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Sample records for bacterial cell envelope

  1. Conjugated gold nanoparticles as a tool for probing the bacterial cell envelope: The case of Shewanella oneidensis MR-1.

    Science.gov (United States)

    Jahnke, Justin P; Cornejo, Jose A; Sumner, James J; Schuler, Andrew J; Atanassov, Plamen; Ista, Linnea K

    2016-03-01

    The bacterial cell envelope forms the interface between the interior of the cell and the outer world and is, thus, the means of communication with the environment. In particular, the outer cell surface mediates the adhesion of bacteria to the surface, the first step in biofilm formation. While a number of ligand-based interactions are known for the attachment process in commensal organisms and, as a result, opportunistic pathogens, the process of nonspecific attachment is thought to be mediated by colloidal, physiochemical, interactions. It is becoming clear, however, that colloidal models ignore the heterogeneity of the bacterial surface, and that the so-called nonspecific attachment may be mediated by specific regions of the cell surface, whether or not the relevant interaction is ligand-mediate. The authors introduce surface functionalized gold nanoparticles to probe the surface chemistry of Shewanella oneidensis MR-1 as it relates to surface attachment to ω-substituted alkanethiolates self-assembled monolayers (SAMs). A linear relationship between the attachment of S. oneidensis to SAM modified planar substrates and the number of similarly modified nanoparticles attached to the bacterial surfaces was demonstrated. In addition, the authors demonstrate that carboxylic acid-terminated nanoparticles attach preferentially to the subpolar region of the S. oneidensis and obliteration of that binding preference corresponds in loss of attachment to carboxylic acid terminated SAMs. Moreover, this region corresponds to suspected functional regions of the S. oneidensis surface. Because this method can be employed over large numbers of cells, this method is expected to be generally applicable for understanding cell surface organization across populations. PMID:26746161

  2. Effect of irradiation on kinetic behavior of Salmonella Typhimurium and Staphylococcus aureus in lettuce and damage of bacterial cell envelope

    Science.gov (United States)

    Shim, Won-Bo; Je, Gil-Soo; Kim, Kyeongyeol; Mtenga, Adelard B.; Lee, Won-Gyeong; Song, Jeong-Un; Chung, Duck-Hwa; Yoon, Yohan

    2012-05-01

    This study evaluated effect of gamma irradiation on survival of Salmonella Typhimurium and Staphylococcus aureus on lettuce and damage of cell envelope. S. Typhimurium and S. aureus were inoculated on red leaf lettuce, and they were irradiated at 0, 0.5, 1, 1.5, 2, 2.5, and 3 kGy, and the samples were then stored at 7 and 25 °C for 7 days. Survival of S. Typhimurium and S. aureus were enumerated on xylose lysine deoxycholate agar and Baird-Parker agar, respectively. D10 value (dose required to reduce 1 log CFU/leaf) was calculated, and kinetic parameters (maximum specific growth rate; μmax and lag phase duration; LPD) were calculated by the modified Gompertz model. In addition, cell envelope damage of the pathogens was observed by scanning electron microscope (SEM) and transmission electron microscope (TEM). D10 values were 0.35 and 0.33 kGy for S. Typhimurium and S. aureus, respectively. During storage at 7 °C, S. Typhimurium and S. aureus had significant (Plettuce by destroying cells of S. Typhimurium and S. aureus.

  3. Effect of irradiation on kinetic behavior of Salmonella Typhimurium and Staphylococcus aureus in lettuce and damage of bacterial cell envelope

    International Nuclear Information System (INIS)

    This study evaluated effect of gamma irradiation on survival of Salmonella Typhimurium and Staphylococcus aureus on lettuce and damage of cell envelope. S. Typhimurium and S. aureus were inoculated on red leaf lettuce, and they were irradiated at 0, 0.5, 1, 1.5, 2, 2.5, and 3 kGy, and the samples were then stored at 7 and 25 °C for 7 days. Survival of S. Typhimurium and S. aureus were enumerated on xylose lysine deoxycholate agar and Baird–Parker agar, respectively. D10 value (dose required to reduce 1 log CFU/leaf) was calculated, and kinetic parameters (maximum specific growth rate; μmax and lag phase duration; LPD) were calculated by the modified Gompertz model. In addition, cell envelope damage of the pathogens was observed by scanning electron microscope (SEM) and transmission electron microscope (TEM). D10 values were 0.35 and 0.33 kGy for S. Typhimurium and S. aureus, respectively. During storage at 7 °C, S. Typhimurium and S. aureus had significant (Pmax, respectively. At 25 °C, cell counts of S. Typhimurium and S. aureus on the samples irradiated at 0 and 0.5 kGy increased (Pmax of both pathogens were higher in 0 kGy (1.08–2.27 log CFU/leaf/day) and 0.5 kGy (0.58–0.92 log CFU/leaf/day), and LPDs ranged from 1.53 to 3.14 day. SEM and TEM observations showed that cells irradiated at 1.5 and 3 kGy showed disrupted cell membrane. These results indicate that gamma irradiation could be a useful decontamination technology to improve food safety of lettuce by destroying cells of S. Typhimurium and S. aureus. - Highlights: ► Low dose of gamma irradiation destroyed cell envelope of the pathogens. ► Gamma irradiation decreased cell counts of the pathogens on lettuce. ► Gamma irradiation could be useful in improving food safety of lettuce.

  4. Virulence Properties of the Legionella Pneumophila Cell Envelope

    OpenAIRE

    Shevchuk, Olga; Jäger, Jens; Steinert, Michael

    2011-01-01

    The bacterial envelope plays a crucial role in the pathogenesis of infectious diseases. In this review, we summarize the current knowledge of the structure and molecular composition of the Legionella pneumophila cell envelope. We describe lipopolysaccharides biosynthesis and the biological activities of membrane and periplasmic proteins and discuss their decisive functions during the pathogen–host interaction. In addition to adherence, invasion, and intracellular survival of L. pneumophila, s...

  5. Virulence properties of the Legionella pneumophila cell envelope

    OpenAIRE

    Olga eShevchuk; Jens eJäger; Michael eSteinert

    2011-01-01

    The bacterial envelope plays a crucial role in the pathogenesis of infectious diseases. In this review, we summarize the current knowledge of the structure and molecular composition of the Legionella pneumophila cell envelope. We describe LPS biosynthesis and the biological activities of membrane and periplasmic proteins and discuss their decisive functions during the pathogen-host interaction. In addition to adherence, invasion and intracellular survival of L. pneumophila, special emphasis i...

  6. Virulence properties of the Legionella pneumophila cell envelope

    Directory of Open Access Journals (Sweden)

    Olga eShevchuk

    2011-04-01

    Full Text Available The bacterial envelope plays a crucial role in the pathogenesis of infectious diseases. In this review, we summarize the current knowledge of the structure and molecular composition of the Legionella pneumophila cell envelope. We describe LPS biosynthesis and the biological activities of membrane and periplasmic proteins and discuss their decisive functions during the pathogen-host interaction. In addition to adherence, invasion and intracellular survival of L. pneumophila, special emphasis is laid on iron acquisition, detoxification, key elicitors of the immune response and the diverse functions of outer membrane vesicles. The critical analysis of the literature reveals that the dynamics and phenotypic plasticity of the Legionella cell surface during the different metabolic stages requires more attention in the future.

  7. Genetic interaction maps in Escherichia coli reveal functional crosstalk among cell envelope biogenesis pathways.

    Directory of Open Access Journals (Sweden)

    Mohan Babu

    2011-11-01

    Full Text Available As the interface between a microbe and its environment, the bacterial cell envelope has broad biological and clinical significance. While numerous biosynthesis genes and pathways have been identified and studied in isolation, how these intersect functionally to ensure envelope integrity during adaptive responses to environmental challenge remains unclear. To this end, we performed high-density synthetic genetic screens to generate quantitative functional association maps encompassing virtually the entire cell envelope biosynthetic machinery of Escherichia coli under both auxotrophic (rich medium and prototrophic (minimal medium culture conditions. The differential patterns of genetic interactions detected among > 235,000 digenic mutant combinations tested reveal unexpected condition-specific functional crosstalk and genetic backup mechanisms that ensure stress-resistant envelope assembly and maintenance. These networks also provide insights into the global systems connectivity and dynamic functional reorganization of a universal bacterial structure that is both broadly conserved among eubacteria (including pathogens and an important target.

  8. Atomic force microscopy analysis of enveloped and non-enveloped viral entry into, and egress from, cultured cells

    International Nuclear Information System (INIS)

    Since its invention, the atomic force microscope has been used to image a wide variety of biological samples, including viruses. Viral entry into, and egress from, cultured cells has been extensively studied using numerous scientific techniques and to a limited extent using atomic force microscopy. One of the main structural differences that can exist between viruses is the absence, or presence, of an envelope and this factor has consequences for the mode of viral entry and egress. In this study, the entry into, and egress from, cultured cells of enveloped and non-enveloped viruses were investigated using atomic force microscopy. No significant cell surface changes were observed following infection with enveloped or non-enveloped viruses. Although roughness analysis of viral entry revealed cell smoothing post-infection, no differences between the roughness values of enveloped and non-enveloped viral entry were observed. Line analysis of viral entry revealed minor differences between cells infected with an enveloped rather than a non-enveloped virus. These differences may represent a distinction between the uptake processes of enveloped and non-enveloped viruses. Studies of viral egress revealed that infected cells were undergoing cytopathic changes. Whilst topographic, height and roughness differences clearly occurred between virally- and mock-infected cells, no significant differences were elucidated between enveloped and non-enveloped viral egress

  9. Mast cells in bacterial infections

    OpenAIRE

    Rönnberg, Elin

    2014-01-01

    Mast cells are implicated in immunity towards bacterial infection, but the molecular mechanisms by which mast cells contribute to the host response are only partially understood. Previous studies have examined how mast cells react to purified bacterial cell wall components, such as peptidoglycan and lipopolysaccharide. To investigate how mast cells react to live bacteria we co-cultured mast cells and the gram-positive bacteria Streptococcus equi (S. equi) and Staphylococcus aureus (S. aureus)...

  10. Polymers in cell encapsulation from an enveloped cell perspective.

    Science.gov (United States)

    de Vos, Paul; Lazarjani, Hamideh Aghajani; Poncelet, Denis; Faas, Marijke M

    2014-04-01

    In the past two decades, many polymers have been proposed for producing immunoprotective capsules. Examples include the natural polymers alginate, agarose, chitosan, cellulose, collagen, and xanthan and synthetic polymers poly(ethylene glycol), polyvinyl alcohol, polyurethane, poly(ether-sulfone), polypropylene, sodium polystyrene sulfate, and polyacrylate poly(acrylonitrile-sodium methallylsulfonate). The biocompatibility of these polymers is discussed in terms of tissue responses in both the host and matrix to accommodate the functional survival of the cells. Cells should grow and function in the polymer network as adequately as in their natural environment. This is critical when therapeutic cells from scarce cadaveric donors are considered, such as pancreatic islets. Additionally, the cell mass in capsules is discussed from the perspective of emerging new insights into the release of so-called danger-associated molecular pattern molecules by clumps of necrotic therapeutic cells. We conclude that despite two decades of intensive research, drawing conclusions about which polymer is most adequate for clinical application is still difficult. This is because of the lack of documentation on critical information, such as the composition of the polymer, the presence or absence of confounding factors that induce immune responses, toxicity to enveloped cells, and the permeability of the polymer network. Only alginate has been studied extensively and currently qualifies for application. This review also discusses critical issues that are not directly related to polymers and are not discussed in the other reviews in this issue, such as the functional performance of encapsulated cells in vivo. Physiological endocrine responses may indeed not be expected because of the many barriers that the metabolites encounter when traveling from the blood stream to the enveloped cells and back to circulation. However, despite these diffusion barriers, many studies have shown optimal

  11. Analysis of Pseudomonas aeruginosa cell envelope proteome by capture of surface-exposed proteins on activated magnetic nanoparticles.

    Directory of Open Access Journals (Sweden)

    Davide Vecchietti

    Full Text Available We report on specific magneto-capturing followed by Multidimensional Protein Identification Technology (MudPIT for the analysis of surface-exposed proteins of intact cells of the bacterial opportunistic pathogen Pseudomonas aeruginosa. The magneto-separation of cell envelope fragments from the soluble cytoplasmic fraction allowed the MudPIT identification of the captured and neighboring proteins. Remarkably, we identified 63 proteins captured directly by nanoparticles and 67 proteins embedded in the cell envelope fragments. For a high number of proteins, our analysis strongly indicates either surface exposure or localization in an envelope district. The localization of most identified proteins was only predicted or totally unknown. This novel approach greatly improves the sensitivity and specificity of the previous methods, such as surface shaving with proteases that was also tested on P. aeruginosa. The magneto-capture procedure is simple, safe, and rapid, and appears to be well-suited for envelope studies in highly pathogenic bacteria.

  12. Microarray Analysis to Monitor Bacterial Cell Wall Homeostasis.

    Science.gov (United States)

    Hong, Hee-Jeon; Hesketh, Andy

    2016-01-01

    Transcriptomics, the genome-wide analysis of gene transcription, has become an important tool for characterizing and understanding the signal transduction networks operating in bacteria. Here we describe a protocol for quantifying and interpreting changes in the transcriptome of Streptomyces coelicolor that take place in response to treatment with three antibiotics active against different stages of peptidoglycan biosynthesis. The results defined the transcriptional responses associated with cell envelope homeostasis including a generalized response to all three antibiotics involving activation of transcription of the cell envelope stress sigma factor σ(E), together with elements of the stringent response, and of the heat, osmotic, and oxidative stress regulons. Many antibiotic-specific transcriptional changes were identified, representing cellular processes potentially important for tolerance to each antibiotic. The principles behind the protocol are transferable to the study of cell envelope homeostatic mechanisms probed using alternative chemical/environmental insults or in other bacterial strains. PMID:27311662

  13. Targeting bactoprenol-coupled cell envelope precursors.

    Science.gov (United States)

    Ulm, Hannah; Schneider, Tanja

    2016-09-01

    Targeting the bactoprenol-coupled cell wall precursor lipid II is a validated antibacterial strategy. In this review, selected prototype lipid II-binding antibiotics of different chemical classes are discussed. Although these compounds attack the same molecular target, they trigger nuanced and diverse cellular effects. Consequently, the mechanisms of antibacterial resistance and the likelihood of resistance development may vary substantially. PMID:27495122

  14. Thioridazine Alters the Cell-Envelope Permeability of Mycobacterium tuberculosis.

    Science.gov (United States)

    de Keijzer, Jeroen; Mulder, Arnout; de Haas, Petra E W; de Ru, Arnoud H; Heerkens, Evy M; Amaral, Leonard; van Soolingen, Dick; van Veelen, Peter A

    2016-06-01

    The increasing occurrence of multidrug resistant tuberculosis exerts a major burden on treatment of this infectious disease. Thioridazine, previously used as a neuroleptic, is active against extensively drug resistant tuberculosis when added to other second- and third-line antibiotics. By quantitatively studying the proteome of thioridazine-treated Mycobacterium tuberculosis, we discovered the differential abundance of several proteins that are involved in the maintenance of the cell-envelope permeability barrier. By assessing the accumulation of fluorescent dyes in mycobacterial cells over time, we demonstrate that long-term drug exposure of M. tuberculosis indeed increased the cell-envelope permeability. The results of the current study demonstrate that thioridazine induced an increase in cell-envelope permeability and thereby the enhanced uptake of compounds. These results serve as a novel explanation to the previously reported synergistic effects between thioridazine and other antituberculosis drugs. This new insight in the working mechanism of this antituberculosis compound could open novel perspectives of future drug-administration regimens in combinational therapy. PMID:27068340

  15. Analysis of Pseudomonas aeruginosa Cell Envelope Proteome by Capture of Surface-Exposed Proteins on Activated Magnetic Nanoparticles

    OpenAIRE

    Davide Vecchietti; Dario Di Silvestre; Matteo Miriani; Francesco Bonomi; Mauro Marengo; Alessandra Bragonzi; Lara Cova; Eleonora Franceschi; Pierluigi Mauri; Giovanni Bertoni

    2012-01-01

    We report on specific magneto-capturing followed by Multidimensional Protein Identification Technology (MudPIT) for the analysis of surface-exposed proteins of intact cells of the bacterial opportunistic pathogen Pseudomonas aeruginosa. The magneto-separation of cell envelope fragments from the soluble cytoplasmic fraction allowed the MudPIT identification of the captured and neighboring proteins. Remarkably, we identified 63 proteins captured directly by nanoparticles and 67 proteins embedde...

  16. Bacterial cell culture

    OpenAIRE

    sprotocols

    2014-01-01

    ### Materials 1. Glass culture tubes with metal caps and labels - Growth medium, from media room or customized - Glass pipette tubes - Parafilm ### Equipment 1. Vortexer - Fireboy or Bunsen burner - Motorized pipette - Micropipettes and sterile tips ### Procedure For a typical liquid culture, use 5 ml of appropriate medium. The amount in each tube does not have to be exact if you are just trying to culture cells for their precious DNA. 1. Streak an a...

  17. Ion bombardment induced formation of micro-craters in plant cell envelopes

    International Nuclear Information System (INIS)

    Ion beam bombardment of biological material has been recently applied for gene transfer in both plant and bacterial cells. A consistent physical mechanism for this significant result has not yet been developed. A fundamental question about the mechanism is the possible formation of pathways due to ion bombardment that are responsible for the gene transfer. We have carried out investigations of the effects of low-energy bombardment by both gaseous and metallic ion species of onion skin cells on their surface microstructure. Our experimental results reveal evidence demonstrating that the formation of micro-crater-like structures on the plant cell envelope surface is a general phenomenon consequent to ion bombardment, no matter what ion species, under certain ion beam conditions. The micro-craters are about 0.1-1 μm in size (diameter) and a few tens of nanometers in depth. The micro-crater formation process seems to be unrelated to the chemical composition of and rapid water evaporation from the cell envelope, but is associated with the special microstructure of the cell wall

  18. A cell cycle and nutritional checkpoint controlling bacterial surface adhesion.

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    Aretha Fiebig

    2014-01-01

    Full Text Available In natural environments, bacteria often adhere to surfaces where they form complex multicellular communities. Surface adherence is determined by the biochemical composition of the cell envelope. We describe a novel regulatory mechanism by which the bacterium, Caulobacter crescentus, integrates cell cycle and nutritional signals to control development of an adhesive envelope structure known as the holdfast. Specifically, we have discovered a 68-residue protein inhibitor of holdfast development (HfiA that directly targets a conserved glycolipid glycosyltransferase required for holdfast production (HfsJ. Multiple cell cycle regulators associate with the hfiA and hfsJ promoters and control their expression, temporally constraining holdfast development to the late stages of G1. HfiA further functions as part of a 'nutritional override' system that decouples holdfast development from the cell cycle in response to nutritional cues. This control mechanism can limit surface adhesion in nutritionally sub-optimal environments without affecting cell cycle progression. We conclude that post-translational regulation of cell envelope enzymes by small proteins like HfiA may provide a general means to modulate the surface properties of bacterial cells.

  19. Localization of adhesins on the surface of a pathogenic bacterial envelope through atomic force microscopy

    Science.gov (United States)

    Arnal, L.; Longo, G.; Stupar, P.; Castez, M. F.; Cattelan, N.; Salvarezza, R. C.; Yantorno, O. M.; Kasas, S.; Vela, M. E.

    2015-10-01

    Bacterial adhesion is the first and a significant step in establishing infection. This adhesion normally occurs in the presence of flow of fluids. Therefore, bacterial adhesins must be able to provide high strength interactions with their target surface in order to maintain the adhered bacteria under hydromechanical stressing conditions. In the case of B. pertussis, a Gram-negative bacterium responsible for pertussis, a highly contagious human respiratory tract infection, an important protein participating in the adhesion process is a 220 kDa adhesin named filamentous haemagglutinin (FHA), an outer membrane and also secreted protein that contains recognition domains to adhere to ciliated respiratory epithelial cells and macrophages. In this work, we obtained information on the cell-surface localization and distribution of the B. pertussis adhesin FHA using an antibody-functionalized AFM tip. Through the analysis of specific molecular recognition events we built a map of the spatial distribution of the adhesin which revealed a non-homogeneous pattern. Moreover, our experiments showed a force induced reorganization of the adhesin on the surface of the cells, which could explain a reinforced adhesive response under external forces. This single-molecule information contributes to the understanding of basic molecular mechanisms used by bacterial pathogens to cause infectious disease and to gain insights into the structural features by which adhesins can act as force sensors under mechanical shear conditions.Bacterial adhesion is the first and a significant step in establishing infection. This adhesion normally occurs in the presence of flow of fluids. Therefore, bacterial adhesins must be able to provide high strength interactions with their target surface in order to maintain the adhered bacteria under hydromechanical stressing conditions. In the case of B. pertussis, a Gram-negative bacterium responsible for pertussis, a highly contagious human respiratory tract

  20. ESCRT III repairs nuclear envelope ruptures during cell migration to limit DNA damage and cell death.

    Science.gov (United States)

    Raab, M; Gentili, M; de Belly, H; Thiam, H R; Vargas, P; Jimenez, A J; Lautenschlaeger, F; Voituriez, Raphaël; Lennon-Duménil, A M; Manel, N; Piel, M

    2016-04-15

    In eukaryotic cells, the nuclear envelope separates the genomic DNA from the cytoplasmic space and regulates protein trafficking between the two compartments. This barrier is only transiently dissolved during mitosis. Here, we found that it also opened at high frequency in migrating mammalian cells during interphase, which allowed nuclear proteins to leak out and cytoplasmic proteins to leak in. This transient opening was caused by nuclear deformation and was rapidly repaired in an ESCRT (endosomal sorting complexes required for transport)-dependent manner. DNA double-strand breaks coincided with nuclear envelope opening events. As a consequence, survival of cells migrating through confining environments depended on efficient nuclear envelope and DNA repair machineries. Nuclear envelope opening in migrating leukocytes could have potentially important consequences for normal and pathological immune responses. PMID:27013426

  1. Fourier and spectral envelope analysis of medically important bacterial and fungal sequences

    OpenAIRE

    Chang, C.; Chan, KKH; Chan, FHY

    2004-01-01

    In this paper, we introduce the Fourier and spectral envelope analysis methods to analyze some biomolecular sequences, particularly medically important bacteria and fungi DNA sequences, to get their interesting frequency properties. Fourier analysis includes mapping character strings into numerical sequences, calculating spectra of DNA sequences and setting and solving optimization problem in order to construct a powerful predictor of exons along the long DNA sequences. The spectral envelope ...

