paper proposed the use of. This paper proposed the use of yield line theory yield line theory develop a computer program called YLRGT a computer program called YLRGT.

  15. Improved identification of viable myocardium using second harmonic imaging during dobutamine stress echocardiography

    OpenAIRE

    Sozzi, Fabiola; Poldermans, Don; Bax, Jeroen; Elhendy, Abdou; Vourvouri, Eleni; Valkema, Roelf; Sutter, J.; Schinkel, Arend; Borghetti, A; Roelandt, Jos

    2001-01-01

    OBJECTIVE—To determine whether, compared with fundamental imaging, second harmonic imaging can improve the accuracy of dobutamine stress echocardiography for identifying viable myocardium, using nuclear imaging as a reference.
PATIENTS—30 patients with chronic left ventricular dysfunction (mean (SD) age, 60 (8) years; 22 men).
METHODS—Dobutamine stress echocardiography was carried out in all patients using both fundamental and second harmonic imaging. All patients underwent dual isotope simul...

  16. Decomposing global crop yield variability

    Science.gov (United States)

    Ben-Ari, Tamara; Makowski, David

    2014-11-01

    Recent food crises have highlighted the need to better understand the between-year variability of agricultural production. Although increasing future production seems necessary, the globalization of commodity markets suggests that the food system would also benefit from enhanced supplies stability through a reduction in the year-to-year variability. Here, we develop an analytical expression decomposing global crop yield interannual variability into three informative components that quantify how evenly are croplands distributed in the world, the proportion of cultivated areas allocated to regions of above or below average variability and the covariation between yields in distinct world regions. This decomposition is used to identify drivers of interannual yield variations for four major crops (i.e., maize, rice, soybean and wheat) over the period 1961-2012. We show that maize production is fairly spread but marked by one prominent region with high levels of crop yield interannual variability (which encompasses the North American corn belt in the USA, and Canada). In contrast, global rice yields have a small variability because, although spatially concentrated, much of the production is located in regions of below-average variability (i.e., South, Eastern and South Eastern Asia). Because of these contrasted land use allocations, an even cultivated land distribution across regions would reduce global maize yield variance, but increase the variance of global yield rice. Intermediate results are obtained for soybean and wheat for which croplands are mainly located in regions with close-to-average variability. At the scale of large world regions, we find that covariances of regional yields have a negligible contribution to global yield variance. The proposed decomposition could be applied at any spatial and time scales, including the yearly time step. By addressing global crop production stability (or lack thereof) our results contribute to the understanding of a key

  17. Cosmic constraint on massive neutrinos in viable f( R) gravity with producing Λ CDM background expansion

    Science.gov (United States)

    Lu, Jianbo; Liu, Molin; Wu, Yabo; Wang, Yan; Yang, Weiqiang

    2016-12-01

    Tensions between several cosmic observations were found recently, such as the inconsistent values of H0 (or σ 8) were indicated by the different cosmic observations. Introducing the massive neutrinos in Λ CDM could potentially solve the tensions. Viable f( R) gravity producing Λ CDM background expansion with massive neutrinos is investigated in this paper. We fit the current observational data: Planck-2015 CMB, RSD, BAO, and SNIa to constrain the mass of neutrinos in viable f( R) theory. The constraint results at 95% confidence level are: Σ m_ν case, m_{ν , sterile}^effcase. For the effects due to the mass of the neutrinos, the constraint results on model parameter at 95% confidence level become f_{R0}× 10^{-6}> -1.89 and f_{R0}× 10^{-6}> -2.02 for two cases, respectively. It is also shown that the fitting values of several parameters much depend on the neutrino properties, such as the cold dark matter density, the cosmological quantities at matter-radiation equality, the neutrino density and the fraction of baryonic mass in helium. Finally, the constraint result shows that the tension between direct and CMB measurements of H_0 gets slightly weaker in the viable f( R) model than that in the base Λ CDM model.

  18. Improved identification of viable myocardium using second harmonic imaging during dobutamine stress echocardiography.

    Science.gov (United States)

    Sozzi, F B; Poldermans, D; Bax, J J; Elhendy, A; Vourvouri, E C; Valkema, R; De Sutter, J; Schinkel, A F; Borghetti, A; Roelandt, J R

    2001-12-01

    To determine whether, compared with fundamental imaging, second harmonic imaging can improve the accuracy of dobutamine stress echocardiography for identifying viable myocardium, using nuclear imaging as a reference. 30 patients with chronic left ventricular dysfunction (mean (SD) age, 60 (8) years; 22 men). Dobutamine stress echocardiography was carried out in all patients using both fundamental and second harmonic imaging. All patients underwent dual isotope simultaneous acquisition single photon emission computed tomography (DISA-SPECT) with (99m)technetium-tetrofosmin/(18)F-fluorodeoxyglucose on a separate day. Myocardial viability was considered present by dobutamine stress echocardiography when segments with severe dysfunction showed a biphasic sustained improvement or an ischaemic response. Viability criteria on DISA-SPECT were normal or mildly reduced perfusion and metabolism, or perfusion/metabolism mismatch. Using fundamental imaging, 330 segments showed severe dysfunction at baseline; 144 (44%) were considered viable. The agreement between dobutamine stress echocardiography by fundamental imaging and DISA-SPECT was 78%, kappa = 0.56. Using second harmonic imaging, 288 segments showed severe dysfunction; 138 (48%) were viable. The agreement between dobutamine stress echocardiography and DISA-SPECT was significantly better when second harmonic imaging was used (89%, kappa = 0.77, p = 0.001 v fundamental imaging). Second harmonic imaging applied during dobutamine stress echocardiography increases the agreement with DISA-SPECT for detecting myocardial viability.