  2. Synthesis of cell envelope glycoproteins of Cryptococcus laurentii.

    Science.gov (United States)

    Schutzbach, John; Ankel, Helmut; Brockhausen, Inka

    2007-05-21

    Fungi of the genus Cryptococcus are encapsulated basidiomycetes that are ubiquitously found in the environment. These organisms infect both lower and higher animals. Human infections that are common in immune-compromised individuals have proven difficult to cure or even control with currently available antimycotics that are quite often toxic to the host. The virulence of Cryptococcus has been linked primarily to its polysaccharide capsule, but also to cell-bound glycoproteins. In this review, we show that Cryptococcus laurentii is an excellent model for studies of polysaccharide and glycoprotein synthesis in the more pathogenic relative C. neoformans. In particular, we will discuss the structure and biosynthesis of O-linked carbohydrates on cell envelope glycoproteins of C. laurentii. These O-linked structures are synthesized by at least four mannosyltransferases, two galactosyltransferases, and at least one xylosyltransferase that have been characterized. These glycosyltransferases have no known homologues in human tissues. Therefore, enzymes involved in the synthesis of cryptococcal glycoproteins, as well as related enzymes involved in capsule synthesis, are potential targets for the development of specific inhibitors for treatment of cryptococcal disease. PMID:17316583

  3. Modeling bacterial chemotaxis inside a cell

    OpenAIRE

    Ouannes, Nesrine; Djedi, Noureddine; Luga, Hervé; Duthen, Yves

    2014-01-01

    This paper describes a bacterial system that reproduces a population of bacteria that behave by simulating the internal reactions of each bacterial cell. The chemotaxis network of a cell is modulated by a hybrid approach that uses an algebraic model for the receptor clusters activity and an ordinary differential equation for the adaptation dynamics. The experiments are defined in order to simulate bacterial growth in an environment where nutrients are regularly added to it. The results show a...

  4. Bacterial cell division proteins as antibiotic targets

    NARCIS (Netherlands)

    T. den Blaauwen; J.M. Andreu; O. Monasterio

    2014-01-01

    Proteins involved in bacterial cell division often do not have a counterpart in eukaryotic cells and they are essential for the survival of the bacteria. The genetic accessibility of many bacterial species in combination with the Green Fluorescence Protein revolution to study localization of protein

  5. Penicillin-binding site on the Escherichia coli cell envelope

    International Nuclear Information System (INIS)

    The binding of 35S-labeled penicillin to distinct penicillin-binding proteins (PBPs) of the cell envelope obtained from the sonication of Escherichia coli was studied at different pHs ranging from 4 to 11. Experiments distinguishing the effect of pH on penicillin binding by PBP 5/6 from its effect on beta-lactamase activity indicated that although substantial binding occurred at the lowest pH, the amount of binding increased with pH, reaching a maximum at pH 10. Based on earlier studies, it is proposed that the binding at high pH involves the formation of a covalent bond between the C-7 of penicillin and free epsilon amino groups of the PBPs. At pHs ranging from 4 to 8, position 1 of penicillin, occupied by sulfur, is considered to be the site that establishes a covalent bond with the sulfhydryl groups of PBP 5. The use of specific blockers of free epsilon amino groups or sulfhydryl groups indicated that wherever the presence of each had little or no effect on the binding of penicillin by PBP 5, the presence of both completely prevented binding. The specific blocker of the hydroxyl group of serine did not affect the binding of penicillin

  6. EXPRESSION EFFECT OF RECOMBINANT ENVELOPE GENE OF AVIAN LEUKOSIS VIRUS SUBGROUP J IN SF 9 CELLS

    Science.gov (United States)

    Expression effect of envelope gene of avian leukosis virus (ALV-J) in Sf9 cells infected with recombinant baculovirus rBac-env was analyzed by immunofluorescent assay and immunoprecipitation. The results showed that recombinant envelope gene product was a glycosylated protein in tunicumycin treatme...

  7. Bacterial Probiotic Modulation of Dendritic Cells

    OpenAIRE

    Drakes, Maureen; Blanchard, Thomas; Czinn, Steven

    2004-01-01

    Intestinal dendritic cells are continually exposed to ingested microorganisms and high concentrations of endogenous bacterial flora. These cells can be activated by infectious agents and other stimuli to induce T-cell responses and to produce chemokines which recruit other cells to the local environment. Bacterial probiotics are of increasing use against intestinal disorders such as inflammatory bowel disease. They act as nonpathogenic stimuli within the gut to regain immunologic quiescence. ...

  8. Identification of bacterial cells by chromosomal painting.

    OpenAIRE

    Lanoil, B. D.; Giovannoni, S J

    1997-01-01

    Chromosomal painting is a technique for the microscopic localization of genetic material. It has been applied at the subcellular level to identify regions of eukaryotic chromosomes. Here we describe the development of bacterial chromosomal painting (BCP), a related technology for the identification of bacterial cells. Purified genomic DNAs from six bacterial strains were labeled by nick translation with the fluorochrome Fluor-X, Cy3, or Cy5. The average size of the labeled fragments was ca. 5...

  9. Detection of an Immunogenic HERV-E Envelope with Selective Expression in Clear Cell Kidney Cancer.

    Science.gov (United States)

    Cherkasova, Elena; Scrivani, Claire; Doh, Susan; Weisman, Quinn; Takahashi, Yoshiyuki; Harashima, Nanae; Yokoyama, Hisayuki; Srinivasan, Ramaprasad; Linehan, W Marston; Lerman, Michael I; Childs, Richard W

    2016-04-15

    VHL-deficient clear cell renal cell carcinomas (ccRCC), the most common form of kidney cancer, express transcripts derived from the novel human endogenous retrovirus HERV-E (named CT-RCC HERV-E). In this study, we define a transcript encoding the entire envelope gene of HERV-E as expressed selectively in ccRCC tumors, as distinct from normal kidney tissues or other tumor types. Sequence analysis of this envelope transcript revealed long open reading frames encoding putative surface and transmembrane envelope proteins. Retroviral envelopes are known to be capable of eliciting immunity in humans. Accordingly, we found that HLA-A*0201-restricted peptides predicted to be products of the CT-RCC HERV-E envelope transcript-stimulated CD8(+) T cells, which could recognize HLA-A*0201-positive HERV-E-expressing kidney tumor cells. Overall, our results offer evidence of unique HERV-E envelope peptides presented on the surface of ccRCC cells, offering potentially useful tumor-restricted targets for T-cell-based immunotherapy of kidney cancer. Cancer Res; 76(8); 2177-85. ©2016 AACR. PMID:26862115

  10. Original features of cell-envelope proteinases of Lactobacillus helveticus. A review.

    Science.gov (United States)

    Sadat-Mekmene, Leila; Genay, Magali; Atlan, Danièle; Lortal, Sylvie; Gagnaire, Valérie

    2011-03-15

    Lactobacillus helveticus is a lactic acid bacterium very used in fermented milks and cheese. The rapid growth of L. helveticus in milk is supported by an efficient cell envelope proteinase (CEP) activity, due to subtilisin-like serine proteases. These enzymes play also crucial roles in texture and flavor formation in dairy products as well as in generating in situ bioactive peptides. In L. helveticus, several genes encoding putative CEPs were detected and characterized by a large intraspecific diversity; little is known about regulation of expression of CEP-encoding genes. Anchored at the bacterial surface, CEPs are large-sized enzymes (> 150 kDa) hydrolyzing β- and α(s1)-casein as well. Substrate cleavages occur after almost all types of amino acids residues, but mass spectrometry analysis revealed L. helveticus strains with specific profiles of substrate hydrolysis, which could explain identification of strains associated with interesting technological properties. In this review, the most recent data regarding CEP-encoding genes, CEP activities toward caseins and L. helveticus strain diversity are discussed. PMID:21354644

  11. Brucella abortus choloylglycine hydrolase affects cell envelope composition and host cell internalization.

    Directory of Open Access Journals (Sweden)

    María Inés Marchesini

    Full Text Available Choloylglycine hydrolase (CGH, E.C. 3.5.1.24 is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization.

  12. Two-component system cross-regulation integrates Bacillus anthracis response to heme and cell envelope stress.

    Directory of Open Access Journals (Sweden)

    Laura A Mike

    2014-03-01

    Full Text Available Two-component signaling systems (TCSs are one of the mechanisms that bacteria employ to sense and adapt to changes in the environment. A prototypical TCS functions as a phosphorelay from a membrane-bound sensor histidine kinase (HK to a cytoplasmic response regulator (RR that controls target gene expression. Despite significant homology in the signaling domains of HKs and RRs, TCSs are thought to typically function as linear systems with little to no cross-talk between non-cognate HK-RR pairs. Here we have identified several cell envelope acting compounds that stimulate a previously uncharacterized Bacillus anthracis TCS. Furthermore, this TCS cross-signals with the heme sensing TCS HssRS; therefore, we have named it HssRS interfacing TCS (HitRS. HssRS reciprocates cross-talk to HitRS, suggesting a link between heme toxicity and cell envelope stress. The signaling between HssRS and HitRS occurs in the parental B. anthracis strain; therefore, we classify HssRS-HitRS interactions as cross-regulation. Cross-talk between HssRS and HitRS occurs at both HK-RR and post-RR signaling junctions. Finally, HitRS also regulates a previously unstudied ABC transporter implicating this transporter in the response to cell envelope stress. This chemical biology approach to probing TCS signaling provides a new model for understanding how bacterial signaling networks are integrated to enable adaptation to complex environments such as those encountered during colonization of the vertebrate host.

  13. Messenger Functions of the Bacterial Cell Wall-derived Muropeptides

    OpenAIRE

    Boudreau, Marc A.; Fisher, Jed F.; Mobashery, Shahriar

    2012-01-01

    Bacterial muropeptides are soluble peptidoglycan structures central to recycling of the bacterial cell wall, and messengers in diverse cell-signaling events. Bacteria sense muropeptides as signals that antibiotics targeting cell-wall biosynthesis are present, and eukaryotes detect muropeptides during the innate immune response to bacterial infection. This review summarizes the roles of bacterial muropeptides as messengers, with a special emphasis on bacterial muropeptide structures and the re...

  14. Disturbance of the bacterial cell wall specifically interferes with biofilm formation.

    Science.gov (United States)

    Bucher, Tabitha; Oppenheimer-Shaanan, Yaara; Savidor, Alon; Bloom-Ackermann, Zohar; Kolodkin-Gal, Ilana

    2015-12-01

    In nature, bacteria communicate via chemical cues and establish complex communities referred to as biofilms, wherein cells are held together by an extracellular matrix. Much research is focusing on small molecules that manipulate and prevent biofilm assembly by modifying cellular signalling pathways. However, the bacterial cell envelope, presenting the interface between bacterial cells and their surroundings, is largely overlooked. In our study, we identified specific targets within the biosynthesis pathways of the different cell wall components (peptidoglycan, wall teichoic acids and teichuronic acids) hampering biofilm formation and the anchoring of the extracellular matrix with a minimal effect on planktonic growth. In addition, we provide convincing evidence that biofilm hampering by transglycosylation inhibitors and D-Leucine triggers a highly specific response without changing the overall protein levels within the biofilm cells or the overall levels of the extracellular matrix components. The presented results emphasize the central role of the Gram-positive cell wall in biofilm development, resistance and sustainment. PMID:26472159

  15. The bacterial carbon-fixing organelle is formed by shell envelopment of preassembled cargo.

    Directory of Open Access Journals (Sweden)

    Anna H Chen

    Full Text Available BACKGROUND: Cyanobacteria play a significant role in the global carbon cycle. In Synechococcuselongatus, the carbon-fixing enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO is concentrated into polyhedral, proteinaceous compartments called carboxysomes. METHODOLOGY/PRINCIPAL FINDINGS: Using live cell fluorescence microscopy, we show that carboxysomes are first detected as small seeds of RuBisCO that colocalize with existing carboxysomes. These seeds contain little or no shell protein, but increase in RuBisCO content over several hours, during which time they are exposed to the solvent. The maturing seed is then enclosed by shell proteins, a rapid process that seals RuBisCO from the cytosol to establish a distinct, solvent-protected microenvironment that is oxidizing relative to the cytosol. These closure events can be spatially and temporally coincident with the appearance of a nascent daughter RuBisCO seed. CONCLUSIONS/SIGNIFICANCE: Carboxysomes assemble in a stepwise fashion, inside-to-outside, revealing that cargo is the principle organizer of this compartment's biogenesis. Our observations of the spatial relationship of seeds to previously formed carboxysomes lead us to propose a model for carboxysome replication via sequential fission, polymerization, and encapsulation of their internal cargo.

  16. Targeting HIV-1 Envelope Glycoprotein Trimers to B Cells by Using APRIL Improves Antibody Responses

    OpenAIRE

    Melchers M; Bontjer I; Tong T; Chung NP; Klasse PJ; Eggink D; Montefiori DC; Gentile M; Cerutti A; Olson WC; Berkhout B; Binley JM; Moore JP; Sanders RW

    2012-01-01

    An HIV-1 vaccine remains elusive, in part because various factors limit the quantity and quality of the antibodies raised against the viral envelope glycoprotein complex (Env). We hypothesized that targeting Env vaccines directly to B cells, by fusing them to molecules that bind and activate these cells, would improve Env-specific antibody responses. Therefore, we fused trimeric Env gp140 to A PRoliferation-Inducing Ligand (APRIL), B-cell Activating Factor (BAFF), and CD40 Ligand (CD40L). The...

  17. Quantitative analysis of the lipidomes of the influenza virus envelope and MDCK cell apical membrane

    OpenAIRE

    Gerl, Mathias J.; Sampaio, Julio L; Urban, Severino; Kalvodova, Lucie; Verbavatz, Jean-Marc; Binnington, Beth; Lindemann, Dirk; Lingwood, Clifford A.; Shevchenko, Andrej; Schroeder, Cornelia; Simons, Kai

    2012-01-01

    The influenza virus (IFV) acquires its envelope by budding from host cell plasma membranes. Using quantitative shotgun mass spectrometry, we determined the lipidomes of the host Madin–Darby canine kidney cell, its apical membrane, and the IFV budding from it. We found the apical membrane to be enriched in sphingolipids (SPs) and cholesterol, whereas glycerophospholipids were reduced, and storage lipids were depleted compared with the whole-cell membranes. The virus membrane exhibited a furthe...

  18. Human immunodeficiency virus-1 gp120 and gp160 envelope proteins modulate mesangial cell gelatinolytic activity.

    OpenAIRE

    Singhal, P. C.; Sagar, S.; D. Chandra; Garg, P

    1995-01-01

    Patients with human immunodeficiency virus (HIV) infection often develop glomerular lesions (mesangial expansion and sclerosis). Modulation of matrix degradation may be important in the expansion of the mesangium. We studied the effect of HIV sera and HIV-1 envelope glycoproteins on gelatinolytic activity of human mesangial cells. HIV serum-treated cells showed lower (P < 0.01) gelatinolytic activity when compared with cells treated with control serum (control serum, 4.3 +/- 0.1 versus HIV se...

  19. A chimeric measles virus with a lentiviral envelope replicates exclusively in CD4+/CCR5+ cells

    International Nuclear Information System (INIS)

    We generated a replicating chimeric measles virus in which the hemagglutinin and fusion surface glycoproteins were replaced with the gp160 envelope glycoprotein of simian immunodeficiency virus (SIVmac239). Based on a previously cloned live-attenuated Schwarz vaccine strain of measles virus (MV), this chimera was rescued at high titers using reverse genetics in CD4+ target cells. Cytopathic effect consisted in the presence of large cell aggregates evolving to form syncytia, as observed during SIV infection. The morphology of the chimeric virus was identical to that of the parent MV particles. The presence of SIV gp160 as the only envelope protein on chimeric particles surface altered the cell tropism of the new virus from CD46+ to CD4+ cells. Used as an HIV candidate vaccine, this MV/SIVenv chimeric virus would mimic transient HIV-like infection, benefiting both from HIV-like tropism and the capacity of MV to replicate in dendritic cells, macrophages and lymphocytes.

  20. Defining the N-linked glycosylation site of Hantaan virus envelope glycoproteins essential for cell fusion.

    Science.gov (United States)

    Zheng, Feng; Ma, Lixian; Shao, Lihua; Wang, Gang; Chen, Fengzhe; Zhang, Ying; Yang, Song

    2007-02-01

    The Hantaan virus (HTNV) is an enveloped virus that is capable of inducing low pH-dependent cell fusion. We molecularly cloned the viral glycoprotein (GP) and nucleocapsid (NP) cDNA of HTNV and expressed them in Vero E6 cells under the control of a CMV promoter. The viral gene expression was assessed using an indirect immunofluorescence assay and immunoprecipitation. The transfected Vero E6 cells expressing GPs, but not those expressing NP, fused and formed a syncytium following exposure to a low pH. Monoclonal antibodies (MAbs) against envelope GPs inhibited cell fusion, whereas MAbs against NP did not. We also investigated the N-linked glycosylation of HTNV GPs and its role in cell fusion. The envelope GPs of HTNV are modified by N-linked glycosylation at five sites: four sites on G1 (N134, N235, N347, and N399) and one site on G2 (N928). Site-directed mutagenesis was used to construct eight GP gene mutants, including five single N-glycosylation site mutants and three double-site mutants, which were then expressed in Vero E6 cells. The oligosaccharide chain on residue N928 of G2 was found to be crucial for cell fusion after exposure to a low pH. These results suggest that G2 is likely to be the fusion protein of HTNV. PMID:17342054

  1. Green Fluorescent Protein-Tagged Retroviral Envelope Protein for Analysis of Virus-Cell Interactions

    Science.gov (United States)

    Spitzer, Dirk; Dittmar, Kurt E. J.; Rohde, Manfred; Hauser, Hansjörg; Wirth, Dagmar

    2003-01-01

    Fluorescent retroviral envelope (Env) proteins were developed for direct visualization of viral particles. By fusing the enhanced green fluorescent protein (eGFP) to the N terminus of the amphotropic 4070A envelope protein, extracellular presentation of eGFP was achieved. Viruses incorporated the modified Env protein and efficiently infected cells. We used the GFP-tagged viruses for staining retrovirus receptor-positive cells, thereby circumventing indirect labeling techniques. By generating cells which conditionally expressed the GFP-tagged Env protein, we could confirm an inverse correlation between retroviral Env expression and infectivity (superinfection). eGFP-tagged virus particles are suitable for monitoring the dynamics of virus-cell interactions. PMID:12719600

  2. Rupture of the Cell Envelope by Decompression of the Deep-Sea Methanogen Methanococcus jannaschii

    OpenAIRE

    Park, Chan Beum; Clark, Douglas S.

    2002-01-01

    The effect of decompression on the structure of Methanococcus jannaschii, an extremely thermophilic deep-sea methanogen, was studied in a novel high-pressure, high-temperature bioreactor. The cell envelope of M. jannaschii appeared to rupture upon rapid decompression (ca. 1 s) from 260 atm of hyperbaric pressure. When decompression from 260 atm was performed over 5 min, the proportion of ruptured cells decreased significantly. In contrast to the effect produced by decompression from hyperbari...