  19. Grapevine canopy reflectance and yield

    Science.gov (United States)

    Minden, K. A.; Philipson, W. R.

    1982-01-01

    Field spectroradiometric and airborne multispectral scanner data were applied in a study of Concord grapevines. Spectroradiometric measurements of 18 experimental vines were collected on three dates during one growing season. Spectral reflectance, determined at 30 intervals from 0.4 to 1.1 microns, was correlated with vine yield, pruning weight, clusters/vine, and nitrogen input. One date of airborne multispectral scanner data (11 channels) was collected over commercial vineyards, and the average radiance values for eight vineyard sections were correlated with the corresponding average yields. Although some correlations were significant, they were inadequate for developing a reliable yield prediction model.

  20. Contagem de células somáticas e produção de leite em vacas holandesas de alta produção Somatic cells count and milk yield in high production Holstein cows

    Directory of Open Access Journals (Sweden)

    Arlei Coldebella

    2003-12-01

    Full Text Available A mastite bovina pode ser clínica, com sinais visíveis, e subclínica, diagnosticada pela contagem das células somáticas. As perdas econômicas causadas pela mastite subclínica devem ser quantificadas para atender à demanda nacional de produtos lácteos. O objetivo deste trabalho foi verificar se as perdas na produção de leite, pelo aumento do número de células somáticas, são proporcionais ou independentes do nível de produção. Foram utilizadas 7.756 observações, colhidas mensalmente de um único rebanho, de setembro de 2000 a junho de 2002. A curva de lactação foi modelada pela função gama incompleta, e os efeitos de ordem de lactação, época do parto, ocorrência de doenças no periparto e escore de condição corporal ao parto também foram considerados. A contagem de células somáticas foi incluída nesse modelo como fator multiplicativo, representando perdas relativas, e como fator aditivo, representando perdas absolutas. A escolha do melhor modelo foi baseada no critério de informação de Schwarz (BIC. As perdas são absolutas, evidentes a partir de 14.270 células/mL e para cada aumento de uma unidade na escala do logaritmo natural a partir desse valor, estimam-se perdas de 184 e 869 g/dia para vacas primíparas e multíparas, respectivamente.The goal of this paper was to evaluate if losses in milk yield due to increase in somatic cells count are proportional or independent of the level of production. A total of 7,756 observations, monthly collected from a single herd from September/2000 up to June/2002, were used. The lactation curve was modeled by the incomplete gamma function, considering the effects of lactation order, calving season, peripartum disorder incidence and body condition score at calving. Somatic cells count was added to the model as a multiplicative factor, representing relative losses, and as an additive factor, representing absolute losses. The best model was chosen based on the information

  1. The dose-fractionation sensitivity of the kidney; assessment of viable tubule cross-sections at 19 months after X irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Jen, Y.-M.; Hendry, J.H. (Christie Hospital and Holt Radium Inst., Manchester (United Kingdom). Paterson Labs.)

    1993-03-01

    The formation of viable tubule cross-sections was assessed in histological sections of murine kidneys at 19 months after fractionated bilateral X-ray doses with 12 h intervals between fractions. The data were analysed using the linear-quadratic model which provides values of [alpha] and [beta] characterizing the slope of the dose-response curve, and the ratio of [alpha] and [beta] indicative of the sparing effect of dose fractionation. The tubule data were characterized by [alpha] = 0.057 [+-] 0.009 Gy[sup -1], [beta] = 0.011 [+-] 0.001 Gy[sup -2], [alpha]/[beta] = 5.0 [+-] 0.9 Gy. Also, the number of cells (per focus region of the nephron) calculated as being capable of producing a viable focus was 2.5 [+-] 0.5, which was confirmed using a separate two-dose approach (2.1 [+-] 0.3). Together with other data, of the order of 1000 regenerative cells per nephron (10[sup 4] total cells) can be deduced. The values of the fractionation sensitivity parameters are similar to values measured previously for cells taken from irradiated kidneys up to a year after irradiation and forming colonies in primary culture, and also similar to values assessed using various functional measures of kidney injury. (Author).

  2. Existence of both culturable and viable but non culturable (VNC) E. coli populations with distinct settling velocities in karst aquifer

    Science.gov (United States)

    Petit, F.; Ratajczak, M.; Massei, N.; Lafite, R.; Clermont, O.; Denamur, E.; Berthe, T.

    2012-12-01

    associated to "non settleable" particles, decreased from the sinkhole to the spring. In contrast in dry period while the turbidity and the contamination by culturable E. coli is low, the three populations of viable, culturable and VNC E. coli are introduced in the karst. Within the population of culturable E. coli, persistent strains mainly belonging to B1 phylogroup and growing at 7°C were introduced in karst aquifer. At the spring, whatever the class of particles up to 96% of viable cells were in VNC state suggesting that a physiological change occurred within aquifer, independently of the association of E. coli with particles. At the well, the pumping of the water induced an input of resident VNC E. coli consecutively to a resuspension of particles previously settled within the karstic network, during a past turbid event.