  3. Analysis of 4070A envelope levels in retroviral preparations and effect on target cell transduction efficiency.

    Science.gov (United States)

    Slingsby, J H; Baban, D; Sutton, J; Esapa, M; Price, T; Kingsman, S M; Kingsman, A J; Slade, A

    2000-07-01

    A number of stable producer cell lines for high-titer Mo-MuLV vectors have been constructed. Development has previously centered on increasing end-point titers by producing maximal levels of Mo-MuLV Gag/Pol, envelope glycoproteins, and retroviral RNA genomes. We describe the production yields and transduction efficiency characteristics of two Mo-MuLV packaging cell lines, FLYA13 and TEFLYA. Although they both produce 4070A-pseudotyped retroviral vectors reproducibly at >1 x 10(6) LFU ml(-1), the transduction efficiency of unconcentrated and concentrated virus from FLYA13 lines is poor compared with vector preparations from TEFLYA lines. A powerful inhibitor of retroviral transduction is secreted by FLYA13 packaging cells. We show that the inhibitory factor does not affect transduction of target cells by RD114-pseudotyped vectors. This suggests that the inhibitory factor functions at the level of envelope-receptor interactions. Phosphate starvation of target cells shows a two-fold increase in Pit2 receptor mRNA and causes some improvement in FLYA13 virus transduction efficiency. Western blots show that FLYA13 viral samples contain an eight-fold higher ratio of 4070A envelope to p30gag than that of virus produced by TEFLYA producer cell lines. This study correlates overexpression of 4070A envelope glycoprotein in retroviral preparations with a reduction of transduction efficiency at high multiplicities of infection. We suggest that TEFLYA packaging cells express preferable levels of 4070A compared with FLYA13, which not only enables high-titer stocks to be generated, but also facilitates a high efficiency of transduction of target cells. PMID:10910141

  4. Random Transposon Mutagenesis for Cell-Envelope Resistant to Phage Infection.

    Science.gov (United States)

    Reyes-Cortés, Ruth; Arguijo-Hernández, Emma S; Carballo-Ontiveros, Marco A; Martínez-Peñafiel, Eva; Kameyama, Luis

    2016-01-01

    In order to identify host components involved in the infective process of bacteriophages, we developed a wide-range strategy to obtain cell envelope mutants, using Escherichia coli W3110 and its specific phage mEp213. The strategy consisted in four steps: (1) random mutagenesis using transposon miniTn10Km(r); (2) selection of phage-resistant mutants by replica-plating; (3) electroporation of the phage-resistant mutants with mEp213 genome, followed by selection of those allowing phage development; and (4) sequencing of the transposon-disrupted genes. This strategy allowed us to distinguish the host factors related to phage development or multiplication within the cell, from those involved in phage infection at the level of the cell envelope. PMID:27311665

  5. Expression of bacterial luciferase in eukaryotic cells

    International Nuclear Information System (INIS)

    Expression of Bacterial luciferase enzyme (lux) in mammalian cells would be a powerful bioreporter protein system for in vivo imaging because eukaryotic luciferases need expensive substrates. However, only a few efforts have been made to express bacterial luciferase enzyme in mammalian cells. As the result of this, we attempted to construct bicistronic vector including two bacterial luciferase genes (LuxA and LuxB) for assessing the potential to be visualized in vitro or in vivo by optical imaging system after transfection to mammalian cells. We designed and synthesized luxA and luxB genes from Photorhabdus Luminescens. To co-express both luxA and luxB genes from a single promoter, we cloned as a bicistronic transcript fused with an internal ribosomal entry site (IRES). This bicistronic transcript was transfected by Superfect to HEK 293T cell line. We also transfected lux A and lux B vector to HEK 293T cells separately. To evaluate gene expression, n-decanal and FMNH2 were supplemented to transfected HEK 293T cell lines which were measured by In Vivo Imaging System. The luxA gene was cloned into the MCS(A) of pIRESGFP via the 5' SalI and 3' EcoRI restriction sites to generate pIRESluxA. The luxB gene was cleaved via a 5' NcoI and 3' NotI site from luxB and cloned into the MCS(B) of pIRESluxA to generate pIRESluxAB. LuxA and B genes was cleaved by 5' EcoRI and 3' SpeI and cloned into the pcDNA3.1 mammalian expression vector to create pcDNALuxA and pcDNALuxB. We constructed bicistronic vector system which is composed of bacterial luciferase genes (lux A and B) on the single reading frame. These results hold a promise of an available development of an autonomous light generating lux reporter system in mammalian cells

  6. GAGE cancer-germline antigens are recruited to the nuclear envelope by germ cell-less (GCL)

    DEFF Research Database (Denmark)

    Gjerstorff, Morten F; Rösner, Heike I; Pedersen, Christina B;

    2012-01-01

    metazoan transcriptional regulator, Germ cell-less (GCL), as an interaction partner of GAGE12I. GCL directly binds LEM-domain proteins (LAP2β, emerin, MAN1) at the nuclear envelope, and we found that GAGE proteins were recruited to the nuclear envelope inner membrane by GCL. Based on yeast two...

  7. Expression of hepatitis C virus envelope protein 2 induces apoptosis in cultured mammalian cells

    Institute of Scientific and Technical Information of China (English)

    Li-Xin Zhu; Jing Liu; You-Hua Xie; Yu-Ying Kong; Ye Ye; Chun-Lin Wang; Guang-Di Li; Yuan Wang

    2004-01-01

    AIM: To explore the role of hepatitis C virus (HCV) envelope protein 2 (E2) in the induction of apoptosis.METHODS: A carboxyterminal truncated E2 (E2-661) was transiently expressed in several cultured mammalian cell lines or stably expressed in Chinese hamster ovary (CHO)cell line. Cell proliferation was assessed by 3H thymidine uptake. Apoptosis was examined by Hoechst 33258staining, flow cytometry and DNA fragmentation analysis.RESULTS: Reduced proliferation was readily observed in the E2-661 expressing cells. These cells manifested the typical features of apoptosis, including cell shrinkage,chromatin condensation and hypodiploid genomic DNA content. Similar apoptotic cell death was observed in an E2-661 stably expressing cell line.CONCLUSION: HCV E2 can induce apoptosis in cultured mammalian cells.

  8. HIV-1 Envelope Induces Memory B Cell Responses That Correlate with Plasma Antibody Levels after Envelope gp120 Protein Vaccination or HIV-1 Infection1

    OpenAIRE

    Bonsignori, Mattia; Moody, M. Anthony; Parks, Robert J.; Holl, T. Matt; Kelsoe, Garnett; Hicks, Charles B.; Vandergrift, Nathan; Tomaras, Georgia D.; Haynes, Barton F.

    2009-01-01

    Successful vaccines (i.e., tetanus and diphtheria) can induce long-lived Ab levels that are maintained by bone marrow plasma cells and plasma Ab levels do not correlate with numbers of blood memory B cells. Destruction of CD4+ T cells early in HIV-1 acute infection may result in insufficient induction of neutralizing Ab responses; thus, an HIV-1 vaccine should elicit high levels of durable Abs by long-lived plasma cells to be protective. We asked if HIV-1 envelope-specific memory responses we...

  9. Translucent load-bearing GFRP envelopes for daylighting and solar cell integration in building construction

    OpenAIRE

    Pascual Agullo, Carlos

    2014-01-01

    This project investigates the light transmittance of load-bearing glass fiber-reinforced polymer (GFRP) laminates with a view to two architectural applications: the daylighting of buildings through load-bearing translucent GFRP envelopes and encapsulation of solar cells into the GFRP building skins of sandwich structures. The total and diffuse visible light transmittances of the laminates were experimentally investigated using a spectrophotometer coupled to an integrating sphere. The refracti...

  10. In vitro and in vivo screening for novel essential cell-envelope proteins in Pseudomonas aeruginosa

    OpenAIRE

    Regina Fernández-Piñar; Alessandra Lo Sciuto; Alice Rossi; Serena Ranucci; Alessandra Bragonzi; Francesco Imperi

    2015-01-01

    The Gram-negative bacterium Pseudomonas aeruginosa represents a prototype of multi-drug resistant opportunistic pathogens for which novel therapeutic options are urgently required. In order to identify new candidates as potential drug targets, we combined large-scale transposon mutagenesis data analysis and bioinformatics predictions to retrieve a set of putative essential genes which are conserved in P. aeruginosa and predicted to encode cell envelope or secreted proteins. By generating unma...

  11. Subfractionation and analysis of the cell envelope (lipo)polysaccharides of Mycobacterium tuberculosis

    OpenAIRE

    Grzegorzewicz, Anna E.; Jackson, Mary

    2013-01-01

    The cell envelope of Mycobacterium tuberculosis, the causative agent of tuberculosis in humans, is the source of carbohydrates of exceptional structure which play essential roles in the physiology of the bacterium and in its interactions with the host during infection. Much of what is known about their biosynthesis was derived from the phenotypic analysis of knock-out or conditional knock-out mutants of Mycobacteria generated by random or specific insertional mutagenesis. Here, we describe th...

  12. Probing bacterial adhesion at the single-cell level

    DEFF Research Database (Denmark)

    Zeng, Guanghong; Müller, Torsten; Meyer, Rikke Louise

    cantilever coated with the commercial cell adhesive CellTakTM. We applied the method to study adhesion of living cells to abiotic surfaces at the single-cell level. Immobilisation of single bacterial cells to the cantilever was stable for several hours, and viability was confirmed by Live/Dead staining and......Bacteria initiate attachment to surfaces with the aid of different extracellular proteins and polymeric adhesins. To quantitatively analyse the cell-cell and cell-surface interactions provided by bacterial adhesins, it is essential to go down to single cell level where cell-to-cell variation can be...... considered. We have developed a simple and versatile method to make single-cell bacterial probes for measuring single cell adhesion by force spectroscopy using atomic force microscopy (AFM). A single-cell probe was readily made by picking up a bacterial cell from a glass surface by approaching a tipless AFM...

  13. Structure of a Pestivirus Envelope Glycoprotein E2 Clarifies Its Role in Cell Entry

    Directory of Open Access Journals (Sweden)

    Kamel El Omari

    2013-01-01

    Full Text Available Enveloped viruses have developed various adroit mechanisms to invade their host cells. This process requires one or more viral envelope glycoprotein to achieve cell attachment and membrane fusion. Members of the Flaviviridae such as flaviviruses possess only one envelope glycoprotein, E, whereas pestiviruses and hepacivirus encode two glycoproteins, E1 and E2. Although E2 is involved in cell attachment, it has been unclear which protein is responsible for membrane fusion. We report the crystal structures of the homodimeric glycoprotein E2 from the pestivirus bovine viral diarrhea virus 1 (BVDV1 at both neutral and low pH. Unexpectedly, BVDV1 E2 does not have a class II fusion protein fold, and at low pH the N-terminal domain is disordered, similarly to the intermediate postfusion state of E2 from sindbis virus, an alphavirus. Our results suggest that the pestivirus and possibly the hepacivirus fusion machinery are unlike any previously observed.

  14. One Bacterial Cell, One Complete Genome

    Energy Technology Data Exchange (ETDEWEB)

    Woyke, Tanja; Tighe, Damon; Mavrommatis, Konstantinos; Clum, Alicia; Copeland, Alex; Schackwitz, Wendy; Lapidus, Alla; Wu, Dongying; McCutcheon, John P.; McDonald, Bradon R.; Moran, Nancy A.; Bristow, James; Cheng, Jan-Fang

    2010-04-26

    While the bulk of the finished microbial genomes sequenced to date are derived from cultured bacterial and archaeal representatives, the vast majority of microorganisms elude current culturing attempts, severely limiting the ability to recover complete or even partial genomes from these environmental species. Single cell genomics is a novel culture-independent approach, which enables access to the genetic material of an individual cell. No single cell genome has to our knowledge been closed and finished to date. Here we report the completed genome from an uncultured single cell of Candidatus Sulcia muelleri DMIN. Digital PCR on single symbiont cells isolated from the bacteriome of the green sharpshooter Draeculacephala minerva bacteriome allowed us to assess that this bacteria is polyploid with genome copies ranging from approximately 200?900 per cell, making it a most suitable target for single cell finishing efforts. For single cell shotgun sequencing, an individual Sulcia cell was isolated and whole genome amplified by multiple displacement amplification (MDA). Sanger-based finishing methods allowed us to close the genome. To verify the correctness of our single cell genome and exclude MDA-derived artifacts, we independently shotgun sequenced and assembled the Sulcia genome from pooled bacteriomes using a metagenomic approach, yielding a nearly identical genome. Four variations we detected appear to be genuine biological differences between the two samples. Comparison of the single cell genome with bacteriome metagenomic sequence data detected two single nucleotide polymorphisms (SNPs), indicating extremely low genetic diversity within a Sulcia population. This study demonstrates the power of single cell genomics to generate a complete, high quality, non-composite reference genome within an environmental sample, which can be used for population genetic analyzes.

  15. One bacterial cell, one complete genome.

    Directory of Open Access Journals (Sweden)

    Tanja Woyke

    Full Text Available While the bulk of the finished microbial genomes sequenced to date are derived from cultured bacterial and archaeal representatives, the vast majority of microorganisms elude current culturing attempts, severely limiting the ability to recover complete or even partial genomes from these environmental species. Single cell genomics is a novel culture-independent approach, which enables access to the genetic material of an individual cell. No single cell genome has to our knowledge been closed and finished to date. Here we report the completed genome from an uncultured single cell of Candidatus Sulcia muelleri DMIN. Digital PCR on single symbiont cells isolated from the bacteriome of the green sharpshooter Draeculacephala minerva bacteriome allowed us to assess that this bacteria is polyploid with genome copies ranging from approximately 200-900 per cell, making it a most suitable target for single cell finishing efforts. For single cell shotgun sequencing, an individual Sulcia cell was isolated and whole genome amplified by multiple displacement amplification (MDA. Sanger-based finishing methods allowed us to close the genome. To verify the correctness of our single cell genome and exclude MDA-derived artifacts, we independently shotgun sequenced and assembled the Sulcia genome from pooled bacteriomes using a metagenomic approach, yielding a nearly identical genome. Four variations we detected appear to be genuine biological differences between the two samples. Comparison of the single cell genome with bacteriome metagenomic sequence data detected two single nucleotide polymorphisms (SNPs, indicating extremely low genetic diversity within a Sulcia population. This study demonstrates the power of single cell genomics to generate a complete, high quality, non-composite reference genome within an environmental sample, which can be used for population genetic analyzes.

  16. Bacterial cell curvature through mechanical control of cell growth

    DEFF Research Database (Denmark)

    Cabeen, M.; Charbon, Godefroid; Vollmer, W.;

    2009-01-01

    The cytoskeleton is a key regulator of cell morphogenesis. Crescentin, a bacterial intermediate filament-like protein, is required for the curved shape of Caulobacter crescentus and localizes to the inner cell curvature. Here, we show that crescentin forms a single filamentous structure that coll...... cell wall insertion to produce curved growth. Our study suggests that bacteria may use the cytoskeleton for mechanical control of growth to alter morphology......The cytoskeleton is a key regulator of cell morphogenesis. Crescentin, a bacterial intermediate filament-like protein, is required for the curved shape of Caulobacter crescentus and localizes to the inner cell curvature. Here, we show that crescentin forms a single filamentous structure that...

  17. Septins regulate bacterial entry into host cells.

    Directory of Open Access Journals (Sweden)

    Serge Mostowy

    Full Text Available BACKGROUND: Septins are conserved GTPases that form filaments and are required in many organisms for several processes including cytokinesis. We previously identified SEPT9 associated with phagosomes containing latex beads coated with the Listeria surface protein InlB. METHODOLOGY/PRINCIPAL FINDINGS: Here, we investigated septin function during entry of invasive bacteria in non-phagocytic mammalian cells. We found that SEPT9, and its interacting partners SEPT2 and SEPT11, are recruited as collars next to actin at the site of entry of Listeria and Shigella. SEPT2-depletion by siRNA decreased bacterial invasion, suggesting that septins have roles during particle entry. Incubating cells with InlB-coated beads confirmed an essential role for SEPT2. Moreover, SEPT2-depletion impaired InlB-mediated stimulation of Met-dependent signaling as shown by FRET. CONCLUSIONS/SIGNIFICANCE: Together these findings highlight novel roles for SEPT2, and distinguish the roles of septin and actin in bacterial entry.

  18. Metabolic Responses of Bacterial Cells to Immobilization.

    Science.gov (United States)

    Żur, Joanna; Wojcieszyńska, Danuta; Guzik, Urszula

    2016-01-01

    In recent years immobilized cells have commonly been used for various biotechnological applications, e.g., antibiotic production, soil bioremediation, biodegradation and biotransformation of xenobiotics in wastewater treatment plants. Although the literature data on the physiological changes and behaviour of cells in the immobilized state remain fragmentary, it is well documented that in natural settings microorganisms are mainly found in association with surfaces, which results in biofilm formation. Biofilms are characterized by genetic and physiological heterogeneity and the occurrence of altered microenvironments within the matrix. Microbial cells in communities display a variety of metabolic differences as compared to their free-living counterparts. Immobilization of bacteria can occur either as a natural phenomenon or as an artificial process. The majority of changes observed in immobilized cells result from protection provided by the supports. Knowledge about the main physiological responses occurring in immobilized cells may contribute to improving the efficiency of immobilization techniques. This paper reviews the main metabolic changes exhibited by immobilized bacterial cells, including growth rate, biodegradation capabilities, biocatalytic efficiency and plasmid stability. PMID:27455220

  19. Metabolic Responses of Bacterial Cells to Immobilization

    Directory of Open Access Journals (Sweden)

    Joanna Żur

    2016-07-01

    Full Text Available In recent years immobilized cells have commonly been used for various biotechnological applications, e.g., antibiotic production, soil bioremediation, biodegradation and biotransformation of xenobiotics in wastewater treatment plants. Although the literature data on the physiological changes and behaviour of cells in the immobilized state remain fragmentary, it is well documented that in natural settings microorganisms are mainly found in association with surfaces, which results in biofilm formation. Biofilms are characterized by genetic and physiological heterogeneity and the occurrence of altered microenvironments within the matrix. Microbial cells in communities display a variety of metabolic differences as compared to their free-living counterparts. Immobilization of bacteria can occur either as a natural phenomenon or as an artificial process. The majority of changes observed in immobilized cells result from protection provided by the supports. Knowledge about the main physiological responses occurring in immobilized cells may contribute to improving the efficiency of immobilization techniques. This paper reviews the main metabolic changes exhibited by immobilized bacterial cells, including growth rate, biodegradation capabilities, biocatalytic efficiency and plasmid stability.

  20. Impact of trehalose and mycolate biosynthesis on the cell envelope of a Corynebacterium glutamicum L-lysine production strain

    OpenAIRE

    Gebhardt, Henrike

    2005-01-01

    In contrast to other Gram-positive bacteria all members of the suborder of Corynebacterineae, including Corynebacterium glutamicum, contain a cell envelope that comprises an outer lipid bilayer, the mycolate layer, which is considered as permeability barrier. Trehalose is an important component of the mycolate layer and involved in the biosynthesis of mycolate. The first step of mycolate biosynthesis, the condensation of trehalose monomycolate was proven to be located in the cell envelope. Th...

  1. Apoptosis of human intestinal epithelial cells after bacterial invasion.

    OpenAIRE

    Kim, J. M.; Eckmann, L; Savidge, T. C.; Lowe, D C; Witthöft, T; Kagnoff, M F

    1998-01-01

    Epithelial cells that line the human intestinal mucosa are the initial site of host invasion by bacterial pathogens. The studies herein define apoptosis as a new category of intestinal epithelial cell response to bacterial infection. Human colon epithelial cells are shown to undergo apoptosis following infection with invasive enteric pathogens, such as Salmonella or enteroinvasive Escherichia coli. In contrast to the rapid onset of apoptosis seen after bacterial infection of mouse monocyte-ma...

  2. Destructive effects of butyrate on the cell envelope of Helicobacter pylori.

    Science.gov (United States)

    Yonezawa, Hideo; Osaki, Takako; Hanawa, Tomoko; Kurata, Satoshi; Zaman, Cynthia; Woo, Timothy Derk Hoong; Takahashi, Motomichi; Matsubara, Sachie; Kawakami, Hayato; Ochiai, Kuniyasu; Kamiya, Shigeru

    2012-04-01

    Helicobacter pylori can be found in the oral cavity and is mostly detected by the use of PCR techniques. Growth of H. pylori is influenced by various factors in the mouth, such as the oral microflora, saliva and other antimicrobial substances, all of which make colonization of the oral cavity by H. pylori difficult. In the present study, we analysed the effect of the cell supernatant of a representative periodontal bacterium Porphyromonas gingivalis on H. pylori and found that the cell supernatant destroyed the H. pylori cell envelope. As P. gingivalis produces butyric acid, we focused our research on the effects of butyrate and found that it significantly inhibited the growth of H. pylori. H. pylori cytoplasmic proteins and DNA were detected in the extracellular environment after treatment with butyrate, suggesting that the integrity of the cell envelope was compromised and indicating that butyrate has a bactericidal effect on H. pylori. In addition, levels of extracellular H. pylori DNA increased following treatment with the cell supernatant of butyric acid-producing bacteria, indicating that the cell supernatant also has a bactericidal effect and that this may be due to its butyric acid content. In conclusion, butyric acid-producing bacteria may play a role in affecting H. pylori colonization of the oral cavity. PMID:22194341

  3. Inflammatory response of endothelial cells to hepatitis C virus recombinant envelope glycoprotein 2 protein exposure

    Directory of Open Access Journals (Sweden)

    Ana Carolina Urbaczek

    2014-09-01

    Full Text Available The hepatitis C virus (HCV encodes approximately 10 different structural and non-structural proteins, including the envelope glycoprotein 2 (E2. HCV proteins, especially the envelope proteins, bind to cell receptors and can damage tissues. Endothelial inflammation is the most important determinant of fibrosis progression and, consequently, cirrhosis. The aim of this study was to evaluate and compare the inflammatory response of endothelial cells to two recombinant forms of the HCV E2 protein produced in different expression systems (Escherichia coli and Pichia pastoris. We observed the induction of cell death and the production of nitric oxide, hydrogen peroxide, interleukin-8 and vascular endothelial growth factor A in human umbilical vein endothelial cells (HUVECs stimulated by the two recombinant E2 proteins. The E2-induced apoptosis of HUVECs was confirmed using the molecular marker PARP. The apoptosis rescue observed when the antioxidant N-acetylcysteine was used suggests that reactive oxygen species are involved in E2-induced apoptosis. We propose that these proteins are involved in the chronic inflammation caused by HCV.

  4. Characterization of the Pattern of αs1- and β-Casein Breakdown and Release of a Bioactive Peptide by a Cell Envelope Proteinase from Lactobacillus delbrueckii subsp. lactis CRL 581▿

    OpenAIRE

    Hebert, Elvira María; Mamone, Gianfranco; Picariello, Gianluca; Raya, Raúl R.; Savoy, Graciela; Ferranti, Pasquale; Addeo, Francesco

    2008-01-01

    The cell envelope-associated proteinases (CEPs) of the lactobacilli have key roles in bacterial nutrition and contribute to the development of the organoleptic properties of fermented milk products as well, as they can release bioactive health-beneficial peptides from milk proteins. The influence of the peptide supply, carbohydrate source, and osmolites on the CEP activity of the cheese starter Lactobacillus delbrueckii subsp. lactis CRL 581 was investigated. The CEP activity levels were cont...

  5. Duck tembusu virus and its envelope protein induce programmed cell death.

    Science.gov (United States)

    Shaozhou, Wulin; Li, Chenxi; Zhang, Qingshan; Meng, Runzhe; Gao, Youlan; Liu, Hongyu; Bai, Xiaofei; Chen, Yuhuan; Liu, Ming; Liu, Siguo; Zhang, Yun

    2015-08-01

    The cytopathic effect produced in cells infected with duck tembusu virus (DTMUV) suggests that this emerging virus may induce apoptosis in primary cultures of duck embryo fibroblasts (DEF). Here, we present evidence that DTMUV infection of cultured cells activates apoptosis and that the ability of DTMUV to induce apoptosis is not restricted to cell type because DTMUV-induced apoptosis in duck and mammalian host cells. We further investigated which viral components induce apoptosis in DTMUV-infected host cells. The major envelope glycoprotein (E) was investigated for its apoptotic activities in expressed cells. Transient expression of the E protein alone triggered apoptosis in DEF, Vero, and BHK cells. Expression of the E protein resulted in activation of caspase-3-like proteases in cultured cells. These results indicate that infection of cells with DTMUV or expression of DTMUV E protein alone induces apoptosis, providing the basis for future to define the molecules that play key roles in the fate of DTMUV-infected cells. PMID:26056013

  6. Participation of ezrin in bacterial uptake by trophoblast giant cells

    Directory of Open Access Journals (Sweden)

    Kim Suk

    2009-09-01

    Full Text Available Abstract Background Trophoblast giant (TG cells are involved in systematic removal of bacterial pathogens from the maternal-fetal interface of the placenta. In particular, TG cells have the ability to take up extracellular antigens by active phagocytosis induced by interferon-gamma (IFN-gamma. We previously reported that heat shock cognate protein 70 (Hsc70 present on the surface of TG cells mediated the uptake of Brucella abortus. However, the mechanism of bacterial uptake by TG cells is not completely understood. Here we identified ezrin, a member of ezrin-radixin-moesin (ERM protein family, as a molecule associated with Hsc70. Methods Mouse TG cells were employed in all experiments, and B. abortus was used as the bacterial antigen. Confirmation of the binding capacity of ERM protein was assessed by pull-down assay and ELISA using recombinant Hsc70 and ERM proteins. Ezrin was depleted using siRNA and the depletion examined by immunoblotting or immunofluorescence staining. Results The expression level of ezrin was higher in TG cells than in trophoblast stem (TS cells, and ezrin knockdown TG cells showed a reduction in bacterial uptake ability. Although tyrosine phosphorylation of ezrin was not related to bacterial uptake activity, localization of Hsc70 on the membrane was affected by the depletion of ezrin in TG cells. Conclusion Ezrin associates with Hsc70 that locates on the membrane of TG cells and participates in the bacterial uptake by TG cells.

  7. Release of the cell-envelope-associated proteinase of Lactobacillus delbrueckii subspecies lactis CRL 581 is dependent upon pH and temperature.

    Science.gov (United States)

    Espeche Turbay, María B; Savoy de Giori, Graciela; Hebert, Elvira M

    2009-09-23

    The cell-envelope-associated proteinase of Lactobacillus delbrueckii subsp. lactis CRL 581 (PrtL) has an essential role in bacterial growth and contributes to the development of the organoleptic properties of hard cheeses and to the release of bioactive health-beneficial peptides from milk proteins. In this study, the effect of environmental pH on PrtL production by L. delbrueckii subsp. lactis CRL 581 in a chemically defined medium and the influence of pH, temperature, and Ca(2+) ions on PrtL activity, stability, and release from the cell envelope were analyzed. The maximum PrtL activity levels were observed in the middle of the exponential growth phase, with the values at constant pH of 5.5 and 6.0 being higher than those observed at pH 4.5 and 5.0. At pH 4.5, PrtL remained mainly associated with the cell envelope, whereas at pH values of 5.5 or higher, approximately 40% of PrtL was found in the medium. In addition, the PrtL activity was stable for 24 h at 4 and 25 degrees C, and its release at 4, 25, and 40 degrees C was time-dependent. PrtL activity, stability, and release were independent of the presence of Ca(2+) ions in the medium. These results indicated that, at pH and temperature conditions found during the manufacture of hard cheeses, PrtL would remain active either bound to the cell or released in the supernatant contributing to the organoleptic characteristics and beneficial health effects of the fermented milk products. PMID:19754175

  8. Quantitative proteomics of the Neisseria gonorrhoeae cell envelope and membrane vesicles for the discovery of potential therapeutic targets.

    Science.gov (United States)

    Zielke, Ryszard A; Wierzbicki, Igor H; Weber, Jacob V; Gafken, Philip R; Sikora, Aleksandra E

    2014-05-01

    Neisseria gonorrhoeae (GC) is a human-specific pathogen, and the agent of a sexually transmitted disease, gonorrhea. There is a critical need for new approaches to study and treat GC infections because of the growing threat of multidrug-resistant isolates and the lack of a vaccine. Despite the implied role of the GC cell envelope and membrane vesicles in colonization and infection of human tissues and cell lines, comprehensive studies have not been undertaken to elucidate their constituents. Accordingly, in pursuit of novel molecular therapeutic targets, we have applied isobaric tagging for absolute quantification coupled with liquid chromatography and mass spectrometry for proteome quantitative analyses. Mining the proteome of cell envelopes and native membrane vesicles revealed 533 and 168 common proteins, respectively, in analyzed GC strains FA1090, F62, MS11, and 1291. A total of 22 differentially abundant proteins were discovered including previously unknown proteins. Among those proteins that displayed similar abundance in four GC strains, 34 were found in both cell envelopes and membrane vesicles fractions. Focusing on one of them, a homolog of an outer membrane protein LptD, we demonstrated that its depletion caused loss of GC viability. In addition, we selected for initial characterization six predicted outer membrane proteins with unknown function, which were identified as ubiquitous in the cell envelopes derived from examined GC isolates. These studies entitled a construction of deletion mutants and analyses of their resistance to different chemical probes. Loss of NGO1985, in particular, resulted in dramatically decreased GC viability upon treatment with detergents, polymyxin B, and chloramphenicol, suggesting that this protein functions in the maintenance of the cell envelope permeability barrier. Together, these findings underscore the concept that the cell envelope and membrane vesicles contain crucial, yet under-explored determinants of GC

  9. Reprogramming of somatic cells induced by fusion of embryonic stem cells using hemagglutinating virus of Japan envelope (HVJ-E)

    International Nuclear Information System (INIS)

    In this research, hemagglutinating virus of Japan envelope (HVJ-E) was used to reprogram somatic cells by fusion with mouse embryonic stem (ES) cells. Neomycin-resistant mouse embryonic fibroblasts (MEFs) were used as somatic cells. Nanog-overexpressing puromycin-resistant EB3 cells were used as mouse ES cells. These two cells were fused by exposing to HVJ-E and the generated fusion cells were selected by puromycin and G418 to get the stable fusion cell line. The fusion cells form colonies in feeder-free culture system. Microsatellite analysis of the fusion cells showed that they possessed genes from both ES cells and fibroblasts. The fusion cells were tetraploid, had alkali phosphatase activity, and expressed stem cell marker genes such as Pou5f1, Nanog, and Sox2, but not the fibroblast cell marker genes such as Col1a1 and Col1a2. The pluripotency of fusion cells was confirmed by their expression of marker genes for all the three germ layers after differentiation induction, and by their ability to form teratoma which contained all the three primary layers. Our results show that HVJ-E can be used as a fusion reagent for reprogramming of somatic cells.

  10. Expanded breadth of the T-cell response to mosaic HIV-1 envelope DNA vaccination

    Energy Technology Data Exchange (ETDEWEB)

    Korber, Bette [Los Alamos National Laboratory; Fischer, William [Los Alamos National Laboratory; Wallstrom, Timothy [Los Alamos National Laboratory

    2009-01-01

    An effective AIDS vaccine must control highly diverse circulating strains of HIV-1. Among HIV -I gene products, the envelope (Env) protein contains variable as well as conserved regions. In this report, an informatic approach to the design of T-cell vaccines directed to HIV -I Env M group global sequences was tested. Synthetic Env antigens were designed to express mosaics that maximize the inclusion of common potential Tcell epitope (PTE) 9-mers and minimize the inclusion of rare epitopes likely to elicit strain-specific responses. DNA vaccines were evaluated using intracellular cytokine staining (ICS) in inbred mice with a standardized panel of highly conserved 15-mer PTE peptides. I, 2 and 3 mosaic sets were developed that increased theoretical epitope coverage. The breadth and magnitude ofT-cell immunity stimulated by these vaccines were compared to natural strain Env's; additional comparisons were performed on mutant Env's, including gpl60 or gpl45 with or without V regions and gp41 deletions. Among them, the 2 or 3 mosaic Env sets elicited the optimal CD4 and CD8 responses. These responses were most evident in CD8 T cells; the 3 mosaic set elicited responses to an average of 8 peptide pools compared to 2 pools for a set of3 natural Env's. Synthetic mosaic HIV -I antigens can therefore induce T-cell responses with expanded breadth and may facilitate the development of effective T -cell-based HIV -1 vaccines.

  11. Osmotic Pressure, Bacterial Cell Walls, and Penicillin: A Demonstration.

    Science.gov (United States)

    Lennox, John E.

    1984-01-01

    An easily constructed apparatus that models the effect of penicillin on the structure of bacterial cells is described. Background information and procedures for using the apparatus during a classroom demonstration are included. (JN)

  12. Quantification of bacterial invasion into adherent cells by flow cytometry

    OpenAIRE

    Pils, Stefan; Schmitter, Tim; Neske, Florian; Hauck, Christof R.

    2006-01-01

    Quantification of invasive, intracellular bacteria is critical in many areas of cellular microbiology and immunology. We describe a novel and fast approach to determine invasion of bacterial pathogens in adherent cell types such as epithelial cells or fibroblasts based on flow cytometry. Using the CEACAM-mediated uptake of Opa-expressing Neisseria gonorrhoeae as a well-characterized model of bacterial invasion, we demonstrate that the flow cytometry-based method yields results comparable to a...

  13. RECOVERY OF POLYHYDROXYALKANOATES (PHAs) FROM BACTERIAL CELLS USING ENZYMATIC PROCESS

    OpenAIRE

    S. Marsudi

    2012-01-01

    Polyhydroxyalkanoates (PHAs) are intracellular material accumulated by several bacteria. Commercial production of PHAs faces the issue of high production cost especially substrate cost and recovery/separation cost. An alternative to reduce the production cost is to use enzyme and or chemical to recover PHAs from bacterial cells. Recovery of PHAs from bacterial cells was done using enzyme, chemical, and a mixture of enzyme and chemical. Enzyme (s) and or chemical(s) were added into culture bro...

  14. Bacterial surface appendages strongly impact nanomechanical and electrokinetic properties of Escherichia coli cells subjected to osmotic stress.

    Directory of Open Access Journals (Sweden)

    Grégory Francius

    Full Text Available The physicochemical properties and dynamics of bacterial envelope, play a major role in bacterial activity. In this study, the morphological, nanomechanical and electrohydrodynamic properties of Escherichia coli K-12 mutant cells were thoroughly investigated as a function of bulk medium ionic strength using atomic force microscopy (AFM and electrokinetics (electrophoresis. Bacteria were differing according to genetic alterations controlling the production of different surface appendages (short and rigid Ag43 adhesins, longer and more flexible type 1 fimbriae and F pilus. From the analysis of the spatially resolved force curves, it is shown that cells elasticity and turgor pressure are not only depending on bulk salt concentration but also on the presence/absence and nature of surface appendage. In 1 mM KNO(3, cells without appendages or cells surrounded by Ag43 exhibit large Young moduli and turgor pressures (∼700-900 kPa and ∼100-300 kPa respectively. Under similar ionic strength condition, a dramatic ∼50% to ∼70% decrease of these nanomechanical parameters was evidenced for cells with appendages. Qualitatively, such dependence of nanomechanical behavior on surface organization remains when increasing medium salt content to 100 mM, even though, quantitatively, differences are marked to a much smaller extent. Additionally, for a given surface appendage, the magnitude of the nanomechanical parameters decreases significantly when increasing bulk salt concentration. This effect is ascribed to a bacterial exoosmotic water loss resulting in a combined contraction of bacterial cytoplasm together with an electrostatically-driven shrinkage of the surface appendages. The former process is demonstrated upon AFM analysis, while the latter, inaccessible upon AFM imaging, is inferred from electrophoretic data interpreted according to advanced soft particle electrokinetic theory. Altogether, AFM and electrokinetic results clearly demonstrate the

  15. Critical role of a ferritin-like protein in the control of Listeria monocytogenes cell envelope structure and stability under β-lactam pressure.

    Directory of Open Access Journals (Sweden)

    Agata Krawczyk-Balska

    Full Text Available The human pathogen Listeria monocytogenes is susceptible to the β-lactam antibiotics penicillin G and ampicillin, and these are the drugs of choice for the treatment of listerial infections. However, these antibiotics exert only a bacteriostatic effect on this bacterium and consequently, L. monocytogenes is regarded as β-lactam tolerant. It is widely accepted that the phenomenon of bacterial tolerance to β-lactams is due to the lack of adequate autolysin activity, but the mechanisms of L. monocytogenes tolerance to this class of antibiotics are poorly characterized. A ferritin-like protein (Fri was recently identified as a mediator of β-lactam tolerance in L. monocytogenes, but its function in this process remains unknown. The present study was undertaken to improve our understanding of L. monocytogenes tolerance to β-lactams and to characterize the role of Fri in this phenomenon. A comparative physiological analysis of wild-type L. monocytogenes and a fri deletion mutant provided evidence of a multilevel mechanism controlling autolysin activity in cells grown under β-lactam pressure, which leads to a reduction in the level and/or activity of cell wall-associated autolysins. This is accompanied by increases in the amount of teichoic acids, cell wall thickness and cell envelope integrity of L. monocytogenes grown in the presence of penicillin G, and provides the basis for the innate β-lactam tolerance of this bacterium. Furthermore, this study revealed the inability of the L. monocytogenes Δ fri mutant to deplete autolysins from the cell wall, to adjust the content of teichoic acids and to maintain their D-alanylation at the correct level when treated with penicillin G, thus providing further evidence that Fri is involved in the control of L. monocytogenes cell envelope structure and stability under β-lactam pressure.

  16. MEASURING THE OPERATING EFFICIENCY OF SOLAR CELL COMPANIES IN TAIWAN WITH DATA ENVELOPMENT ANALYSIS

    Directory of Open Access Journals (Sweden)

    Jui-Min Hsiao

    2012-01-01

    Full Text Available This study discusses the Data Envelopment Analysis (DEA approach to measure the operating efficiency of solar cell companies. With active and constructive advancement of the government, competition in the solar cell industry became even tenser. When drawing up the competitive strategies, one firm should identify the key indicators of its operating efficiency. This study incorporates three inputs (the number of employees, fixed asset, operating expenses and three outputs (fund and investment, shareholders equity, sales revenue, gross margin to measure the operating efficiency of 12 DMUs (solar cell companies to provide reference for these companies in determining competitive strategies. Data are gathered from the market observation post system in Taiwan. The result indicates that there are six efficient DMUs and six inefficient DMUs. For the relatively inefficient DMUs, a slack variable analysis is performed and the efficiency scores to understand the usage of inputs and performance improvement of inefficient DMUs. The findings could benefit company operators seeking performance improvement in which they could benchmark practices being adapted by the most efficient companies. Together with Malmquist productivity index analysis, companies are able to assess the productivity change of DMUs over time. Finally, managerial implications are provided.

  17. Expression of the human immunodeficiency virus envelope glycoprotein is restricted to basolateral surfaces of polarized epithelial cells

    International Nuclear Information System (INIS)

    Polarized epithelial cells exhibit apical (lumenal) and basolateral (serosal) membrane domains that are separated by circumferential tight junctions. In such cells, enveloped viruses that mature by budding at cell surfaces are released at particular membrane domains. The authors have used a vaccinia virus recombinant to investigate the site of surface expression of the human immunodeficiency virus type 1 envelope glycoprotein in Madin-Darby canine kidney cells. Cells were infected with the vaccinia virus recombinant, and surface expression of the glycoprotein was analyzed by indirect immunofluorescence, 125I-protein A binding, and immunoelectron microscopy. The glycoprotein appeared exclusively at the basolateral surface as early as 2 h postinfection and reached a maximum level at 8 h postinfection. The gp120 glycoprotein was found to be secreted efficiently into culture medium, and this secretion occurred exclusively at the basolateral surface

  18. System-level design of bacterial cell cycle control

    OpenAIRE

    McAdams, Harley H.; Shapiro, Lucy

    2009-01-01

    Understanding of the cell cycle control logic in Caulobacter has progressed to the point where we now have an integrated view of the operation of an entire bacterial cell cycle system functioning as a state machine. Oscillating levels of a few temporally-controlled master regulator proteins in a cyclical circuit drive cell cycle progression. To a striking degree, the cell cycle regulation is a whole cell phenomenon. Phospho-signaling proteins and proteases dynamically deployed to specific loc...

  19. Ultrastructural Study of Salmonella typhimurium Treated with Membrane-Active Agents: Specific Reaction of Dansylchloride with Cell Envelope Components

    Science.gov (United States)

    Schindler, Peter R. G.; Teuber, Michael

    1978-01-01

    Amino groups of cell envelope proteins, lipids, and lipopolysaccharides cannot be labeled in intact cells of Salmonella typhimurium G 30 by using 5-dimethylaminonaphthalene-1-sulfonylchloride incorporated in lecithin-cholesterol vesicles. However, application of membrane-interacting agents like tris(hydroxymethyl)aminomethane (Tris)-hydrochloride, ethylenediaminetetraacetate (Na salt) (EDTA), divalent cations, and sublethal doses of the cationic antibacterial agents polymyxin B and chlorhexidine induced specific fluorescent labeling of envelope proteins and lipids but not of cytoplasmic compounds, with the exception of a soluble protein with a molecular weight of 46,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Treatment with Tris-hydrochloride buffer produced labeling of the heat-modifiable protein B/B+ and of proteins with molecular weights of 26,000, 22,000, and below 17,000. A combination of Tris-hydrochloride and EDTA induced additional dansylation of the major protein A and of proteins of molecular weights 80,000, 60,000, and 44,000. Polymyxin B and chlorhexidine caused similar labeling patterns. In every case, except with divalent cation treatment, protein B/B+ was the most prominently labeled species. Phosphatidylethanolamine was dansylated up to 30%. Lipopolysaccharide was not reactive under any condition or treatment. In addition, the peptidoglycan-bound lipoprotein did not react with dansylchloride in either intact or Tris-hydrochloride-treated cells. The results are discussed with regard to a possible localization of labeled and unlabeled compounds of the cell envelope on the basis of a model placing cell envelope amino groups into ion-ion interactions with anionic components of other envelope compounds like phosphate and carboxyl groups. Images PMID:97268

  20. Conductivity and Dielectric Dispersion of Gram-Positive Bacterial Cells

    Science.gov (United States)

    van der Wal A; Minor; Norde; Zehnder; Lyklema

    1997-02-01

    The conductivity of bacterial cell suspensions has been studied over a wide range of ionic strengths and is interpreted in terms of their cell wall properties. The experimental data have been analyzed after improving the high kappaa double-layer theory of Fixman, by accounting for ionic mobility in the hydrodynamically stagnant layer, i.e., in the bacterial wall. Static conductivity and dielectric dispersion measurements both show that the counterions in the porous gel-like cell wall give rise to a considerable surface conductance. From a comparison of the mobile charge with the total cell wall charge it is inferred that the mobilities of the ions in the bacterial wall are of the same order but somewhat lower than those in the bulk electrolyte solution. The occurrence of surface conductance reduces the electrophoretic mobility in electrophoresis studies. If this effect is not taken into account, the zeta-potential will be underestimated, especially at low electrolyte concentrations. PMID:9056304

  1. RECOVERY OF POLYHYDROXYALKANOATES (PHAs FROM BACTERIAL CELLS USING ENZYMATIC PROCESS

    Directory of Open Access Journals (Sweden)

    S. Marsudi

    2012-02-01

    Full Text Available Polyhydroxyalkanoates (PHAs are intracellular material accumulated by several bacteria. Commercial production of PHAs faces the issue of high production cost especially substrate cost and recovery/separation cost. An alternative to reduce the production cost is to use enzyme and or chemical to recover PHAs from bacterial cells. Recovery of PHAs from bacterial cells was done using enzyme, chemical, and a mixture of enzyme and chemical. Enzyme (s and or chemical(s were added into culture broth to disrupt cells after adjusting pH and temperature of the culture broth. Treatment by adding enzyme or chemical only into culture broth showed a low level of PHAs recovered from bacterial cells. Treatment by adding a mixture of enzymes and chemicals showed the best result among 22 examined combinations, i.e. a mixture of EDTA, lisozyme, papain enzyme, and SDS. This combination gave a PHA recovery of 65 % w/w.

  2. [Cashmere goat bacterial artificial chromosome recombination and cell transfection system].