  3. Malignant small round cell tumors

    Science.gov (United States)

    Rajwanshi, Arvind; Srinivas, Radhika; Upasana, Gautam

    2009-01-01

    Malignant small round cell tumors are characterised by small, round, relatively undifferentiated cells. They generally include Ewing's sarcoma, peripheral neuroectodermal tumor, rhabdomyosarcoma, synovial sarcoma, non-Hodgkin's lymphoma, retinoblastoma, neuroblastoma, hepatoblastoma, and nephroblastoma or Wilms’ tumor. Other differential diagnoses of small round cell tumors include small cell osteogenic sarcoma, undifferentiated hepatoblastoma, granulocytic sarcoma, and intraabdominal desmoplastic small round cell tumor. Differential diagnosis of small round cell tumors is particularly difficult due to their undifferentiated or primitive character. Tumors that show good differentiation are generally easy to diagnose, but when a tumor is poorly differentiated, identification of the diagnostic, morphological features is difficult and therefore, no definitive diagnosis may be possible. As seen in several study reports, fine needle aspiration cytology (FNAC) has become an important modality of diagnosis for these tumors. The technique yields adequate numbers of dissociated, viable cells, making it ideally suitable for ancillary techniques. Typically, a multimodal approach is employed and the principal ancillary techniques that have been found to be useful in classification are immunohistochemistry and immunophenotyping by flow cytometry, reverse transcriptase polymerase chain reaction (RT-PCR), fluorescence in situ hybridization (FISH), and electron microscopy. However, the recent characterization of chromosomal breakpoints and the corresponding genes involved in malignant small round cell tumors means that it is possible to use molecular genetic approaches for detection. PMID:21938141

  4. Fission yield measurements at IGISOL

    Science.gov (United States)

    Lantz, M.; Al-Adili, A.; Gorelov, D.; Jokinen, A.; Kolhinen, V. S.; Mattera, A.; Moore, I.; Penttilä, H.; Pomp, S.; Prokofiev, A. V.; Rakopoulos, V.; Rinta-Antila, S.; Simutkin, V.; Solders, A.

    2016-06-01

    The fission product yields are an important characteristic of the fission process. In fundamental physics, knowledge of the yield distributions is needed to better understand the fission process. For nuclear energy applications good knowledge of neutroninduced fission-product yields is important for the safe and efficient operation of nuclear power plants. With the Ion Guide Isotope Separator On-Line (IGISOL) technique, products of nuclear reactions are stopped in a buffer gas and then extracted and separated by mass. Thanks to the high resolving power of the JYFLTRAP Penning trap, at University of Jyväskylä, fission products can be isobarically separated, making it possible to measure relative independent fission yields. In some cases it is even possible to resolve isomeric states from the ground state, permitting measurements of isomeric yield ratios. So far the reactions U(p,f) and Th(p,f) have been studied using the IGISOL-JYFLTRAP facility. Recently, a neutron converter target has been developed utilizing the Be(p,xn) reaction. We here present the IGISOL-technique for fission yield measurements and some of the results from the measurements on proton induced fission. We also present the development of the neutron converter target, the characterization of the neutron field and the first tests with neutron-induced fission.

  5. Fission yield measurements at IGISOL

    Directory of Open Access Journals (Sweden)

    Lantz M.

    2016-01-01

    Full Text Available The fission product yields are an important characteristic of the fission process. In fundamental physics, knowledge of the yield distributions is needed to better understand the fission process. For nuclear energy applications good knowledge of neutroninduced fission-product yields is important for the safe and efficient operation of nuclear power plants. With the Ion Guide Isotope Separator On-Line (IGISOL technique, products of nuclear reactions are stopped in a buffer gas and then extracted and separated by mass. Thanks to the high resolving power of the JYFLTRAP Penning trap, at University of Jyväskylä, fission products can be isobarically separated, making it possible to measure relative independent fission yields. In some cases it is even possible to resolve isomeric states from the ground state, permitting measurements of isomeric yield ratios. So far the reactions U(p,f and Th(p,f have been studied using the IGISOL-JYFLTRAP facility. Recently, a neutron converter target has been developed utilizing the Be(p,xn reaction. We here present the IGISOL-technique for fission yield measurements and some of the results from the measurements on proton induced fission. We also present the development of the neutron converter target, the characterization of the neutron field and the first tests with neutron-induced fission.

  6. Detection of viable enterotoxin-producing Bacillus cereus and analysis of toxigenicity from ready-to-eat foods and infant formula milk powder by multiplex PCR.

    Science.gov (United States)

    Zhang, Zhihong; Feng, Lixia; Xu, Hengyi; Liu, Chengwei; Shah, Nagendra P; Wei, Hua

    2016-02-01

    Bacillus cereus is responsible for several outbreaks of foodborne diseases due to its emetic toxin and enterotoxin. Enterotoxins, cytotoxin K (CytK), nonhemolytic enterotoxin (Nhe), and hemolysin BL (Hbl), have been recorded in several diarrheal cases due to food poisoning from B. cereus. The objective of this study was to develop a rapid and accurate method that combines multiplex PCR with propidium monoazide to selectively detect viable cells of enterotoxin-producing B. cereus in milk powder, noodles, and rice, and investigate the distribution of enterotoxins in 62 strains of B. cereus in Jiangxi province, China. The specificity of primers of 3 enterotoxins (i.e., cytK, nheA, and hblD) of B. cereus was verified by inclusivity and exclusivity tests using single PCR. Upon optimization of multiplex PCR conditions, it was found that the detection limit of viable cells was 10(2) cfu/mL of B. cereus in pure culture. By enrichment for 3 or 4 h and propidium monoazide pretreatment, a protocol for detection of viable cells as low as 2.2×10(1) cfu/g in spiked food (e.g., milk powder, noodles, and rice) was established and proved valid even under the interference of non-Bacillus cereus at as high as 10(5) cfu/g. Moreover, the protocol based on multiplex PCR for detection was applied for the analysis of distribution of toxin gene of B. cereus, and the results showed a regional feature for toxin gene distribution, indicating that potential toxigenicity of B. cereus should be evaluated further. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  7. Cell structure and percent viability by a slide centrifuge technique.