    Science.gov (United States)

    Huang, Tian; Cao, Zhongyang; Yang, Yaohui; Cao, Gengsheng

    2016-03-01

    The Cashmere goat is mainly used to produce cashmere, which is very popular for its delicate fiber, luscious softness and natural excellent warm property. Keratin associated protein (KAP) and bone morphogenetic protein (BMP) of the Cashmere goat play an important role in the proliferation and development of cashmere fiber follicle cells. Bacterial artificial chromosome containing kap6.3, kap8.1 and bmp4 genes were used to increase the production and quality of Cashmere. First, we constructed bacterial artificial chromosomes by homology recombination. Then Tol2 transposon was inserted into bacterial artificial chromosomes that were then transfected into Cashmere goat fibroblasts by Amaxa Nucleofector technology according to the manufacture's instructions. We successfully constructed the BAC-Tol2 vectors containing target genes. Each vector contained egfp report gene with UBC promoter, Neomycin resistant gene for cell screening and two loxp elements for resistance removing after transfected into cells. The bacterial artificial chromosome-Tol2 vectors showed a high efficiency of transfection that can reach 1% to 6% with a highest efficiency of 10%. We also obtained Cashmere goat fibroblasts integrated exogenous genes (kap6.3, kap8.1 and bmp4) preparing for the clone of Cashmere goat in the future. Our research demonstrates that the insertion of Tol2 transposons into bacterial artificial chromosomes improves the transfection efficiency and accuracy of bacterial artificial chromosome error-free recombination. PMID:27349114

  3. The HP0256 gene product is involved in motility and cell envelope architecture of Helicobacter pylori

    LENUS (Irish Health Repository)

    Douillard, Francois P

    2010-04-08

    Abstract Background Helicobacter pylori is the causative agent for gastritis, and peptic and duodenal ulcers. The bacterium displays 5-6 polar sheathed flagella that are essential for colonisation and persistence in the gastric mucosa. The biochemistry and genetics of flagellar biogenesis in H. pylori has not been fully elucidated. Bioinformatics analysis suggested that the gene HP0256, annotated as hypothetical, was a FliJ homologue. In Salmonella, FliJ is a chaperone escort protein for FlgN and FliT, two proteins that themselves display chaperone activity for components of the hook, the rod and the filament. Results Ablation of the HP0256 gene in H. pylori significantly reduced motility. However, flagellin and hook protein synthesis was not affected in the HP0256 mutant. Transmission electron transmission microscopy revealed that the HP0256 mutant cells displayed a normal flagellum configuration, suggesting that HP0256 was not essential for assembly and polar localisation of the flagella in the cell. Interestingly, whole genome microarrays of an HP0256 mutant revealed transcriptional changes in a number of genes associated with the flagellar regulon and the cell envelope, such as outer membrane proteins and adhesins. Consistent with the array data, lack of the HP0256 gene significantly reduced adhesion and the inflammatory response in host cells. Conclusions We conclude that HP0256 is not a functional counterpart of FliJ in H. pylori. However, it is required for full motility and it is involved, possibly indirectly, in expression of outer membrane proteins and adhesins involved in pathogenesis and adhesion.

  4. Human monoclonal antibody directed against an envelope glycoprotein of human T-cell leukemia virus type I.

    OpenAIRE

    Matsushita, S; Robert-Guroff, M; Trepel, J. (Jane); Cossman, J; Mitsuya, H; Broder, S

    1986-01-01

    We report the production and characterization of a human monoclonal antibody reactive against the major envelope glycoprotein of human T-cell leukemia virus type I (HTLV-I), a virus linked to the etiology of adult T-cell leukemia. We exposed lymph-node cells derived from a patient with adult T-cell leukemia to the Epstein-Barr virus in vitro and obtained a B-cell clone (designated 0.5 alpha) by a limiting dilution technique. The secreted product of 0.5 alpha is a monoclonal antibody (also des...

  5. Electroporation of Functional Bacterial Effectors into Mammalian Cells

    Energy Technology Data Exchange (ETDEWEB)

    Sontag, Ryan L.; Mihai, Cosmin; Orr, Galya; Savchenko, Alexei; Skarina, Tatiana; Cui, Hong; Cort, John R.; Adkins, Joshua N.; Brown, Roslyn N.

    2015-01-01

    Electroporation was used to insert purified bacterial virulence effector proteins directly into living eukaryotic cells. Protein localization was monitored by confocal immunofluorescence microscopy. This method allows for studies on trafficking, function, and protein-protein interactions using active exogenous proteins, avoiding the need for heterologous expression in eukaryotic cells.

  6. Bacterial-induced cell reprogramming to stem cell-like cells: new premise in host-pathogen interactions

    OpenAIRE

    Hess, Samuel; Rambukkana, Anura

    2014-01-01

    Bacterial pathogens employ a myriad of strategies to alter host tissue cell functions for bacterial advantage during infection. Recent advances revealed a fusion of infection biology with stem cell biology by demonstrating developmental reprogramming of lineage committed host glial cells to progenitor/stem cell-like cells by an intracellular bacterial pathogen Mycobacterium leprae. Acquisition of migratory and immunomodulatory properties of such reprogrammed cells provides an added advantage ...

  7. Nuclear envelope proteins modulate proliferation of vascular smooth muscle cells during cyclic stretch application

    Science.gov (United States)

    Qi, Ying-Xin; Yao, Qing-Ping; Huang, Kai; Shi, Qian; Zhang, Ping; Wang, Guo-Liang; Han, Yue; Bao, Han; Wang, Lu; Li, Hai-Peng; Shen, Bao-Rong; Wang, Yingxiao; Chien, Shu; Jiang, Zong-Lai

    2016-01-01

    Cyclic stretch is an important inducer of vascular smooth muscle cell (VSMC) proliferation, which is crucial in vascular remodeling during hypertension. However, the molecular mechanism remains unclear. We studied the effects of emerin and lamin A/C, two important nuclear envelope proteins, on VSMC proliferation in hypertension and the underlying mechano-mechanisms. In common carotid artery of hypertensive rats in vivo and in cultured cells subjected to high (15%) cyclic stretch in vitro, VSMC proliferation was increased significantly, and the expression of emerin and lamin A/C was repressed compared with normotensive or normal (5%) cyclic stretch controls. Using targeted siRNA to mimic the repressed expression of emerin or lamin A/C induced by 15% stretch, we found that VSMC proliferation was enhanced under static and 5%-stretch conditions. Overexpression of emerin or lamin A/C reversed VSMC proliferation induced by 15% stretch. Hence, emerin and lamin A/C play critical roles in suppressing VSMC hyperproliferation induced by hyperstretch. ChIP-on-chip and MOTIF analyses showed that the DNAs binding with emerin contain three transcription factor motifs: CCNGGA, CCMGCC, and ABTTCCG; DNAs binding with lamin A/C contain the motifs CVGGAA, GCCGCYGC, and DAAGAAA. Protein/DNA array proved that altered emerin or lamin A/C expression modulated the activation of various transcription factors. Furthermore, accelerating local expression of emerin or lamin A/C reversed cell proliferation in the carotid artery of hypertensive rats in vivo. Our findings establish the pathogenetic role of emerin and lamin A/C repression in stretch-induced VSMC proliferation and suggest mechanobiological mechanism underlying this process that involves the sequence-specific binding of emerin and lamin A/C to specific transcription factor motifs. PMID:27114541

  8. Proteomic analysis of Brucella abortus cell envelope and identification of immunogenic candidate proteins for vaccine development.

    Science.gov (United States)

    Connolly, Joseph P; Comerci, Diego; Alefantis, Timothy G; Walz, Alexander; Quan, Marian; Chafin, Ryan; Grewal, Paul; Mujer, Cesar V; Ugalde, Rodolfo A; DelVecchio, Vito G

    2006-07-01

    Brucella abortus is the etiologic agent of bovine brucellosis and causes a chronic disease in humans known as undulant fever. In livestock the disease is characterized by abortion and sterility. Live, attenuated vaccines such as S19 and RB51 have been used to control the spread of the disease in animals; however, they are considered unsafe for human use and they induce abortion in pregnant cattle. For the development of a safer and equally efficacious vaccine, immunoproteomics was utilized to identify novel candidate proteins from B. abortus cell envelope (CE). A total of 163 proteins were identified using 2-DE with MALDI-TOF MS and LC-MS/MS. Some of the major protein components include outer-membrane protein (OMP) 25, OMP31, Omp2b porin, and 60 kDa chaperonin GroEL. 2-DE Western blot analyses probed with antiserum from bovine and a human patient infected with Brucella identified several new immunogenic proteins such as fumarate reductase flavoprotein subunit, F0F1-type ATP synthase alpha subunit, and cysteine synthase A. The elucidation of the immunome of B. abortus CE identified a number of candidate proteins for developing vaccines against Brucella infection in bovine and humans. PMID:16739129

  9. Micro-magnet arrays for specific single bacterial cell positioning

    Energy Technology Data Exchange (ETDEWEB)

    Pivetal, Jérémy, E-mail: jeremy.piv@netcmail.com [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France); Royet, David [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France); Ciuta, Georgeta [Univ. Grenoble Alpes, Inst NEEL, F-38042 Grenoble (France); CNRS, Inst NEEL, F-38042 Grenoble (France); Frenea-Robin, Marie [Université de Lyon, Université Lyon 1, CNRS UMR 5005, Laboratoire Ampère, F-69622 Villeurbanne (France); Haddour, Naoufel [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France); Dempsey, Nora M. [Univ. Grenoble Alpes, Inst NEEL, F-38042 Grenoble (France); CNRS, Inst NEEL, F-38042 Grenoble (France); Dumas-Bouchiat, Frédéric [Univ Limoges, CNRS, SPCTS UMR 7513, 12 Rue Atlantis, F-87068 Limoges (France); Simonet, Pascal [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France)

    2015-04-15

    In various contexts such as pathogen detection or analysis of microbial diversity where cellular heterogeneity must be taken into account, there is a growing need for tools and methods that enable microbiologists to analyze bacterial cells individually. One of the main challenges in the development of new platforms for single cell studies is to perform precise cell positioning, but the ability to specifically target cells is also important in many applications. In this work, we report the development of new strategies to selectively trap single bacterial cells upon large arrays, based on the use of micro-magnets. Escherichia coli bacteria were used to demonstrate magnetically driven bacterial cell organization. In order to provide a flexible approach adaptable to several applications in the field of microbiology, cells were magnetically and specifically labeled using two different strategies, namely immunomagnetic labeling and magnetic in situ hybridization. Results show that centimeter-sized arrays of targeted, isolated bacteria can be successfully created upon the surface of a flat magnetically patterned hard magnetic film. Efforts are now being directed towards the integration of a detection tool to provide a complete micro-system device for a variety of microbiological applications. - Highlights: 1.We report a new approach to selectively micropattern bacterial cells individually upon micro-magnet arrays. 2.Permanent micro-magnets of a size approaching that of bacteria could be fabricated using a Thermo-Magnetic Patterning process. 3.Bacterial cells were labeled using two different magnetic labeling strategies providing flexible approach adaptable to several applications in the field of microbiology.

  10. Micro-magnet arrays for specific single bacterial cell positioning

    International Nuclear Information System (INIS)

    In various contexts such as pathogen detection or analysis of microbial diversity where cellular heterogeneity must be taken into account, there is a growing need for tools and methods that enable microbiologists to analyze bacterial cells individually. One of the main challenges in the development of new platforms for single cell studies is to perform precise cell positioning, but the ability to specifically target cells is also important in many applications. In this work, we report the development of new strategies to selectively trap single bacterial cells upon large arrays, based on the use of micro-magnets. Escherichia coli bacteria were used to demonstrate magnetically driven bacterial cell organization. In order to provide a flexible approach adaptable to several applications in the field of microbiology, cells were magnetically and specifically labeled using two different strategies, namely immunomagnetic labeling and magnetic in situ hybridization. Results show that centimeter-sized arrays of targeted, isolated bacteria can be successfully created upon the surface of a flat magnetically patterned hard magnetic film. Efforts are now being directed towards the integration of a detection tool to provide a complete micro-system device for a variety of microbiological applications. - Highlights: 1.We report a new approach to selectively micropattern bacterial cells individually upon micro-magnet arrays. 2.Permanent micro-magnets of a size approaching that of bacteria could be fabricated using a Thermo-Magnetic Patterning process. 3.Bacterial cells were labeled using two different magnetic labeling strategies providing flexible approach adaptable to several applications in the field of microbiology

  11. Single Cell Analysis of a Bacterial Sender-Receiver System.

    Science.gov (United States)

    Ramalho, Tiago; Meyer, Andrea; Mückl, Andrea; Kapsner, Korbinian; Gerland, Ulrich; Simmel, Friedrich C

    2016-01-01

    Monitoring gene expression dynamics on the single cell level provides important information on cellular heterogeneity and stochasticity, and potentially allows for more accurate quantitation of gene expression processes. We here study bacterial senders and receivers genetically engineered with components of the quorum sensing system derived from Aliivibrio fischeri on the single cell level using microfluidics-based bacterial chemostats and fluorescence video microscopy. We track large numbers of bacteria over extended periods of time, which allows us to determine bacterial lineages and filter out subpopulations within a heterogeneous population. We quantitatively determine the dynamic gene expression response of receiver bacteria to varying amounts of the quorum sensing inducer N-3-oxo-C6-homoserine lactone (AHL). From this we construct AHL response curves and characterize gene expression dynamics of whole bacterial populations by investigating the statistical distribution of gene expression activity over time. The bacteria are found to display heterogeneous induction behavior within the population. We therefore also characterize gene expression in a homogeneous bacterial subpopulation by focusing on single cell trajectories derived only from bacteria with similar induction behavior. The response at the single cell level is found to be more cooperative than that obtained for the heterogeneous total population. For the analysis of systems containing both AHL senders and receiver cells, we utilize the receiver cells as 'bacterial sensors' for AHL. Based on a simple gene expression model and the response curves obtained in receiver-only experiments, the effective AHL concentration established by the senders and their 'sending power' is determined. PMID:26808777

  12. Cell fusion by the envelope glycoproteins of persistent measles viruses which caused lethal human brain disease.

    OpenAIRE

    Cattaneo, R.; Rose, J. K.

    1993-01-01

    Measles virus (MV) rarely induces lethal diseases of the human central nervous system characterized by reduced expression of the viral envelope proteins and by lack of viral budding. The MV envelope contains two integral membrane proteins, termed fusion (F) protein and hemagglutinin (H) protein, and a membrane-associated matrix (M) protein. Previously, analysis of MV genes from autopsy material indicated that the M protein and the F protein intracellular domain are often drastically altered b...

  13. Myeloid-Derived Suppressor Cells in Bacterial Infections

    Science.gov (United States)

    Ost, Michael; Singh, Anurag; Peschel, Andreas; Mehling, Roman; Rieber, Nikolaus; Hartl, Dominik

    2016-01-01

    Myeloid-derived suppressor cells (MDSCs) comprise monocytic and granulocytic innate immune cells with the capability of suppressing T- and NK-cell responses. While the role of MDSCs has been studied in depth in malignant diseases, the understanding of their regulation and function in infectious disease conditions has just begun to evolve. Here we summarize and discuss the current view how MDSCs participate in bacterial infections and how this knowledge could be exploited for potential future therapeutics. PMID:27066459

  14. 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 divi

  15. Aerotactic Cell Density Variations in Bacterial Turbulence

    Science.gov (United States)

    Fernandez, Vicente; Smriga, Steven; Menolascina, Filippo; Rusconi, Roberto; Stocker, Roman

    2015-11-01

    Concentrated suspensions of motile bacteria such as Bacillus subtilis exhibit group dynamics much larger than the scale of an individual bacterium, visual similar to high Reynolds number turbulence. These suspensions represent a microscale realization of active matter. Individually, B. subtilis are also aerotactic, and will accumulate near oxygen sources. Using a microfluidic device for generating oxygen gradients, we investigate the relationship between individuals' attraction to oxygen and the collective motion resultant from hydrodynamic interactions. We focus on changes in density revealed by a fluorescently labeled sub-population of B. subtilis in the dense suspension. This approach allows us to examine changes in density during the onset of collective motion as well as fully developed bacterial turbulence.

  16. Infection with the oncogenic human papillomavirus type 59 alters protein components of the cornified cell envelope

    International Nuclear Information System (INIS)

    Infection of the genital tract with human papillomaviruses (HPVs) leads to proliferative and dysplastic epithelial lesions. The mechanisms used by the virus to escape the infected keratinocyte are not well understood. Infection of keratinocytes with HPV does not cause lysis, the mechanism used by many viruses to release newly formed virions. For HPV 11, a type associated with a low risk of neoplastic disease, the cornified cell envelope (CCE) of infected keratinocytes is thin and fragile, and transcription of loricrin, the major CCE protein, is reduced. The effects of high-risk HPV infection on components of the CCE have not been previously reported. HPV 59, an oncogenic genital type related to HPV types 18 and 45 was identified in a condylomata acuminata lesion. An extract of this lesion was used to infect human foreskin fragments, which were grown in athymic mice as xenografts. Continued propagation using extracts of xenografts permitted growth of additional HPV 59-infected xenografts. CCEs purified from HPV 59-infected xenografts displayed subtle morphologic abnormalities compared to those derived from uninfected xenografts. HPV 59-infected xenografts revealed dysplastic-appearing cells with mitotic figures. Detection of loricrin, involucrin, and cytokeratin 10 was reduced in HPV 59-infected epithelium, while small proline-rich protein 3 (SPR3) was increased. Reduction in loricrin was most apparent in regions of epithelium containing abundant HPV 59 DNA. Compared to uninfected epithelium, loricrin transcription was decreased in HPV 59-infected epithelium. We conclude that HPV 59 shares with HPV 11 the ability to alter CCE components and to specifically reduce transcription of the loricrin gene. Because loricrin is the major CCE protein, a reduction in this component could alter the physical properties of the CCE, thus facilitating virion release

  17. Efficient Gene Transfer in Bacterial Cell Chains

    OpenAIRE

    Babic, Ana; Berkmen, Melanie B.; Lee, Catherine A.; Grossman, Alan D.

    2011-01-01

    Horizontal gene transfer contributes to evolution and the acquisition of new traits. In bacteria, horizontal gene transfer is often mediated by conjugative genetic elements that transfer directly from cell to cell. Integrative and conjugative elements (ICEs; also known as conjugative transposons) are mobile genetic elements that reside within a host genome but can excise to form a circle and transfer by conjugation to recipient cells. ICEs contribute to the spread of genes involved in pathoge...

  18. Association of a Myosin Immunoanalogue with Cell Envelopes of Aspergillus fumigatus Conidia and Its Participation in Swelling and Germination

    OpenAIRE

    Esnault, Karine; el Moudni, Brahim; Bouchara, Jean-Philippe; Chabasse, Dominique; Tronchin, Guy

    1999-01-01

    A myosin immunoanalogue was identified in conidia of Aspergillus fumigatus by Western blotting, indirect immunofluorescence assay, and gold immunoelectron microscopy with two different antimyosin antibodies. The distribution pattern of this protein was followed during the early stages of germination. A single 180-kDa polypeptide, detected predominantly in a cell envelope extract, was found to cross-react with monoclonal and polyclonal antibodies raised against vertebrate muscle myosin. Immuno...

  19. Cellular Architecture of Treponema pallidum: Novel Flagellum, Periplasmic Cone, and Cell Envelope as Revealed by Cryo-Electron Tomography

    OpenAIRE

    Liu, Jun; Howell, Jerrilyn K.; Bradley, Sherille D.; Zheng, Yesha; Zhou, Z. Hong; Norris, Steven J

    2010-01-01

    High resolution cryo-electron tomography (cryo-ET) was utilized to visualize Treponema pallidum, the causative agent of syphilis, at the molecular level. Three-dimensional (3-D) reconstructions from 304 infectious organisms revealed unprecedented cellular structures of this unusual member in the spirochetal family. High resolution cryo-ET reconstructions provided the detailed structures of the cell envelope, which is significantly different from that of gram-negative bacteria. The 4 nm lipid ...

  20. Temporal expression of HIV-1 envelope proteins in baculovirus-infected insect cells: Implications for glycosylation and CD4 binding

    International Nuclear Information System (INIS)

    Three different human immunodeficiency virus type I (HIV-1) envelope derived recombinant proteins and the full length human CD4 polypeptide were expressed in Spodoptera frugiperda (Sf9) cells. DNA constructs encoding CD4, gp120, gp160, and gp160 delta were cloned into the baculovirus expression vector pVL941 or a derivative and used to generate recombinant viruses in a cotransfection with DNA from Autographa californica nuclear polyhedrosis virus (AcMNPV). Western blotting of cell extracts of the recombinant HIV-1 proteins showed that for each construct two major bands specifically reacted with anti-HIV-1 envelope antiserum. These bands corresponded to glycosylated and nonglycosylated versions of the HIV proteins as determined by 3H-mannose labeling and tunicamycin treatment of infected cells. A time course of HIV envelope expression revealed that at early times post-infection (24 hours) the proteins were fully glycosylated and soluble in nonionic detergents. However, at later times postinfection (48 hours), expression levels of recombinant protein reached a maximum but most of the increase was due to a rise in the level of the nonglycosylated species, which was largely insoluble in nonionic detergents. Thus, it appears that Sf9 cells cannot process large amounts of glycosylated recombinant proteins efficiently. As a measure of biological activity, the CD4 binding ability of both glycosylated and nonglycosylated recombinant HIV envelope proteins was tested in a coimmunoprecipitation assay. The results showed that CD4 and the glycosylated versions of recombinant gp120 or gp160 delta specifically associated with one another in this analysis. Nonglycosylated gp120 or gp160 delta proteins from tunicamycin-treated cultures did immunoprecipitate with anti-HIV-1 antiserum but did not interact with CD4

  1. Isolation of biologically active nanomaterial (inclusion bodies from bacterial cells

    Directory of Open Access Journals (Sweden)

    Peternel Špela

    2010-09-01

    Full Text Available Abstract Background In recent years bacterial inclusion bodies (IBs were recognised as highly pure deposits of active proteins inside bacterial cells. Such active nanoparticles are very interesting for further downstream protein isolation, as well as for many other applications in nanomedicine, cosmetic, chemical and pharmaceutical industry. To prepare large quantities of a high quality product, the whole bioprocess has to be optimised. This includes not only the cultivation of the bacterial culture, but also the isolation step itself, which can be of critical importance for the production process. To determine the most appropriate method for the isolation of biologically active nanoparticles, three methods for bacterial cell disruption were analyzed. Results In this study, enzymatic lysis and two mechanical methods, high-pressure homogenization and sonication, were compared. During enzymatic lysis the enzyme lysozyme was found to attach to the surface of IBs, and it could not be removed by simple washing. As this represents an additional impurity in the engineered nanoparticles, we concluded that enzymatic lysis is not the most suitable method for IBs isolation. During sonication proteins are released (lost from the surface of IBs and thus the surface of IBs appears more porous when compared to the other two methods. We also found that the acoustic output power needed to isolate the IBs from bacterial cells actually damages proteins structures, thereby causing a reduction in biological activity. High-pressure homogenization also caused some damage to IBs, however the protein loss from the IBs was negligible. Furthermore, homogenization had no side-effects on protein biological activity. Conclusions The study shows that among the three methods tested, homogenization is the most appropriate method for the isolation of active nanoparticles from bacterial cells.