    Science.gov (United States)

    Fitzgerald, M G; Hosking, C S

    1982-01-01

    It was found that a slide centrifuge (Cytospin) preparation of a cell suspension allowed a reliable assessment of not only cell structure but also the percentage of non-viable cells. The non-viable cells appeared as "smear" cells and paralleled in number the cells taking up trypan blue. Direct experiment showed the unstained viable cells in a trypan blue cell suspension remained intact in a Cytospin preparation while the cells taking up trypan blue were the "smear" cells. The non-viability of the "smear" cells was confirmed by their inability to survive in culture. Images PMID:7040483

  8. GROWTH, GAS EXCHANGE AND YIELD OF CORN WHEN FERTIGATED WITH BOVINE BIOFERTILIZER

    OpenAIRE

    THALES VINÍCIUS DE ARAÚJO VIANA; JOÃO GUILHERME ARAÚJO LIMA; GEOCLEBER GOMES DE SOUSA; LUIS GONZAGA PINHEIRO NETO; BENITO MOREIRA DE AZEVEDO

    2014-01-01

    The bovine biofertilizer applied through irrigation water in the soil (bio fertigation), can be a viable organic source to maintain fertility levels in agricultural production systems. So, this work was aimed at evaluating the effects of different concentrations of bovine biofertilizer applied by fertigation on corn growth, gas exchange and yield. The experiment was conducted under full sun exposure, in Fortaleza, Ceara, in 100 liter (100 L) vessels. The experimental design was that of random...

  9. A PCR-Based Method for Monitoring Legionella pneumophila in Water Samples Detects Viable but Noncultivable Legionellae That Can Recover Their Cultivability▿

    Science.gov (United States)

    Dusserre, Eric; Ginevra, Christophe; Hallier-Soulier, Sylvie; Vandenesch, François; Festoc, Gabriel; Etienne, Jerome; Jarraud, Sophie; Molmeret, Maëlle

    2008-01-01

    Legionella pneumophila is the causative agent of Legionnaires' disease. This bacterium is ubiquitous in aqueous environments and uses amoebae as an intracellular replicative niche. Real-time PCR has been developed for rapid detection of Legionella DNA in water samples. In addition to culturable bacteria, this method may also detect dead and viable but noncultivable (VBNC) legionellae. In order to understand the significance of positive PCR results in this setting, we prepared water samples containing known concentrations of L. pneumophila and analyzed them comparatively by means of conventional culture, real-time PCR, viability labeling, and immunodetection (solid-phase cytometry). We also examined the influence of chlorination on the results of the four methods. The different techniques yielded similar results for nonchlorinated water samples but not for chlorinated samples. After treatment for 24 h with 0.5 and 1 ppm chlorine, all cultures were negative, PCR and immunodetection showed about 106 genome units and bacteria/ml, and total-viable-count (TVC) labeling detected 105 and 102 metabolically active bacteria/ml, respectively. Thus, PCR also detected bacteria that were VBNC. The recoverability of VBNC forms was confirmed by 5 days of coculture with Acanthamoeba polyphaga. Therefore, some TVC-positive bacteria were potentially infective. These data show that L. pneumophila PCR detects not only culturable bacteria but also VBNC forms and dead bacterial DNA at low chlorine concentrations. PMID:18515476

  10. Persistence and potential Viable but Non-culturable state of pathogenic bacteria during storage of digestates from agricultural biogas plants

    Directory of Open Access Journals (Sweden)

    Geraldine Maynaud

    2016-09-01

    Full Text Available Despite the development of on-farm anaerobic digestion as a process for making profitable use of animal by-products, factors leading to the inactivation of pathogenic bacteria during storage of digestates remain poorly described. Here, a microcosm approach was used to evaluate the persistence of three pathogenic bacteria (Salmonella enterica Derby, Campylobacter coli and Listeria monocytogenes in digestates from farms, stored for later land spreading. Nine samples, including raw digestates, liquid fractions of digestate and composted digestates, were inoculated with each pathogen and maintained for 40 days at 24°C. Concentrations of pathogens were monitored using culture and qPCR methods. The persistence of L. monocytogenes, detected up to 20 days after inoculation, was higher than that of Salmonella Derby, detected for 7-20 days, and of C. coli (not detected after 7 days. In some digestates, the concentration of the pathogens by qPCR assay was several orders of magnitude higher than the concentration of culturable cells, suggesting a potential loss of culturability and induction of Viable but Non-Culturable (VBNC state. The potential VBNC state which was generally not observed in the same digestate for the three pathogens, occurred more frequently for C. coli and L. monocytogenes than for Salmonella Derby. Composting a digestate reduced the persistence of seeded L. monocytogenes but promoted the maintenance of Salmonella Derby. The effect of NH4+/NH3 on the culturability of C. coli and Salmonella Derby was also shown.The loss of culturability may be the underlying mechanism for the regrowth of pathogens. We have also demonstrated the importance of using molecular tools to monitor pathogens in environmental samples since culture methods may underestimate cell concentration. Our results underline the importance of considering VBNC cells when evaluating the sanitary effect of an anaerobic digestion process and the persistence of pathogens during

  11. Isolation of dental pulp stem cells from a single donor and characterization of their ability to differentiate after 2 years of cryopreservation.

    Science.gov (United States)

    Alsulaimani, Reem S; Ajlan, Sumaiah A; Aldahmash, Abdullah M; Alnabaheen, May S; Ashri, Nahid Y

    2016-05-01

    To investigate the viability and differentiation capacity of dental pulp stem cells (DPSCs) isolated from single donors after 2 years of cryopreservation.   This prospective study was conducted between October 2010 and February 2014 in the Stem Unit, College of Medicine, King Saud University, Riyadh, Saudi Arabia. Seventeen teeth extracted from 11 participants were processed separately to assess the minimum tissue weight needed to yield cells for culturing in vitro. Cell stemness was evaluated before passage 4 using the colony forming unit assay, immunofluorescence staining, and bi-lineage differentiation. Dental pulp stem cells  were cryopreserved for 2 years. Post-thaw DPSCs were cultured until senescence and differentiated toward osteogenic, odontogenic, adipogenic, and chondrogenic lineages.   Viable cells were isolated successfully from 6 of the 11 participants. Three of these 6 cultured cell lines were identified as DPSCs. A minimum of 0.2 g of dental pulp tissue was required for successful isolation of viable cells from a single donor. Post-thaw  DPSCs successfully differentiated towards osteogenic, odontogenic, chondrogenic, and adipogenic lineages. The post-thaw DPSCs were viable in vitro up to 70 days before senescence. There was no significant difference between the cells.   Within the limitations of this investigation, viable cells from dental pulp tissue were isolated successfully from the same donor using a minimum of 2 extracted teeth. Not all isolated cells from harvested dental pulp tissue had the characteristics of DPSCs. Post-thaw DPSCs maintained their multi-lineage differentiation capacity.

  12. Rice yield prediction from yield components and limiting factors

    NARCIS (Netherlands)

    Casanova, D.; Goudriaan, J.; Catala Former, M.M.; Withagen, J.C.M.

    2002-01-01

    This article aims to quantify growth at field level in relation to crop status and soil properties in irrigated direct-seeded rice. Forty fields were selected in the Ebro Delta (Spain). Rice growth was monitored and soil properties measured. Yield was related to soil properties by a deductive

  13. Evaluation of Yield and Yield Attributes of Five Sweet Potato ...

    African Journals Online (AJOL)

    0087, and TIS 2532.OP.1.13) were evaluated for yield and agronomic performance in Imo State University Farm, Owerri. The experiment was laid out in a randomised complete block design with three replications. The planting density was 33,000 ...

  14. Yield and yield component association of some capsicum genotypes ...

    African Journals Online (AJOL)

    The result showed that all the genotypes of pepper used were significantly different (P = O.O 5) in plant height, number of branches, fruit length, fruit breadth, number of fruit per plant and total fruit yield but no significant differences was observed in days to 50% flowering and leaf area. The cultivar Tabasco pepper gave the ...

  15. Mapping quantitative trait loci associated with yield and yield ...

    Indian Academy of Sciences (India)

    Both parental lines contributed additively for QTLs identified for the yield components. A majority of the QTLs detected in our study are reported for the first time for reproductive stage salinity stress. Fine-mapping of selected putative QTLs will be the next step to facilitate marker-assisted backcrossing and to detect useful ...

  16. Effect of Integrated Nutrient Management on Yield and Yield ...

    African Journals Online (AJOL)

    ... of combined application of FYM (5 t ha-1) with 75% of recommended rates of inorganic NP in the study areas and other locations with similar agro-ecologies can significantly increase food barley yield and provide high economic return. Keywords: Farm Yard Manure; Food barley; Nitrogen; Phosphorous; Vermicompost ...

  17. yield and yield component association of some capsicum genotypes

    African Journals Online (AJOL)

    2015-07-24

    Jul 24, 2015 ... dried, ground to powder and use as ingredient in curry powder (Dewitt and Bosland 2009). World production of pepper is estimated at 2.4 million tons harvested from 1.65 million hectares giving an average yield of 1.4 tons/ha (FAO, 2005). In Nigeria, pepper is very important and widely cultivated in every.

  18. An Extension of the Hawkins and Simon Condition Characterizing Viable Techniques

    Directory of Open Access Journals (Sweden)

    Alberto Benítez Sánchez

    2015-01-01

    Full Text Available This paper discusses an extended version of the Hawkins and Simon condition which constitutes a synthetic formulation of the mathematical properties that viable economies must satisfy in single production models. The new version is implicit in the economic interpretations offered by them of the Hawkins and Simon condition, once a correction is introduced in one of those interpretations. Moreover, the paper details the meaning of the extended version following the interpretation of the original version proposed by Dorfman, Samuelson, and Solow. It also introduces a characteristic property of indecomposable matrices that has not previously been published.

  19. THE CIVILIZATIONAL PROJECT AND lTS DISCONTENTS: TOWARD A VIABLE GLOBAL MARKET SOCIETY?

    Directory of Open Access Journals (Sweden)

    Volker Bornschier

    2015-08-01

    Full Text Available I agree with the point of an anonymous reviewer arguing that a pro-found discussion on the prerequisites of a viable global market society has too many facets to be contained within the bounds of a single journal article. Yet, in order to enter the debate now we should not wait until book length treatments become available-often only after years. This is, therfore, an essay attempting to overview various conflicts and contradictions within the global social system. It sythesizes arguments developed in more detail else-where.

  20. Fake Journals: Their Features and Some Viable Ways to Distinguishing Them

    DEFF Research Database (Denmark)

    Hemmat Esfe, Mohammad; Wongwises, Somchai; Asadi, Amin

    2015-01-01

    In this paper, we aim to discuss the fake journals and their advertisement and publication techniques. These types of journals mostly start and continue their activities by using the name of some indexed journals and establishing fake websites. The fake journals and publishers, while asking...... the authors for a significant amount of money for publishing their papers, have no peer-review process, publish the papers without any revision on the fake sites, and put the scientific reputation and prestige of the researchers in jeopardy. In the rest of the paper, we present some viable techniques in order...

  1. Information Warfare: using the viable system model as a framework to attack organisations

    Directory of Open Access Journals (Sweden)

    Bill Hutchinson

    2002-05-01

    Full Text Available Information is the glue in any organization. It is needed for policy, decision-making, control, and co-ordination. If an organisation's information systems are disrupted or destroyed, then damage to the whole inevitably follows. This paper uses a proven systemic, analytic framework the Viable System Model (VSM - in a functionalist mode, to analyse the vulnerabilities of an organisation's information resources to this form of aggression. It examines the tactics available, and where they can be used to effectively attack an organisation.