  2. Heavy metal biosorption by bacterial cells

    Energy Technology Data Exchange (ETDEWEB)

    Vecchio, A.; Finoli, C.; Di Simine, D.; Andreoni, V. [Department of Food Science and Microbiology, State University, Milan (Italy)

    1998-06-01

    Microbial biomass provides available ligand groups on which metal ions bind by different mechanisms. Biosorption of these elements from aqueous solutions represents a remediation technology suitable for the treatment of metal-contaminated effluents. The purpose of the present investigation was the assessment of the capability of Brevibacterium sp. cells to remove bivalent ions, when present alone or in pairs, from aqueous solutions, using immobilized polyacrylamide cells of the microorganism in a flow-through system. The biosorption capacity of Brevibacterium cells was studied for lead, cadmium and copper. The metal cell binding capacity followed the order Cu > Pb > Cd, based on estimated q{sub max}. These values, expressed as mmol metal/g dry weight cells, were 0.54 for Cu, 0.36 for Pb and 0.14 for Cd. Polyacrylamide-gel immobilized cells were effective in Pb, Cu and Cd removal. Lead removal was not affected by the presence of Cd and Cu; lead instead inhibited Cd and Cu removal. The desorption of the metal, by fluxing a chelating solution, restored the metal binding capacity of the cells, thus affording the multiple use of the same biomass in the remediation treatment. (orig.) (orig.) With 5 figs., 4 tabs., 23 refs.

  3. Expression of Bacterial β-Galactosidase in Animal Cells

    OpenAIRE

    An, Gynheung; Hidaka, Katsuhiko; Siminovitch, Louis

    1982-01-01

    A recombinant plasmid containing the gene for bacterial β-galactosidase, situated close to the simian virus 40 early promoter, has been constructed. Transfection of CHO, L, and COS-1 cells with this plasmid led to the expression and appearance of the enzyme. Using this system, we have developed a series of promoter cloning vehicles capable of accepting promoter signals for animal genes.

  4. The Nuclear Envelope

    OpenAIRE

    Watson, Michael L.

    2010-01-01

    An electron microscope study of thin sections of interphase cells has revealed the following:— Circular pores are formed in the double nuclear envelope by continuities between the inner and outer membranes which permit contact between the nucleoplasm and the cytoplasm unmediated by a well defined membrane. The pores, seen in sections normal to the nuclear envelope, are profiles of the ring-shaped structures described by others and seen in tangential section. The inner and outer nuclear membra...

  5. Fluid dynamics and noise in bacterial cell-cell and cell-surface scattering

    CERN Document Server

    Drescher, Knut; Cisneros, Luis H; Ganguly, Sujoy; Goldstein, Raymond E; 10.1073/pnas.1019079108

    2011-01-01

    Bacterial processes ranging from gene expression to motility and biofilm formation are constantly challenged by internal and external noise. While the importance of stochastic fluctuations has been appreciated for chemotaxis, it is currently believed that deterministic long-range fluid dynamical effects govern cell-cell and cell-surface scattering - the elementary events that lead to swarming and collective swimming in active suspensions and to the formation of biofilms. Here, we report the first direct measurements of the bacterial flow field generated by individual swimming Escherichia coli both far from and near to a solid surface. These experiments allowed us to examine the relative importance of fluid dynamics and rotational diffusion for bacteria. For cell-cell interactions it is shown that thermal and intrinsic stochasticity drown the effects of long-range fluid dynamics, implying that physical interactions between bacteria are determined by steric collisions and near-field lubrication forces. This dom...

  6. Cell Wall Nonlinear Elasticity and Growth Dynamics: How Do Bacterial Cells Regulate Pressure and Growth?

    Science.gov (United States)

    Deng, Yi

    In my thesis, I study intact and bulging Escherichia coli cells using atomic force microscopy to separate the contributions of the cell wall and turgor pressure to the overall cell stiffness. I find strong evidence of power--law stress--stiffening in the E. coli cell wall, with an exponent of 1.22±0.12, such that the wall is significantly stiffer in intact cells (E = 23±8 MPa and 49±20 MPa in the axial and circumferential directions) than in unpressurized sacculi. These measurements also indicate that the turgor pressure in living cells E. coli is 29±3 kPa. The nonlinearity in cell elasticity serves as a plausible mechanism to balance the mechanical protection and tension measurement sensitivity of the cell envelope. I also study the growth dynamics of the Bacillus subtilis cell wall to help understand the mechanism of the spatiotemporal order of inserting new cell wall material. High density fluorescent markers are used to label the entire cell surface to capture the morphological changes of the cell surface at sub-cellular to diffraction-limited spatial resolution and sub-minute temporal resolution. This approach reveals that rod-shaped chaining B. subtilis cells grow and twist in a highly heterogeneous fashion both spatially and temporally. Regions of high growth and twisting activity have a typical length scale of 5 μm, and last for 10-40 minutes. Motivated by the quantification of the cell wall growth dynamics, two microscopy and image analysis techniques are developed and applied to broader applications beyond resolving bacterial growth. To resolve densely distributed quantum dots, we present a fast and efficient image analysis algorithm, namely Spatial Covariance Reconstruction (SCORE) microscopy that takes into account the blinking statistics of the fluorescence emitters. We achieve sub-diffraction lateral resolution of 100 nm from 5 to 7 seconds of imaging, which is at least an order of magnitude faster than single-particle localization based methods

  7. Bounds on bacterial cell growth rates

    CERN Document Server

    Landy, Jonathan

    2013-01-01

    Recent experiments have shown that rod-like bacteria in nutrient-rich media grow in length at an exponential rate. Here, I point out that it is the elongated shape of these bacteria that allows for this behavior. Further, I show that when a bacterium's growth is limited by some nutrient -- taken in by the cell through a diffusion-to-capture process -- its growth is suppressed: In three-dimensional geometries, the length $L$ is bounded by $\\log L \\lesssim t^{1/2}$, while in two dimensions the length is bounded by a power-law form. Fits of experimental growth curves to these predicted, sub-exponential forms could allow for direct measures of quantities relating to cellular metabolic rates.

  8. Bursting the bubble on bacterial biofilms: a flow cell methodology

    OpenAIRE

    Shanika A. Crusz; Popat, Roman; Rybtke, Morten Theil; Cámara, Miguel; Givskov, Michael; Tolker-Nielsen, Tim; Diggle, Stephen P.; Williams, Paul

    2012-01-01

    The flow cell biofilm system is an important and widely used tool for the in vitro cultivation and evaluation of bacterial biofilms under hydrodynamic conditions of flow. This paper provides an introduction to the background and use of such systems, accompanied by a detailed guide to the assembly of the apparatus including the description of new modifications which enhance its performance. As such, this is an essential guide for the novice biofilm researcher as well as providing valuable trou...

  9. Role of the Gram-Negative Envelope Stress Response in the Presence of Antimicrobial Agents.

    Science.gov (United States)

    Guest, Randi L; Raivio, Tracy L

    2016-05-01

    Bacterial survival necessitates endurance of many types of antimicrobial compound. Many Gram-negative envelope stress responses, which must contend with an outer membrane and a dense periplasm containing the cell wall, have been associated with the status of protein folding, membrane homeostasis, and physiological functions such as efflux and the proton motive force (PMF). In this review, we discuss evidence that indicates an emerging role for Gram-negative envelope stress responses in enduring exposure to diverse antimicrobial substances, focusing on recent studies of the γ-proteobacterial Cpx envelope stress response. PMID:27068053

  10. H2-O2 fuel cell and advanced battery power systems for autonomous underwater vehicles: performance envelope comparisons

    International Nuclear Information System (INIS)

    Autonomous underwater vehicles have traditionally been powered by low energy density lead-acid batteries. Recently, advanced battery technologies and H2-O2 fuel cells have become available, offering significant improvements in performance. This paper compares the solid polymer fuel cell to the lithium-thionyl chloride primary battery, sodium-sulfur battery, and lead acid battery for a variety of missions. The power system performance is simulated using computer modelling techniques. Performance envelopes are constructed, indicating domains of preference for competing power system technologies. For most mission scenarios, the solid polymer fuel cell using liquid reactant storage is the preferred system. Nevertheless, the advanced battery systems are competitive with the fuel cell systems using gaseous hydrogen storage, and they illustrate preferred performance for missions requiring high power density. 11 figs., 4 tabs., 15 refs

  11. A recombinant West Nile virus envelope protein vaccine candidate produced in Spodoptera frugiperda expresSF+ cells

    OpenAIRE

    Bonafé, Nathalie; Rininger, Joseph A.; Chubet, Richard G.; Foellmer, Harald G.; Fader, Stacey; Anderson, John F.; Bushmich, Sandra L.; Anthony, Karen; Ledizet, Michel; Fikrig, Erol; Koski, Raymond A.; Kaplan, Paul

    2008-01-01

    In this study, a recombinant truncated West Nile virus envelope protein antigen (rWNV-E) was produced in serum-free cultures of the expresSF+ insect cell line via baculovirus infection. This production system was selected based on its use in the production of candidate human and animal vaccine antigens. A defined fermentation and purification process for the rWNV-E antigen was established to control for purity and immunogenicity of each protein batch. The material formulated with aluminum hyd...

  12. The Role of Lipid Domains in Bacterial Cell Processes

    Directory of Open Access Journals (Sweden)

    Katarína Muchová

    2013-02-01

    Full Text Available Membranes are vital structures for cellular life forms. As thin, hydrophobic films, they provide a physical barrier separating the aqueous cytoplasm from the outside world or from the interiors of other cellular compartments. They maintain a selective permeability for the import and export of water-soluble compounds, enabling the living cell to maintain a stable chemical environment for biological processes. Cell membranes are primarily composed of two crucial substances, lipids and proteins. Bacterial membranes can sense environmental changes or communication signals from other cells and they support different cell processes, including cell division, differentiation, protein secretion and supplementary protein functions. The original fluid mosaic model of membrane structure has been recently revised because it has become apparent that domains of different lipid composition are present in both eukaryotic and prokaryotic cell membranes. In this review, we summarize different aspects of phospholipid domain formation in bacterial membranes, mainly in Gram-negative Escherichia coli and Gram-positive Bacillus subtilis. We describe the role of these lipid domains in membrane dynamics and the localization of specific proteins and protein complexes in relation to the regulation of cellular function.

  13. Membrane topology and mutational analysis of the TolQ protein of Escherichia coli required for the uptake of macromolecules and cell envelope integrity.

    OpenAIRE

    Vianney, A; Lewin, T M; Beyer, W F; Lazzaroni, J C; Portalier, R; Webster, R E

    1994-01-01

    TolQ is a 230-amino-acid protein required to maintain the integrity of the bacterial envelope and to facilitate the import of both filamentous bacteriophage and group A colicins. Cellular fractionation experiments showed TolQ to be localized to the cytoplasmic membrane. Bacteria expressing a series of TolQ-beta-galactosidase and TolQ-alkaline phosphatase fusion proteins were analyzed for the appropriate enzyme activity, membrane location, and sensitivity to exogenously added protease. The res...

  14. Bacterial-mediated DNA delivery to tumour associated phagocytic cells.

    Science.gov (United States)

    Byrne, W L; Murphy, C T; Cronin, M; Wirth, T; Tangney, M

    2014-12-28

    Phagocytic cells including macrophages, dendritic cells and neutrophils are now recognised as playing a negative role in many disease settings including cancer. In particular, macrophages are known to play a pathophysiological role in multiple diseases and present a valid and ubiquitous therapeutic target. The technology to target these phagocytic cells in situ, both selectively and efficiently, is required in order to translate novel therapeutic modalities into clinical reality. We present a novel delivery strategy using non-pathogenic bacteria to effect gene delivery specifically to tumour-associated phagocytic cells. Non-invasive bacteria lack the ability to actively enter host cells, except for phagocytic cells. We exploit this natural property to effect 'passive transfection' of tumour-associated phagocytic cells following direct administration of transgene-loaded bacteria to tumour regions. Using an in vitro-differentiated human monocyte cell line and two in vivo mouse models (an ovarian cancer ascites and a solid colon tumour model) proof of delivery is demonstrated with bacteria carrying reporter constructs. The results confirm that the delivery strategy is specific for phagocytic cells and that the bacterial vector itself recruits more phagocytic cells to the tumour. While proof of delivery to phagocytic cells is demonstrated in vivo for solid and ascites tumour models, this strategy may be applied to other settings, including non-cancer related disease. PMID:25466954

  15. The Membrane Steps of Bacterial Cell Wall Synthesis as Antibiotic Targets.

    Science.gov (United States)

    Liu, Yao; Breukink, Eefjan

    2016-01-01

    Peptidoglycan is the major component of the cell envelope of virtually all bacteria. It has structural roles and acts as a selective sieve for molecules from the outer environment. Peptidoglycan synthesis is therefore one of the most important biogenesis pathways in bacteria and has been studied extensively over the last twenty years. The pathway starts in the cytoplasm, continues in the cytoplasmic membrane and finishes in the periplasmic space, where the precursor is polymerized into the peptidoglycan layer. A number of proteins involved in this pathway, such as the Mur enzymes and the penicillin binding proteins (PBPs), have been studied and regarded as good targets for antibiotics. The present review focuses on the membrane steps of peptidoglycan synthesis that involve two enzymes, MraY and MurG, the inhibitors of these enzymes and the inhibition mechanisms. We also discuss the challenges of targeting these two cytoplasmic membrane (associated) proteins in bacterial cells and the perspectives on how to overcome the issues. PMID:27571111

  16. The Membrane Steps of Bacterial Cell Wall Synthesis as Antibiotic Targets

    Directory of Open Access Journals (Sweden)

    Yao Liu

    2016-08-01

    Full Text Available Peptidoglycan is the major component of the cell envelope of virtually all bacteria. It has structural roles and acts as a selective sieve for molecules from the outer environment. Peptidoglycan synthesis is therefore one of the most important biogenesis pathways in bacteria and has been studied extensively over the last twenty years. The pathway starts in the cytoplasm, continues in the cytoplasmic membrane and finishes in the periplasmic space, where the precursor is polymerized into the peptidoglycan layer. A number of proteins involved in this pathway, such as the Mur enzymes and the penicillin binding proteins (PBPs, have been studied and regarded as good targets for antibiotics. The present review focuses on the membrane steps of peptidoglycan synthesis that involve two enzymes, MraY and MurG, the inhibitors of these enzymes and the inhibition mechanisms. We also discuss the challenges of targeting these two cytoplasmic membrane (associated proteins in bacterial cells and the perspectives on how to overcome the issues.

  17. Virus and Bacterial Cell Chemical Analysis by NanoSIMS

    Energy Technology Data Exchange (ETDEWEB)

    Weber, P; Holt, J

    2008-07-28

    In past work for the Department of Homeland Security, the LLNL NanoSIMS team has succeeded in extracting quantitative elemental composition at sub-micron resolution from bacterial spores using nanometer-scale secondary ion mass spectrometry (NanoSIMS). The purpose of this task is to test our NanoSIMS capabilities on viruses and bacterial cells. This initial work has proven successful. We imaged Tobacco Mosaic Virus (TMV) and Bacillus anthracis Sterne cells using scanning electron microscopy (SEM) and then analyzed those samples by NanoSIMS. We were able resolve individual viral particles ({approx}18 nm by 300 nm) in the SEM and extract correlated elemental composition in the NanoSIMS. The phosphorous/carbon ratio observed in TMV is comparable to that seen in bacterial spores (0.033), as was the chlorine/carbon ratio (0.11). TMV elemental composition is consistent from spot to spot, and TMV is readily distinguished from debris by NanoSIMS analysis. Bacterial cells were readily identified in the SEM and relocated in the NanoSIMS for elemental analysis. The Ba Sterne cells were observed to have a measurably lower phosphorous/carbon ratio (0.005), as compared to the spores produced in the same run (0.02). The chlorine/carbon ratio was approximately 2.5X larger in the cells (0.2) versus the spores (0.08), while the fluorine/carbon ratio was approximately 10X lower in the cells (0.008) than the spores (0.08). Silicon/carbon ratios for both cells and spores encompassed a comparable range. The initial data in this study suggest that high resolution analysis is useful because it allows the target agent to be analyzed separate from particulates and other debris. High resolution analysis would also be useful for trace sample analysis. The next step in this work is to determine the potential utility of elemental signatures in these kinds of samples. We recommend bulk analyses of media and agent samples to determine the range of media compositions in use, and to determine how

  18. The immunosuppressive domain of the transmembrane envelope protein gp41 of HIV-1 binds to human monocytes and B cells.

    Science.gov (United States)

    Mühle, Michael; Kroniger, Tobias; Hoffmann, Kerstin; Denner, Joachim

    2016-06-01

    The induction of the acquired immunodeficiency syndrome by the human immunodeficiency virus-1 (HIV-1) is a complex process which is not yet understood in full detail. Still open is the question whether the highly conserved so-called immunosuppressive (Isu) domain in the transmembrane envelope (TM) protein gp41 of HIV-1 is actively participating in immunopathogenesis. Inactivated virus particles, recombinant gp41 and peptides corresponding to the Isu domain have been reported to inhibit lymphocyte proliferation, as well as to alter cytokine release and gene expression. Here we demonstrate, using fluorescence-activated cell sorting and competition experiments, that homopolymers of the Isu peptide of HIV-1 are binding specifically to human peripheral blood mononuclear cells, mainly to monocytes and B cells. These data suggest that a putative receptor might be involved in the immunomodulatory effects observed previously. PMID:26754765

  19. Cellular Architecture of Treponema pallidum: Novel Flagellum, Periplasmic Cone, and Cell Envelope as Revealed by Cryo-Electron Tomography

    Science.gov (United States)

    Liu, Jun; Howell, Jerrilyn K.; Bradley, Sherille D.; Zheng, Yesha; Zhou, Z. Hong; Norris, Steven J.

    2010-01-01

    High resolution cryo-electron tomography (cryo-ET) was utilized to visualize Treponema pallidum, the causative agent of syphilis, at the molecular level. Three-dimensional (3-D) reconstructions from 304 infectious organisms revealed unprecedented cellular structures of this unusual member in the spirochetal family. High resolution cryo-ET reconstructions provided the detailed structures of the cell envelope, which is significantly different from that of gram-negative bacteria. The 4 nm lipid bilayer of both outer and cytoplasmic membranes resolved in 3-D reconstructions, providing an important marker for interpreting membrane-associated structures. Abundant lipoproteins cover the outer leaflet of the cytoplasmic membrane, in contrast to the rare outer membrane proteins visible by scanning probe microscopy. High resolution cryo-ET images also provided the first observation of T. pallidum chemoreceptor arrays, as well as structural details of the periplasmically located, cone-shaped structure at both ends of bacterium. Furthermore, 3-D subvolume averages of the periplasmic flagellar motors and filaments from living organisms revealed the novel flagellar architectures that may facilitate their rotation within the confining periplasmic space. Together, our findings provide the most detailed structural understanding of the periplasmic flagella and the surrounding cell envelope, which enable this enigmatic bacterium to efficiently penetrate tissue and escape host immune responses. PMID:20850455

  20. Colon-targeted delivery of live bacterial cell biotherapeutics including microencapsulated live bacterial cells

    OpenAIRE

    Satya Prakash; Aleksandra Malgorzata Urbanska

    2008-01-01

    Satya Prakash, Aleksandra Malgorzata UrbanskaBiomedical Technology and Cell Therapy Research Laboratory, Departments of Biomedical Engineering and Physiology, Artificial Cells and Organs Research Center, Faculty of Medicine, McGill University, Montreal, Quebec, CanadaAbstract: There has been an ample interest in delivery of therapeutic molecules using live cells. Oral delivery has been stipulated as best way to deliver live cells to humans for therapy. Colon, in particular, is a part of gastr...

  1. Differential sensitivity of bat cells to infection by enveloped RNA viruses: coronaviruses, paramyxoviruses, filoviruses, and influenza viruses.

    Directory of Open Access Journals (Sweden)

    Markus Hoffmann

    Full Text Available Bats (Chiroptera host major human pathogenic viruses including corona-, paramyxo, rhabdo- and filoviruses. We analyzed six different cell lines from either Yinpterochiroptera (including African flying foxes and a rhinolophid bat or Yangochiroptera (genera Carollia and Tadarida for susceptibility to infection by different enveloped RNA viruses. None of the cells were sensitive to infection by transmissible gastroenteritis virus (TGEV, a porcine coronavirus, or to infection mediated by the Spike (S protein of SARS-coronavirus (SARS-CoV incorporated into pseudotypes based on vesicular stomatitis virus (VSV. The resistance to infection was overcome if cells were transfected to express the respective cellular receptor, porcine aminopeptidase N for TGEV or angiotensin-converting enzyme 2 for SARS-CoV. VSV pseudotypes containing the S proteins of two bat SARS-related CoV (Bg08 and Rp3 were unable to infect any of the six tested bat cell lines. By contrast, viral pseudotypes containing the surface protein GP of Marburg virus from the family Filoviridae infected all six cell lines though at different efficiency. Notably, all cells were sensitive to infection by two paramyxoviruses (Sendai virus and bovine respiratory syncytial virus and three influenza viruses from different subtypes. These results indicate that bat cells are more resistant to infection by coronaviruses than to infection by paramyxoviruses, filoviruses and influenza viruses. Furthermore, these results show a receptor-dependent restriction of the infection of bat cells by CoV. The implications for the isolation of coronaviruses from bats are discussed.

  2. Biomimetic Envelopes

    Directory of Open Access Journals (Sweden)

    Ilaria Mazzoleni

    2010-06-01

    Full Text Available How to translate the lessons learned from the analysis and observation of the animal world is the design learning experience presented in this article. Skin is a complex and incredibly sophisticated organ that performs various functions, including protection, sensation and heat and water regulation. In a similar way building envelopes serve multiple roles, as they are the interface between the building inhabitants and environmental elements. The resulting architectural building envelopes proto-architectural research and design projects here presented, inspired by the study of animal skins, perform and respond; they take into consideration various dynamic local environmental conditions, enhancing and supporting them rather than exploiting them, creating a more sustainable way of building and living.