  2. Current progress and challenges in engineering viable artificial leaf for solar water splitting

    Directory of Open Access Journals (Sweden)

    Phuc D. Nguyen

    2017-12-01

    Full Text Available Large scale production of H2, a clean fuel, can be realized with just water and solar light energy by employing a viable energy conversion device called artificial leaf. In this tutorial review, we discuss on advances achieved recently and technical challenges remained toward the creation of such a leaf. Development of key components like catalysts for water electrolysis process and light harvester for harvesting solar energy as well as strategies being developed for assembling these components to create a complete artificial leaf will be highlighted.

  3. Emergency total thyroidectomy for bleeding anaplastic thyroid carcinoma: A viable option for palliation

    Directory of Open Access Journals (Sweden)

    Sunil Kumar

    2011-01-01

    Full Text Available Anaplastic thyroid carcinoma (ATC is a rare and highly aggressive thyroid neoplasm. Bleeding from tumor is an uncommon, but potentially life-threatening complication requiring sophisticated intervention facilities which are not usually available at odd hours in emergency. We report the case of a 45-year-old woman who presented with exsanguinating hemorrhage from ATC and was treated by emergency total thyroidectomy. The patient is well three months postoperatively. Emergency total thyroidectomy is a viable option for palliation in ATC presenting with bleeding.

  4. Yield Improvement and Energy Savings Uing Phosphonates as Additives in Kraft pulping

    Energy Technology Data Exchange (ETDEWEB)

    Ulrike W. Tschirner; Timothy Smith

    2007-03-31

    Project Objective: Develop a commercially viable modification to the Kraft process resulting in energy savings, increased yield and improved bleachability. Evaluate the feasibility of this technology across a spectrum of wood species used in North America. Develop detailed fundamental understanding of the mechanism by which phosphonates improve KAPPA number and yield. Evaluate the North American market potential for the use of phosphonates in the Kraft pulping process. Examine determinants of customer perceived value and explore organizational and operational factors influencing attitudes and behaviors. Provide an economic feasibility assessment for the supply chain, both suppliers (chemical supply companies) and buyers (Kraft mills). Provide background to most effectively transfer this new technology to commercial mills.

  5. Texture analysis of cardiac cine magnetic resonance imaging to detect non-viable segments in patients with chronic myocardial infarction.

    Science.gov (United States)

    Larroza, Andrés; López-Lereu, María P; Monmeneu, José V; Gavara, Jose; Chorro, Francisco J; Bodí, Vicente; Moratal, David

    2018-02-01

    To investigate the ability of texture analysis to differentiate between infarcted non-viable, viable, and remote segments on cardiac cine magnetic resonance imaging (MRI). This retrospective study included 50 patients suffering chronic myocardial infarction. The data was randomly split into training (30 patients) and testing (20 patients) sets. The left ventricular myocardium was segmented according to the 17-segment model in both cine and late gadolinium enhancement (LGE) MRI. Infarcted myocardium regions were identified on LGE in short-axis views. Non-viable segments were identified as those showing LGE ≥ 50%, and viable segments those showing 0 cine images. A support vector machine (SVM) classifier was trained with different combination of texture features to obtain a model that provided optimal classification performance. The best classification on testing set was achieved with local binary patterns features using a 2D + t approach, in which the features are computed by including information of the time dimension available in cine sequences. The best overall area under the receiver operating characteristic curve (AUC) were: 0.849, sensitivity of 92% to detect non-viable segments, 72% to detect viable segments, and 85% to detect remote segments. Non-viable segments can be detected on cine MRI using texture analysis and this may be used as hypothesis for future research aiming to detect the infarcted myocardium by means of a gadolinium-free approach. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  6. Reduction of viable Enterococcus faecalis in human radicular dentin treated with 1% cetrimide and conventional intracanal medicaments.

    Science.gov (United States)

    Carbajal Mejía, Jeison B; Aguilar Arrieta, Angela

    2016-08-01

    The purpose of this study was to evaluate the viability of E. faecalis after a 14-day exposure to 1% cetrimide (CET), triantibiotic paste (i.e., metronidazole, minocycline, and ciprofloxacin), 2% chlorhexidine (CHX) gel, and calcium hydroxide (Ca[OH]2 ) in an infected dentine model. A total of 75 roots of extracted uniradicular human teeth were chemomechanically prepared, sterilized, and infected for 21 days with E. faecalis. Samples were divided into five groups (n = 15) to apply intracanal medicaments, namely saline solution (negative control), 1% CET, triantibiotic paste (TRIA), 2% CHX gel, and Ca(OH)2 during 14 days. Dentine samples were collected and stained with the SYTO 9/propidium iodide technique (Live/Dead, Bacligth, Invitrogen, Eugene, OR, USA) for fluorescence microscopy to obtain the percentage of viable cells. Statistical analysis was performed using one-way analysis of variance followed by Tukey's multiple comparison test (P faecalis when compared with the control group. In addition, the least cytotoxic medicament was Ca(OH)2 followed by CHX. There was no significant difference between 1% CET and TRIA (P = 0.98). Both 1% CET and TRIA significantly reduced the viability of E. faecalis in dentine of extracted teeth in comparison with 2% CHX gel and calcium hydroxide paste. Further laboratory and clinical investigations should be carried out to validate findings of the beneficial use of 1% CET as an intracanal medicament against E. faecalis. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. Isolation of Viable but Non-culturable Bacteria from Printing and Dyeing Wastewater Bioreactor Based on Resuscitation Promoting Factor.