  3. Enveloping algebras

    International Nuclear Information System (INIS)

    Since the works of Gelfand, Harish-Chandra, Kostant and Duflo, a new theory has earned its place in the field of mathematics, due to the abundance of its results and the coherence of its methods: the theory of enveloping algebras. This study is the first to present the whole subject in textbook form. The most recent results are included, as well as complete proofs, starting from the elementary theory of Lie algebras. (Auth.)

  4. INTERNAL ENVELOPES

    CERN Multimedia

    Mail Office

    2001-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration.

  5. Structure of a Bacterial Cell Surface Decaheme Electron Conduit

    Energy Technology Data Exchange (ETDEWEB)

    Clarke, Thomas A.; Edwards, Marcus; Gates, Andrew J.; Hall, Andrea; White, Gaye; Bradley, Justin; Reardon, Catherine L.; Shi, Liang; Beliaev, Alex S.; Marshall, Matthew J.; Wang, Zheming; Watmough, Nicholas; Fredrickson, Jim K.; Zachara, John M.; Butt, Julea N.; Richardson, David J.

    2011-05-23

    Some bacterial species are able to utilize extracellular mineral forms of iron and manganese as respiratory electron acceptors. In Shewanella oneidensis this involves deca-heme cytochromes that are located on the bacterial cell surface at the termini of trans-outermembrane (OM) electron transfer conduits. The cell surface cytochromes can potentially play multiple roles in mediating electron transfer directly to insoluble electron sinks, catalyzing electron exchange with flavin electron shuttles or participating in extracellular inter-cytochrome electron exchange along ‘nanowire’ appendages. We present a 3.2 Å crystal structure of one of these deca-heme cytochromes, MtrF, that allows the spatial organization of the ten hemes to be visualized for the first time. The hemes are organized across four domains in a unique crossed conformation, in which a staggered 65 Å octa-heme chain transects the length of the protein and is bisected by a planar 45 Å tetra-heme chain that connects two extended Greek key split β-barrel domains. The structure provides molecular insight into how reduction of insoluble substrate (e.g. minerals), soluble substrates (e.g. flavins) and cytochrome redox partners might be possible in tandem at different termini of a trifurcated electron transport chain on the cell surface.

  6. Formation of infectious hybrid virions with gibbon ape leukemia virus and human T-cell leukemia virus retroviral envelope glycoproteins and the gag and pol proteins of Moloney murine leukemia virus.

    OpenAIRE

    C. Wilson; Reitz, M S; Okayama, H; Eiden, M V

    1989-01-01

    The gibbon ape leukemia virus, SEATO strain, and human T-cell leukemia virus type I envelope glycoproteins can be functionally assembled with a Moloney murine leukemia virus core into infectious particles. The envelope-host cell receptor interaction is the major determinant of the host cell specificity for these hybrid virions.

  7. Lung Dendritic Cells Facilitate Extrapulmonary Bacterial Dissemination during Pneumococcal Pneumonia

    Directory of Open Access Journals (Sweden)

    Alva eRosendahl

    2013-06-01

    Full Text Available Streptococcus pneumoniae is a leading cause of bacterial pneumonia worldwide. Given the critical role of dendritic cells (DCs in regulating and modulating the immune response to pathogens, we investigated here the role of DCs in S. pneumoniae lung infections. Using a well-established transgenic mouse line which allows the conditional transient depletion of DCs, we showed that ablation of DCs resulted in enhanced resistance to intranasal challenge with S. pneumoniae. DC-depleted mice exhibited delayed bacterial systemic dissemination, significantly reduced bacterial loads in the infected organs and lower levels of serum inflammatory mediators than non-depleted animals. The increased resistance of DC-depleted mice to S. pneumoniae was associated with a better capacity to restrict pneumococci extrapulmonary dissemination. Furthermore, we demonstrated that S. pneumoniae disseminated from the lungs into the regional lymph nodes in a cell-independent manner and that this direct way of dissemination was much more efficient in the presence of DCs. We also provide evidence that S. pneumoniae induces expression and activation of matrix metalloproteinase-9 (MMP-9 in cultured bone marrow-derived DCs. MMP-9 is a protease involved in the breakdown of extracellular matrix proteins and is critical for DC trafficking across extracellular matrix and basement membranes during the migration from the periphery to the lymph nodes. MMP-9 was also significantly up-regulated in the lungs of mice after intranasal infection with S. pneumoniae. Notably, the expression levels of MMP-9 in the infected lungs were significantly decreased after depletion of DCs suggesting the involvement of DCs in MMP-9 production during pneumococcal pneumonia. Thus, we propose that S. pneumoniae can exploit the DC-derived proteolysis to open tissue barriers thereby facilitating its own dissemination from the local site of infection.

  8. Fate of deposited cells in an aerobic binary bacterial biofilm

    International Nuclear Information System (INIS)

    A biofilm is a matrix of microbial cells and their extracellular products that is associated with a solid surface. Previous studies on biofilm development have employed only dissolved compounds as growth limiting substrates, without the influence of microbial species invading from the bulk liquid. The goal of this research project was to quantify the kinetics of processes governing suspended biomass turnover in biofilm systems, and the accompanying effects of suspended cell deposition on biofilm population dynamics. Experiments were conducted with two species of bacteria, Pseudomonas putida ATCC 11172 grown on glucose, and Hyphomicrobium ZV620 grown on methanol. Cryptic growth and particulate hydrolysis studies were evaluated, using combinations of these two bacteria, by measuring the uptake of radiolabelled cell lysis products, under batch conditions. Biofilms studies were performed to investigate bacterial deposition, continual biofilm removal by shear induced erosion, and biofilm ecology. Biofilms were developed in a flow cell reactor, under laminar flow conditions. Bacterial species were differentiated by radioactively labelling each species with their carbon substrate. A mathematical model was developed to predict the biofilm ecology of mixed cultures. The equations developed predict biofilm accumulation, as well as substrate and oxygen consumption. Results indicate that cryptic growth will occur for bacteria growing on their own species soluble lysis products and in some cases, bacteria growing on the soluble lysis products of other species. Particulate hydrolysis only occurred for Pseudomonas putida growing on Pseudomonas putida lysis products, but the lack of particulate hydrolysis occurring in the other studies may have been due to the short experimental period

  9. Softness of the bacterial cell wall of Streptococcus mitis as probed by micro-electrophoresis

    NARCIS (Netherlands)

    Vadillo-Rodriguez, V.; Busscher, H.J.; Norde, W.; Mei, van der H.C.

    2002-01-01

    Chemical and structural complexity of bacterial cell surfaces complicate accurate quantification of cell surfaces properties. The presence of fibrils, fimbriae or other surface appendages on bacterial cell surfaces largely influence those properties and would therefore play a major function in inter

  10. Studies on penetration of antibiotic in bacterial cells in space conditions (7-IML-1)

    Science.gov (United States)

    Tixador, R.

    1992-01-01

    The Cytos 2 experiment was performed aboard Salyut 7 in order to test the antibiotic sensitivity of bacteria cultivated in vitro in space. An increase of the Minimal Inhibitory Concentration (MIC) in the inflight cultures (i.e., an increase of the antibiotic resistance) was observed. Complementary studies of the ultrastructure showed a thickening of the cell envelope. In order to confirm the results of the Cytos 2 experiment, we performed the ANTIBIO experiment during the D1 mission to try to differentiate, by means of the 1 g centrifuge in the Biorack, between the biological effects of cosmic rays and those caused by microgravity conditions. The originality of this experiment was in the fact that it was designed to test the antibiotic sensitivity of bacteria cultivated in vitro during the orbital phase of the flight. The results show an increase in resistance to Colistin in in-flight bacteria. The MIC is practically double in the in-flight cultures. A cell count of living bacteria in the cultures containing the different Colistin concentrations showed a significant difference between the cultures developed during space flight and the ground based cultures. The comparison between the 1 g and 0 g in-flight cultures show similar behavior for the two sets. Nevertheless, a small difference between the two sets of ground based control cultures was noted. The cultures developed on the ground centrifuge (1.4 g) present a slight decrease in comparison with the cultures developed in the static rack (1 g). In order to approach the mechanisms of the increase of antibiotic resistance on bacteria cultivated in vitro in space, we have proposed the study on penetration of antibiotics in bacterial cells in space conditions. This experiment was selected for the International Microgravity Laboratory 1 (IML-1) mission.

  11. Deficiency of a Sinorhizobium meliloti bacA Mutant in Alfalfa Symbiosis Correlates with Alteration of the Cell Envelope

    OpenAIRE

    Ferguson, Gail P.; Roop II, R. Martin; Walker, Graham C.

    2002-01-01

    The BacA protein is essential for the long-term survival of Sinorhizobium meliloti and Brucella abortus within acidic compartments in plant and animal cells, respectively. Since both the S. meliloti and B. abortus bacA mutants have an increased resistance to bleomycin, it was hypothesized that BacA was a transporter of bleomycin and bleomycin-like compounds into the bacterial cell. However, our finding that the S. meliloti bacA mutant also has an increased sensitivity to detergents, a hydroph...

  12. Principles of bacterial cell-size determination revealed by cell wall synthesis perturbations

    OpenAIRE

    Carolina Tropini; Timothy K. Lee; Jen Hsin; Samantha M. Desmarais; Tristan Ursell; Russell D. Monds; Kerwyn Casey Huang

    2014-01-01

    Although bacterial cell morphology is tightly controlled, the principles of size regulation remain elusive. In Escherichia coli, perturbation of cell-wall synthesis often results in similar morphologies, making it difficult to deconvolve the complex genotype-phenotype relationships underlying morphogenesis. Here we modulated cell width through heterologous expression of sequences encoding the essential enzyme PBP2 and through sublethal treatments with drugs that inhibit PBP2 and the MreB cyto...

  13. (p)ppGpp and the bacterial cell cycle

    Indian Academy of Sciences (India)

    Aanisa Nazir; Rajendran Harinarayanan

    2016-06-01

    Genes of the Rel/Spo homolog (RSH) superfamily synthesize and/or hydrolyse the modified nucleotides pppGpp/ppGpp (collectively referred to as (p)ppGpp) and are prevalent across diverse bacteria and in plant chloroplasts. Bacteria accumulate (p)ppGpp in response to nutrient deprivation (generically called the stringent response) and elicit appropriate adaptive responses mainly through the regulation of transcription. Although at different concentrations (p)ppGpp affect the expression of distinct set of genes, the two well-characterized responses are reduction in expression of the protein synthesis machinery and increase in the expression of genes coding for amino acid biosynthesis. In Escherichia coli, the cellular (p)ppGpp level inversely correlates with the growth rate and increasing its concentration decreases the steady state growth rate in a defined growth medium. Since change in growth rate must be accompanied by changes in cell cycle parameters set through the activities of the DNA replication and cell division apparatus, (p)ppGpp could coordinate protein synthesis (cell mass increase) with these processes. Here we review the role of (p)ppGpp in bacterial cell cycle regulation.

  14. Depletion of the surface CD4 molecule by the envelope protein of human immunodeficiency virus expressed in a human CD4+ monocytoid cell line

    International Nuclear Information System (INIS)

    A CD4+ human monocytoid cell line, U937, was transfected with a constructed plasmid which has the envelope gene of human immunodeficiency virus under the transcriptional control of the human metallothionein IIA promoter and was cloned thereafter. These cloned cell lines (EH and EL cells) expressed the viral gp160 in the cytoplasm. The expression of surface CD4 antigen examined by Leu3a and OKT4 monoclonal antibodies, however, disappeared completely in EH cells, which produce a larger amount of gp160, while diminishing only partly in EL cells, which produce a smaller amount of gp160. These results indicate that the level of expression of surface CD4 antigen correlates inversely with the amount of intracellular gp160. Moreover, immunoprecipitation studies using lysate from EH cells showed that OKT4 monoclonal antibody precipitated a significant number of CD4 molecules even after surface CD4 disappeared. However, Leu3a monoclonal antibody, which recognizes the binding site for envelope protein, could not precipitate any CD4 molecules in the same cell lysate. Taken together, these results suggested that CD4 molecules are still synthesized normally after the augmented production of gp160 in the cells but form a complex with the envelope protein in the cytoplasm and become unable to be transported to the cell surface, resulting in the observed depletion of surface CD4 antigen. This mechanism may explain the decrease or absence of surface CD4 antigens in human lymphocytes infected with human immunodeficiency virus

  15. Phase Diagram of Collective Motion of Bacterial Cells in a Shallow Circular Pool

    OpenAIRE

    Wakita, Jun-ichi; Tsukamoto, Shota; Yamamoto, Ken; Katori, Makoto; Yamada, Yasuyuki

    2015-01-01

    The collective motion of bacterial cells in a shallow circular pool is systematically studied using the bacterial species $Bacillus$ $subtilis$. The ratio of cell length to pool diameter (i.e., the reduced cell length) ranges from 0.06 to 0.43 in our experiments. Bacterial cells in a circular pool show various types of collective motion depending on the cell density in the pool and the reduced cell length. The motion is classified into six types, which we call random motion, turbulent motion,...

  16. Human Cytomegalovirus Nuclear Egress Proteins Ectopically Expressed in the Heterologous Environment of Plant Cells are Strictly Targeted to the Nuclear Envelope.

    Science.gov (United States)

    Lamm, Christian E; Link, Katrin; Wagner, Sabrina; Milbradt, Jens; Marschall, Manfred; Sonnewald, Uwe

    2016-01-01

    In all eukaryotic cells, the nucleus forms a prominent cellular compartment containing the cell's nuclear genome. Although structurally similar, animal and plant nuclei differ substantially in details of their architecture. One example is the nuclear lamina, a layer of tightly interconnected filament proteins (lamins) underlying the nuclear envelope of metazoans. So far no orthologous lamin genes could be detected in plant genomes and putative lamin-like proteins are only poorly described in plants. To probe for potentially conserved features of metazoan and plant nuclear envelopes, we ectopically expressed the core nuclear egress proteins of human cytomegalovirus pUL50 and pUL53 in plant cells. pUL50 localizes to the inner envelope of metazoan nuclei and recruits the nuclear localized pUL53 to it, forming heterodimers. Upon expression in plant cells, a very similar localization pattern of both proteins could be determined. Notably, pUL50 is specifically targeted to the plant nuclear envelope in a rim-like fashion, a location to which coexpressed pUL53 becomes strictly corecruited from its initial nucleoplasmic distribution. Using pUL50 as bait in a yeast two-hybrid screening, the cytoplasmic re-initiation supporting protein RISP could be identified. Interaction of pUL50 and RISP could be confirmed by coexpression and coimmunoprecipitation in mammalian cells and by confocal laser scanning microscopy in plant cells, demonstrating partial pUL50-RISP colocalization in areas of the nuclear rim and other intracellular compartments. Thus, our study provides strong evidence for conserved structural features of plant and metazoan nuclear envelops and identifies RISP as a potential pUL50-interacting plant protein. PMID:26978388

  17. Bacterial glycosidases for the production of universal red blood cells.

    Science.gov (United States)

    Liu, Qiyong P; Sulzenbacher, Gerlind; Yuan, Huaiping; Bennett, Eric P; Pietz, Greg; Saunders, Kristen; Spence, Jean; Nudelman, Edward; Levery, Steven B; White, Thayer; Neveu, John M; Lane, William S; Bourne, Yves; Olsson, Martin L; Henrissat, Bernard; Clausen, Henrik

    2007-04-01

    Enzymatic removal of blood group ABO antigens to develop universal red blood cells (RBCs) was a pioneering vision originally proposed more than 25 years ago. Although the feasibility of this approach was demonstrated in clinical trials for group B RBCs, a major obstacle in translating this technology to clinical practice has been the lack of efficient glycosidase enzymes. Here we report two bacterial glycosidase gene families that provide enzymes capable of efficient removal of A and B antigens at neutral pH with low consumption of recombinant enzymes. The crystal structure of a member of the alpha-N-acetylgalactosaminidase family reveals an unusual catalytic mechanism involving NAD+. The enzymatic conversion processes we describe hold promise for achieving the goal of producing universal RBCs, which would improve the blood supply while enhancing the safety of clinical transfusions. PMID:17401360

  18. Envelope-specific B-cell populations in African green monkeys chronically infected with simian immunodeficiency virus.

    Science.gov (United States)

    Zhang, Ruijun; Martinez, David R; Nguyen, Quang N; Pollara, Justin; Arifin, Trina; Stolarchuk, Christina; Foulger, Andrew; Amos, Josh D; Parks, Robert; Himes, Jonathon E; Wang, Minyue; Edwards, Regina W; Trama, Ashley M; Vandergrift, Nathan; Colvin, Lisa; Dewar, Ken; Juretic, Nikoleta; Wasserscheid, Jessica; Ferrari, Guido; Liao, Hua-Xin; Permar, Sallie R

    2016-01-01

    African green monkeys (AGMs) are natural primate hosts of simian immunodeficiency virus (SIV). Interestingly, features of the envelope-specific antibody responses in SIV-infected AGMs are distinct from that of HIV-infected humans and SIV-infected rhesus monkeys, including gp120-focused responses and rapid development of autologous neutralization. Yet, the lack of genetic tools to evaluate B-cell lineages hinders potential use of this unique non-human primate model for HIV vaccine development. Here we define features of the AGM Ig loci and compare the proportion of Env-specific memory B-cell populations to that of HIV-infected humans and SIV-infected rhesus monkeys. AGMs appear to have a higher proportion of Env-specific memory B cells that are mainly gp120 directed. Furthermore, AGM gp120-specific monoclonal antibodies display robust antibody-dependent cellular cytotoxicity and CD4-dependent virion capture activity. Our results support the use of AGMs to model induction of functional gp120-specific antibodies by HIV vaccine strategies. PMID:27381634

  19. Bacterial Cell Wall-Induced Arthritis: Chemical Composition and Tissue Distribution of Four Lactobacillus Strains

    OpenAIRE

    Šimelyte, Egle; Rimpiläinen, Marja; Lehtonen, Leena; Zhang, Xiang; Toivanen, Paavo

    2000-01-01

    To study what determines the arthritogenicity of bacterial cell walls, cell wall-induced arthritis in the rat was applied, using four strains of Lactobacillus. Three of the strains used proved to induce chronic arthritis in the rat; all were Lactobacillus casei. The cell wall of Lactobacillus fermentum did not induce chronic arthritis. All arthritogenic bacterial cell walls had the same peptidoglycan structure, whereas that of L. fermentum was different. Likewise, all arthritogenic cell walls...

  20. Human Cytomegalovirus Nuclear Egress Proteins Ectopically Expressed in the Heterologous Environment of Plant Cells are Strictly Targeted to the Nuclear Envelope

    Science.gov (United States)

    Lamm, Christian E.; Link, Katrin; Wagner, Sabrina; Milbradt, Jens; Marschall, Manfred; Sonnewald, Uwe

    2016-01-01

    In all eukaryotic cells, the nucleus forms a prominent cellular compartment containing the cell’s nuclear genome. Although structurally similar, animal and plant nuclei differ substantially in details of their architecture. One example is the nuclear lamina, a layer of tightly interconnected filament proteins (lamins) underlying the nuclear envelope of metazoans. So far no orthologous lamin genes could be detected in plant genomes and putative lamin-like proteins are only poorly described in plants. To probe for potentially conserved features of metazoan and plant nuclear envelopes, we ectopically expressed the core nuclear egress proteins of human cytomegalovirus pUL50 and pUL53 in plant cells. pUL50 localizes to the inner envelope of metazoan nuclei and recruits the nuclear localized pUL53 to it, forming heterodimers. Upon expression in plant cells, a very similar localization pattern of both proteins could be determined. Notably, pUL50 is specifically targeted to the plant nuclear envelope in a rim-like fashion, a location to which coexpressed pUL53 becomes strictly corecruited from its initial nucleoplasmic distribution. Using pUL50 as bait in a yeast two-hybrid screening, the cytoplasmic re-initiation supporting protein RISP could be identified. Interaction of pUL50 and RISP could be confirmed by coexpression and coimmunoprecipitation in mammalian cells and by confocal laser scanning microscopy in plant cells, demonstrating partial pUL50-RISP colocalization in areas of the nuclear rim and other intracellular compartments. Thus, our study provides strong evidence for conserved structural features of plant and metazoan nuclear envelops and identifies RISP as a potential pUL50-interacting plant protein. PMID:26978388

  1. Human Cytomegalovirus Nuclear Egress Proteins Ectopically Expressed in the Heterologous Environment of Plant Cells are Strictly Targeted to the Nuclear Envelope

    Directory of Open Access Journals (Sweden)

    Christian E. Lamm

    2016-03-01

    Full Text Available In all eukaryotic cells, the nucleus forms a prominent cellular compartment containing the cell’s nuclear genome. Although structurally similar, animal and plant nuclei differ substantially in details of their architecture. One example is the nuclear lamina, a layer of tightly interconnected filament proteins (lamins underlying the nuclear envelope of metazoans. So far no orthologous lamin genes could be detected in plant genomes and putative lamin-like proteins are only poorly described in plants. To probe for potentially conserved features of metazoan and plant nuclear envelopes, we ectopically expressed the core nuclear egress proteins of human cytomegalovirus pUL50 and pUL53 in plant cells. pUL50 localizes to the inner envelope of metazoan nuclei and recruits the nuclear localized pUL53 to it, forming heterodimers. Upon expression in plant cells, a very similar localization pattern of both proteins could be determined. Notably, pUL50 is specifically targeted to the plant nuclear envelope in a rim-like fashion, a location to which coexpressed pUL53 becomes strictly corecruited from its initial nucleoplasmic distribution. Using pUL50 as bait in a yeast two-hybrid screening, the cytoplasmic re-initiation supporting protein RISP could be identified. Interaction of pUL50 and RISP could be confirmed by coexpression and coimmunoprecipitation in mammalian cells and by confocal laser scanning microscopy in plant cells, demonstrating partial pUL50-RISP colocalization in areas of the nuclear rim and other intracellular compartments. Thus, our study provides strong evidence for conserved structural features of plant and metazoan nuclear envelops and identifies RISP as a potential pUL50-interacting plant protein.