    Science.gov (United States)

    Jin, Yi; Gan, Guojuan; Yu, Xiaoyun; Wu, Dongdong; Zhang, Li; Yang, Na; Hu, Jiadan; Liu, Zhiheng; Zhang, Lixin; Hong, Huachang; Yan, Xiaoqing; Liang, Yan; Ding, Linxian; Pan, Yonglong

    2017-07-01

    Printing and dyeing wastewater with high content of organic matters, high colority, and poor biochemical performance is hard to be degraded. In this study, we isolated viable but non-culturable (VBNC) bacteria from printing and dyeing wastewater with the culture media contained resuscitation promoting factor (Rpf) protein secreted by Micrococcus luteus, counted the culturable cells number with the most probable number, sequenced 16S rRNA genes, and performed polymerase chain reaction-denaturing gradient gel electrophoresis. It is obviously that the addition of Rpf in the enrichment culture could promote growth and resuscitation of bacteria in VBNC state to obtain more fastidious bacteria significantly. The identified bacteria were assigned to nine genera in the treatment group, while the two strains of Ochrobactrum anthropi and Microbacterium sp. could not be isolated from the control group. The function of isolated strains was explored and these strains could degrade the dye of Congo red. This study provides a new sight into the further study including the present state, composition, formation mechanism, and recovery mechanism about VBNC bacteria in printing and dyeing wastewater, which would promote to understand bacterial community in printing and dyeing wastewater, and to obtain VBNC bacteria from ecological environment.

  8. Are big potassium-type Ca(2+)-activated potassium channels a viable target for the treatment of epilepsy?

    Science.gov (United States)

    Leo, Antonio; Citraro, Rita; Constanti, Andrew; De Sarro, Giovambattista; Russo, Emilio

    2015-07-01

    BK (big potassium) channels are Ca(2+)-activated K(+) channels widely expressed in mammalian cells. They are extensively distributed in the CNS, the most abundant level being found in brain areas largely involved in epilepsy, namely cortex, hippocampus, piriform cortex, and other limbic structures. BK channels control action potential shape/duration, thereby regulating membrane excitability and Ca(2+) signaling. The potassium channel superfamily represents a rich source of potential targets for therapeutic intervention in epilepsy. Some studies have identified alterations in BK channel function, therefore, supporting the development of drugs acting on these channels for epilepsy treatment. The actual sketch is intriguing and controversial, since mechanisms altering the physiological role of BK channels leading to either a loss- or gain-of-function have both been linked to seizure onset. Not many studies have been performed to unravel the efficacy of drugs acting on these channels as potential antiepileptics; however, paradoxically, efficacy has been demonstrated for both BK channel openers and blockers. Furthermore, their potential usefulness in preventing epileptogenesis has not been investigated at all. Substantial data on risks and benefits of modulating these channels are urgently needed to draw a definitive conclusion on whether BK channels are a viable future target for the treatment of epilepsy.

  9. Structure based modification of Bluetongue virus helicase protein VP6 to produce a viable VP6-truncated BTV

    Energy Technology Data Exchange (ETDEWEB)

    Matsuo, Eiko [Microbiology and Immunology, Division of Animal Science, Department of Bioresource Science, Graduate School of Agricultural Science, Kobe University, 1-1, Rokkodai, Nada-ku, Kobe-City 657-8501 (Japan); Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT (United Kingdom); Leon, Esther; Matthews, Steve J. [Division of Molecular Biosciences, Centre for Structural Biology, Imperial College London, South Kensington, London SW7 2AZ (United Kingdom); Roy, Polly, E-mail: polly.roy@lshtm.ac.uk [Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT (United Kingdom)

    2014-09-05

    Highlights: • NMR analysis on BTV VP6 reveals two large loop regions. • The loss of a loop (aa 34–130) does not affect the overall fold of the protein. • A region of VP6 (aa 34–92) is not required for BTV replication. • A region of VP6 (aa 93–130) plays an essential role in the virus replication. - Abstract: Bluetongue virus core protein VP6 is an ATP hydrolysis dependent RNA helicase. However, despite much study, the precise role of VP6 within the viral capsid and its structure remain unclear. To investigate the requirement of VP6 in BTV replication, we initiated a structural and biological study. Multinuclear nuclear magnetic resonance spectra were assigned on his-tagged full-length VP6 (329 amino acid residues) as well as several truncated VP6 variants. The analysis revealed a large structured domain with two large loop regions that exhibit significant conformational exchange. One of the loops (amino acid position 34–130) could be removed without affecting the overall fold of the protein. Moreover, using a BTV reverse genetics system, it was possible to demonstrate that the VP6-truncated BTV was viable in BHK cells in the absence of any helper VP6 protein, suggesting that a large portion of this loop region is not absolutely required for BTV replication.

  10. Bacteriophage treatment significantly reduces viable Clostridium difficile and prevents toxin production in an in vitro model system.

    Science.gov (United States)

    Meader, Emma; Mayer, Melinda J; Gasson, Michael J; Steverding, Dietmar; Carding, Simon R; Narbad, Arjan

    2010-12-01

    Clostridium difficile is primarily a nosocomial pathogen, causing thousands of cases of antibiotic-associated diarrhoea in the UK each year. In this study, we used a batch fermentation model of a C. difficile colonised system to evaluate the potential of a prophylactic and a remedial bacteriophage treatment regime to control the pathogen. It is shown that the prophylaxis regime was effective at preventing the growth of C. difficile (p = <0.001) and precluded the production of detectable levels of toxins A and B. The remedial treatment regime caused a less profound and somewhat transient decrease in the number of viable C. difficile cells (p = <0.0001), but still resulted in a lower level of toxin production relative to the control. The numbers of commensal bacteria including total aerobes and anaerobes, Bifidobacterium sp., Bacteroides sp., Lactobacillus sp., total Clostridium sp., and Enterobacteriaceae were not significantly decreased by this therapy, whereas significant detrimental effects were observed with metronidazole treatment. Our study indicates that phage therapy has potential to be used for the control of C. difficile; it highlights the main benefits of this approach, and some future challenges. Copyright © 2010 Elsevier Ltd. All rights reserved.

  11. Isolation and Culture of Single Cell Types from Rat Liver.

    Science.gov (United States)

    Zhang, Qidi; Qu, Ying; Li, Zhenghong; Zhang, Qingqing; Xu, Mingyi; Cai, Xiaobo; Li, Fei; Lu, Lungen

    2016-01-01

    There have been few reports on the simultaneous isolation of multiple liver cell populations thus far. As such, this study was aimed at establishing a protocol for the simultaneous separation of hepatocytes (HCs), hepatic stellate cells (HSCs), liver sinusoidal endothelial cells (LSECs) and Kupffer cells (KCs) from the rat liver and assessing the in vitro culture of these cells. Single-cell suspensions from the liver were obtained by ethylene glycol tetraacetic acid/collagenase perfusion. After low-speed centrifugal separation of HCs, pronase was added to the nonparenchymal cell fraction to eliminate the remaining HCs. Subsequently, HSCs, LSECs and KCs were purified by two steps of density gradient centrifugation using Nycodenz and Percoll in addition to selective attachment. Pronase treatment increased the HSC yield (1.5 ± 0.2 vs. 0.7 ± 0.3 cells/g liver, p cultured in vitro. LSEC apoptosis began on day 3 and reached a maximum on day 7. A few surviving LSECs began proliferating and split to form a cobblestone, sheet-like appearance on day 14. The LSECs on day 14 lost fenestrations but retained scavenger function. Thus, viable and purified liver cells were obtained with a high yield from the rat liver using the developed method, which may be useful for studying the physiology and pathology of the liver in the future. © 2016 S. Karger AG, Basel.

  12. Cultivar mixtures: a meta-analysis of the effect of intraspecific diversity on crop yield.

    Science.gov (United States)

    Reiss, Emily R; Drinkwater, Laurie E

    2017-09-22

    Extensive research has shown that greater plant community diversity leads to higher levels of productivity and other ecosystem services, and such increased diversity has been suggested as a way to improve yield and agricultural sustainability. Increasing intraspecific diversity with cultivar mixtures is one way to increase diversity in agricultural systems. We examined the relationship between intraspecific diversity and yield in cultivar mixtures using a meta-analysis of 91 studies and >3,600 observations. Additionally, we investigated how environmental and management factors might influence this relationship, and if the yield stability of cultivar mixtures differed from that of monocultures. We found that the yield increased by 2.2% overall in cultivar mixtures relative to their monoculture components. Mixtures with more cultivars and those with more functional trait diversity showed higher relative yields. Under biotic stressors, such as disease pressure, and abiotic stressors, such as low levels of soil organic matter and nutrient availability, this diversity effect was stronger, resulting in higher relative yields. Finally, cultivar mixtures generally showed higher yield stability compared to monocultures, especially in response to annual weather variability at a site over time. This practice of mixing cultivars can be integrated into intensified cropping systems where species monocultures dominate, as well as in smallholder cropping systems where low-cost improvements are in demand. Overall, these results suggest that cultivar mixtures are a viable strategy to increase diversity in agroecosystems, promoting increased yield and yield stability, with minimal environmental impact. © 2017 by the Ecological Society of America.

  13. Efeito da densidade de mudas por célula e do volume da célula na produção de mudas e cultivo da rúcula Effect of seedlings density per cells and cells volume on seedlings production and yield of rocket press

    Directory of Open Access Journals (Sweden)

    Marie Yamamoto Reghin

    2004-04-01

    úcula. Houve diminuição no comprimento da parte aérea, número de folhas, peso da matéria fresca e seca produzida por planta, em resposta à pressão de competição promovida pelo aumento da densidade. Considerando o rendimento de matéria fresca, a densidade contribuiu diretamente para o incremento dessa característica. Observou-se a média de 1,4 kg m-2, quando se usou a densidade de uma muda e de 6,4 kg m-2 com quatro mudas. Para os dois tipos de bandejas, à medida que houve aumento da densidade de mudas, houve resposta linear crescente na matéria fresca por área. Como a forma de comercialização predominante para a rúcula é em maços, o rendimento promovido pela densidade de quatro mudas, tanto na bandeja de 200 como na de 288 células, apresentou-se como a alternativa mais adequada.The experiment was realized in Ponta Grossa (PR aiming at avaluating seedlings density/cells using different trays and its effects on seedlings production and yield of rocket press. The experimental design was a randomised blocks with four replications; the treatments followed a factorial arrangement 4 x 2, being 1, 2 , 3 and 4 seedlings/cell on trays with 200 and 288 cells. Seeds were sowed on 28/03/2003, with the cultivar Cultivada on trays with commercial substrate Plantmax under protected cultivation. Before seedlings transplanting it was avaluated the characteristics of leaves number, lenght of upper part and root, fresh and dry weight of up ground material and root. The seedlings transplanting was carried out through with seventeen days in plots containing 40 plants in 0,20 x 0,10 m arrangement. At harvesting, 20 days of the seedlings transplanting, it was avaluated the following characteristics: lenght of upper part, leaves number, fresh and dry weight of up ground material. On the seedlings stage it was observed a significative effect of the interaction. The cell size was an important factor on seedlings production and it depended also on seedlings densities/cell. The best