  2. Infection of human and non-human cells by a highly fusogenic primary CD4-independent HIV-1 isolate with a truncated envelope cytoplasmic tail

    International Nuclear Information System (INIS)

    Truncation of the envelope cytoplasmic tail has enabled FIV, SIV, and some laboratory HIV-1 strains to acquire broader cellular tropism and enhanced fusogenicity. Here we have characterized a primary CD4-independent HIV-1 isolate (92UG046-T8) with a truncated cytoplasmic tail that was able to infect and induce syncytia in primary lymphocytes from human, chimpanzee, and monkey, as well as CD4-negative cell lines from human and monkey. Increased syncytia were also noticeable with 293 cells expressing the cloned envelope from the 92UG046-T8 isolate suggesting envelope-mediated cellular fusion. Except pooled serum from HIV-1-infected individuals, monoclonal anti-envelope antibodies or antibodies/antagonists against CD4, CXCR4, and CCR5 were not able to prevent infection by the 92UG046-T8 isolate. This is the first report showing a primary HIV-1 variant with truncated cytoplasmic tail which is highly fusogenic and can infect a broad range of cells from human and non-human origins. In vivo evolution of similar HIV-1 mutants may have important implications in AIDS pathogenesis

  3. Infection of human and non-human cells by a highly fusogenic primary CD4-independent HIV-1 isolate with a truncated envelope cytoplasmic tail.

    Science.gov (United States)

    Saha, Kunal; Yan, Hui; Nelson, Julie A E; Zerhouni-Layachi, Bouchra

    2005-06-20

    Truncation of the envelope cytoplasmic tail has enabled FIV, SIV, and some laboratory HIV-1 strains to acquire broader cellular tropism and enhanced fusogenicity. Here we have characterized a primary CD4-independent HIV-1 isolate (92UG046-T8) with a truncated cytoplasmic tail that was able to infect and induce syncytia in primary lymphocytes from human, chimpanzee, and monkey, as well as CD4-negative cell lines from human and monkey. Increased syncytia were also noticeable with 293 cells expressing the cloned envelope from the 92UG046-T8 isolate suggesting envelope-mediated cellular fusion. Except pooled serum from HIV-1-infected individuals, monoclonal anti-envelope antibodies or antibodies/antagonists against CD4, CXCR4, and CCR5 were not able to prevent infection by the 92UG046-T8 isolate. This is the first report showing a primary HIV-1 variant with truncated cytoplasmic tail which is highly fusogenic and can infect a broad range of cells from human and non-human origins. In vivo evolution of similar HIV-1 mutants may have important implications in AIDS pathogenesis. PMID:15914218

  4. Applying Data Envelopment Analysis to Evaluation of Taiwanese Solar Cell Industry Operational Performance

    OpenAIRE

    Hao-En Chueh; Jie-Yi Jheng

    2012-01-01

    In the Taiwanese solar power industry, the upstream industry’s lack of silicon raw materials and thedownstream’s underdeveloped systematic manufacturing status have led industrial development to continueconcentrating on cell and module research and development manufacturing and production. Taiwan’ssolar power industry has developed midstream cell manufacturers holding a share of the global market.The research period for this study was between 2010 and 2011. This study constructed a performanc...

  5. Influence of Disulfide-Stabilized Structure on the Specificity of Helper T-Cell and Antibody Responses to HIV Envelope Glycoprotein gp120▿ †

    OpenAIRE

    Mirano-Bascos, Denise; Steede, N. Kalaya; Robinson, James E.; Landry, Samuel J.

    2010-01-01

    CD4+ helper T cells specific for human immunodeficiency virus type 1 (HIV-1) are associated with control of viremia. Nevertheless, vaccines have had limited effectiveness thus far, in part because sequence variability and other structural features of the HIV envelope glycoprotein deflect the immune response. Previous studies indicated that CD4+ T-cell epitope dominance is controlled by antigen three-dimensional structure through its influence on antigen processing and presentation. In this wo...

  6. Expression of particulate-form of Japanese encephalitis virus envelope protein in a stably transfected Drosophila cell line

    Directory of Open Access Journals (Sweden)

    Zhang Li

    2007-02-01

    Full Text Available Abstract Background Japanese encephalitis virus (JEV, a member of the family Flaviviridae, is an important mosquito-borne human pathogen. Its envelope glycoprotein (E is the major determinant of the pathogenicity and host immune responses. In the present study, we explored the feasibility of producing recombinant JEV E protein in the virus-free Drosophila expression system. Results The coding sequence for the signal sequence of premembrane and E protein was cloned into the Drosophila expression vector pAc5.1/V5-His. A Drosophila cell line S2 was cotransfected with this construct as well as a plasmid providing hygromycin B resistance. A cell line expressing the JEV E protein was selected by immunofluoresence, confocal microscopy, and western blot analysis using three different monoclonal antibodies directed against JEV E protein. This cell line was stable in the yield of JEV E protein during two months in vitro maintenance in the presence of hygromycin B. The results showed that the recombinant E protein had an expected molecular weight of about 50 kilodalton, was immunoreactive with all three monoclonal antibodies, and found in both the cytoplasm and culture supernatant. Sucrose gradient ultracentrifugation analysis revealed that the secreted E protein product was in a particulate form. It migrated to the sucrose fraction with a density of 1.13 g/ml. Balb/c mice immunised with the sucrose fraction containing the E protein particles developed specific antibodies. These data show that functioning JEV E protein was expressed in the stable S2 cell line. Conclusion The Drosophila expression system is a more convenient, cheaper and safer approach to the production of vaccine candidates and diagnostic reagents for JEV.

  7. Positive selection of mutants with cell envelope defects of a Salmonella typhimurium strain hypersensitive to the products of genes hisF and hisH

    International Nuclear Information System (INIS)

    Strain SB564 and its derivative DA78 are hypersensitive to the inhibitory action of the proteins coded for by genes hisF and hisH on cell division. Transduction of hisO1242, a regulatory mutation that elicits a very high level of expression of the histidine operon, into these strains resulted in the production of long filamentous cells carrying large balloons and in growth failure. Forty-one hisO1242 derivatives that escaped inhibition were isolated. These strains showed a large variety of alterations, many of which were related to the cell envelope. The more-frequent alterations included: changes in cell shape, increased sensitivity to one or more of several drugs (deoxycholate, cycloserine, penicillin, novobiocin, acridine orange), increased autolytic activity in alkaline buffer, anomalous fermentation of maltose on eosin--methylene blue plates, and temperature-conditional cell division. The alterations are produced, in some of the strains, by pleiotropic mutations in gene envB. Strains affected in divC, divD, and rodA loci have also been identified. Genetic analaysis has shown that several strains carry more than one envelope mutation. It is assumed that envelope mutations are positively selected because they somehow alleviate the particularly severe inhibition of cell division caused, in strains SB564 and DA78, by the excessive synthesis of hisF and hisH gene products

  8. Antigen 43-mediated autotransporter display, a versatile bacterial cell surface presentation system

    DEFF Research Database (Denmark)

    Kjærgaard, Kristian; Hasman, Henrik; Schembri, Mark; Klemm, Per

    2002-01-01

    the outer membrane and secretion through the cell envelope is contained within the protein itself. Ag43 consists of two subunits (alpha and beta), where the beta-subunit forms an integral outer membrane translocator to which the alpha-subunit is noncovalently attached. The simplicity of the Ag43...

  9. Common polysaccharide antigens from the cell envelope of Clostridium perfringens type A.

    OpenAIRE

    Dayalu, K I; Cherniak, R; Hatheway, C L

    1981-01-01

    Soluble antigens were obtained by extracting five serotype strains of Clostridium perfringens type A with water at 100 degrees C. The type-specific polysaccharides were precipitated with ethanol, and the common antigens were recovered from the ethanol supernatants by concentration, dialysis, and lyophilization. Refluxing the water-extracted cell residues with 1% acetic acid followed by concentration, dialysis, and lyophilization gave additional common antigen fractions. A comprehensive, side-...

  10. Construction of an Active Façade Envelope with Peltier Cells

    OpenAIRE

    Martín-Gómez, C. (César); Ibáñez-Puy, M. (María); Sacristán-Fernández, J.A. (José Antonio)

    2013-01-01

    The team researchers have been investigating on alternative ways for buildings to waste less energy. The result is the consecution of a new facade system with Peltier cells, that is to say, a new system of air conditioning that works both as a machine as a facade. That means the application in the field of construction of a technology that is already in use in other areas, fundamentally the military and aerospace. The new system has to be a prefabricated element that perfectly fix between the...

  11. CLA-1 and its splicing variant CLA-2 mediate bacterial adhesion and cytosolic bacterial invasion in mammalian cells

    OpenAIRE

    Vishnyakova, Tatyana G.; Kurlander, Roger; Bocharov, Alexander V.; Baranova, Irina N.; CHEN, ZHIGANG; Abu-Asab, Mones S.; Tsokos, Maria; Malide, Daniela; Basso, Federica; Remaley, Alan; Csako, Gyorgy; Eggerman, Thomas L.; Patterson, Amy P.

    2006-01-01

    CD36 and LIMPII analog 1, CLA-1, and its splicing variant, CLA-2 (SR-BI and SR-BII in rodents), are human high density lipoprotein receptors with an identical extracellular domain which binds a spectrum of ligands including bacterial cell wall components. In this study, CLA-1- and CLA-2-stably transfected HeLa and HEK293 cells demonstrated several-fold increases in the uptake of various bacteria over mock-transfected cells. All bacteria tested, including both Gram-negatives (Escherichia coli ...

  12. Cooperative effects of the human immunodeficiency virus type 1 envelope variable loops V1 and V3 in mediating infectivity for T cells.

    OpenAIRE

    A. Carrillo; Ratner, L

    1996-01-01

    Insertion of T-cell line-tropic V3 and V4 loops from the HXB2 strain into the macrophage-tropic YU-2 envelope resulted in a virus with delayed infectivity for HUT78 and Jurkat cells compared with HXB2. Sequence analysis of viral DNA derived from long-term cultures of Jurkat cells revealed a specific mutation that changed a highly conserved Asn residue in the V1 loop of Env to an Asp residue (N-136-->D). Introduction of this mutation into clones containing a T-cell line-tropic V3 loop, either ...

  13. Resonant mass biosensor for ultrasensitive detection of bacterial cells

    Science.gov (United States)

    Gupta, Amit; Akin, Demir; Bashir, Rashid

    2003-01-01

    This paper describes a surface micromachined cantilever beam based oscillator detector for biological applications. This study used a novel microfabrication technique of merged epitaxial lateral overgrowth (MELO) and chemical mechanical polishing (CMP) to fabricate thin, low stress, single-crystal silicon cantilever beams. Vibration spectra of the cantilever beams, excited by thermal and ambient noise, was measured in air using a Digital Instrument Dimension 3100 Series scanning probe microscope (SPM). The cantilever beams were calibrated by obtaining the spring constant using the added-mass method. The sensors were used to detect the presence of Listeria innocua bacteria by applying increasing concentration of bacteria suspension on the same cantilever beam and measuring the resonant frequency changes in air. Cantilever beams were also used to detect the mass of the adsorbed antibodies and used to show selective capture of bacterial cells. The results indicate that the developed biosensor is capable of rapid and ultra-sensitive detection of bacteria and promises significant potential for enhancement of microbiological research and diagnostics.

  14. Bacterial colonization of host cells in the absence of cholesterol.

    Directory of Open Access Journals (Sweden)

    Stacey D Gilk

    2013-01-01

    Full Text Available Reports implicating important roles for cholesterol and cholesterol-rich lipid rafts in host-pathogen interactions have largely employed sterol sequestering agents and biosynthesis inhibitors. Because the pleiotropic effects of these compounds can complicate experimental interpretation, we developed a new model system to investigate cholesterol requirements in pathogen infection utilizing DHCR24(-/- mouse embryonic fibroblasts (MEFs. DHCR24(-/- MEFs lack the Δ24 sterol reductase required for the final enzymatic step in cholesterol biosynthesis, and consequently accumulate desmosterol into cellular membranes. Defective lipid raft function by DHCR24(-/- MEFs adapted to growth in cholesterol-free medium was confirmed by showing deficient uptake of cholera-toxin B and impaired signaling by epidermal growth factor. Infection in the absence of cholesterol was then investigated for three intracellular bacterial pathogens: Coxiella burnetii, Salmonella enterica serovar Typhimurium, and Chlamydia trachomatis. Invasion by S. Typhimurium and C. trachomatis was unaltered in DHCR24(-/- MEFs. In contrast, C. burnetii entry was significantly decreased in -cholesterol MEFs, and also in +cholesterol MEFs when lipid raft-associated α(Vβ(3 integrin was blocked, suggesting a role for lipid rafts in C. burnetii uptake. Once internalized, all three pathogens established their respective vacuolar niches and replicated normally. However, the C. burnetii-occupied vacuole within DHCR24(-/- MEFs lacked the CD63-positive material and multilamellar membranes typical of vacuoles formed in wild type cells, indicating cholesterol functions in trafficking of multivesicular bodies to the pathogen vacuole. These data demonstrate that cholesterol is not essential for invasion and intracellular replication by S. Typhimurium and C. trachomatis, but plays a role in C. burnetii-host cell interactions.

  15. The bacterial tubulin FtsZ requires its intrinsically disordered linker to direct robust cell wall construction.

    Science.gov (United States)

    Sundararajan, Kousik; Miguel, Amanda; Desmarais, Samantha M; Meier, Elizabeth L; Casey Huang, Kerwyn; Goley, Erin D

    2015-01-01

    The bacterial GTPase FtsZ forms a cytokinetic ring at midcell, recruits the division machinery and orchestrates membrane and peptidoglycan cell wall invagination. However, the mechanism for FtsZ regulation of peptidoglycan metabolism is unknown. The FtsZ GTPase domain is separated from its membrane-anchoring C-terminal conserved (CTC) peptide by a disordered C-terminal linker (CTL). Here we investigate CTL function in Caulobacter crescentus. Strikingly, production of FtsZ lacking the CTL (ΔCTL) is lethal: cells become filamentous, form envelope bulges and lyse, resembling treatment with β-lactam antibiotics. This phenotype is produced by FtsZ polymers bearing the CTC and a CTL shorter than 14 residues. Peptidoglycan synthesis still occurs downstream of ΔCTL; however, cells expressing ΔCTL exhibit reduced peptidoglycan crosslinking and longer glycan strands than wild type. Importantly, midcell proteins are still recruited to sites of ΔCTL assembly. We propose that FtsZ regulates peptidoglycan metabolism through a CTL-dependent mechanism that extends beyond simple protein recruitment. PMID:26099469

  16. Isolated receptor binding domains of HTLV-1 and HTLV-2 envelopes bind Glut-1 on activated CD4+ and CD8+ T cells

    Directory of Open Access Journals (Sweden)

    Montel-Hagen Amélie

    2007-05-01

    Full Text Available Abstract Background We previously identified the glucose transporter Glut-1, a member of the multimembrane-spanning facilitative nutrient transporter family, as a receptor for both HTLV-1 and HTLV-2. However, a recent report concluded that Glut-1 cannot serve as a receptor for HTLV-1 on CD4 T cells: This was based mainly on their inability to detect Glut-1 on this lymphocyte subset using the commercial antibody mAb1418. It was therefore of significant interest to thoroughly assess Glut-1 expression on CD4 and CD8 T cells, and its association with HTLV-1 and -2 envelope binding. Results As previously reported, ectopic expression of Glut-1 but not Glut-3 resulted in significantly augmented binding of tagged proteins harboring the receptor binding domains of either HTLV-1 or HTLV-2 envelope glycoproteins (H1RBD or H2RBD. Using antibodies raised against the carboxy-terminal peptide of Glut-1, we found that Glut-1 expression was significantly increased in both CD4 and CD8 cells following TCR stimulation. Corresponding increases in the binding of H1RBD as well as H2RBD, not detected on quiescent T cells, were observed following TCR engagement. Furthermore, increased Glut-1 expression was accompanied by a massive augmentation in glucose uptake in TCR-stimulated CD4 and CD8 lymphocytes. Finally, we determined that the apparent contradictory results obtained by Takenouchi et al were due to their monitoring of Glut-1 with a mAb that does not bind cells expressing endogenous Glut-1, including human erythrocytes that harbor 300,000 copies per cell. Conclusion Transfection of Glut-1 directly correlates with the capacities of HTLV-1 and HTLV-2 envelope-derived ligands to bind cells. Moreover, Glut-1 is induced by TCR engagement, resulting in massive increases in glucose uptake and binding of HTLV-1 and -2 envelopes to both CD4 and CD8 T lymphocytes. Therefore, Glut-1 is a primary binding receptor for HTLV-1 and HTLV-2 envelopes on activated CD4 as well as CD8

  17. EEVD motif of heat shock cognate protein 70 contributes to bacterial uptake by trophoblast giant cells

    Directory of Open Access Journals (Sweden)

    Kim Suk

    2009-12-01

    Full Text Available Abstract Background The uptake of abortion-inducing pathogens by trophoblast giant (TG cells is a key event in infectious abortion. However, little is known about phagocytic functions of TG cells against the pathogens. Here we show that heat shock cognate protein 70 (Hsc70 contributes to bacterial uptake by TG cells and the EEVD motif of Hsc70 plays an important role in this. Methods Brucella abortus and Listeria monocytogenes were used as the bacterial antigen in this study. Recombinant proteins containing tetratricopeptide repeat (TPR domains were constructed and confirmation of the binding capacity to Hsc70 was assessed by ELISA. The recombinant TPR proteins were used for investigation of the effect of TPR proteins on bacterial uptake by TG cells and on pregnancy in mice. Results The monoclonal antibody that inhibits bacterial uptake by TG cells reacted with the EEVD motif of Hsc70. Bacterial TPR proteins bound to the C-terminal of Hsc70 through its EEVD motif and this binding inhibited bacterial uptake by TG cells. Infectious abortion was also prevented by blocking the EEVD motif of Hsc70. Conclusions Our results demonstrate that surface located Hsc70 on TG cells mediates the uptake of pathogenic bacteria and proteins containing the TPR domain inhibit the function of Hsc70 by binding to its EEVD motif. These molecules may be useful in the development of methods for preventing infectious abortion.

  18. STD NMR spectroscopy: a case study of fosfomycin binding interactions in living bacterial cells

    Energy Technology Data Exchange (ETDEWEB)

    Milagre, Cintia D.F.; Cabeca, Luis Fernando; Martins, Lucas G.; Marsaioli, Anita J., E-mail: anita@iq [Universidade Estadual de Campinas (IQ/UNICAMP), SP (Brazil). Inst. de Quimica

    2011-07-01

    A saturation transfer difference (STD) NMR experiment was successfully employed to observe the binding interactions of fosfomycin resistant and non-resistant bacterial strains using living cell suspensions, without the need for isotopic labelling of the ligand or receptor. (author)

  19. Facile method to stain the bacterial cell surface for super-resolution fluorescence microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Gunsolus, Ian L.; Hu, Dehong; Mihai, Cosmin; Lohse, Samuel E.; Lee, Chang-Soo; Torelli, Marco; Hamers, Robert J.; Murphy, Catherine; Orr, Galya; Haynes, Christy L.

    2014-01-01

    A method to fluorescently stain the surfaces of both Gram-negative and Gram-positive bacterial cells compatible with super-resolution fluorescence microscopy is presented. This method utilizes a commercially-available fluorescent probe to label primary amines at the surface of the cell. We demonstrate efficient staining of two bacterial strains, the Gram-negative Shewanella oneidensis MR-1 and the Gram-positive Bacillus subtilis 168. Using structured illumination microscopy and stochastic optical reconstruction microscopy, which require high quantum yield or specialized dyes, we show that this staining method may be used to resolve the bacterial cell surface with sub-diffraction-limited resolution. We further use this method to identify localization patterns of nanomaterials, specifically cadmium selenide quantum dots, following interaction with bacterial cells.

  20. Phase Diagram of Collective Motion of Bacterial Cells in a Shallow Circular Pool

    CERN Document Server

    Wakita, Jun-ichi; Yamamoto, Ken; Katori, Makoto; Yamada, Yasuyuki

    2015-01-01

    The collective motion of bacterial cells in a shallow circular pool is systematically studied using the bacterial species $Bacillus$ $subtilis$. The ratio of cell length to pool diameter (i.e., the reduced cell length) ranges from 0.06 to 0.43 in our experiments. Bacterial cells in a circular pool show various types of collective motion depending on the cell density in the pool and the reduced cell length. The motion is classified into six types, which we call random motion, turbulent motion, one-way rotational motion, two-way rotational motion, random oscillatory motion, and ordered oscillatory motion. Two critical values of reduced cell lengths are evaluated, at which drastic changes in collective motion are induced. A phase diagram is proposed in which the six phases are arranged.

  1. Insights into Substrate Specificity of NlpC/P60 Cell Wall Hydrolases Containing Bacterial SH3 Domains

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Qingping; Mengin-Lecreulx, Dominique; Liu, Xueqian W.; Patin, Delphine; Farr, Carol L.; Grant, Joanna C.; Chiu, Hsiu-Ju; Jaroszewski, Lukasz; Knuth, Mark W.; Godzik, Adam; Lesley, Scott A.; Elsliger, Marc-André; Deacon, Ashley M.; Wilson, Ian A.

    2015-09-15

    ABSTRACT

    Bacterial SH3 (SH3b) domains are commonly fused with papain-like Nlp/P60 cell wall hydrolase domains. To understand how the modular architecture of SH3b and NlpC/P60 affects the activity of the catalytic domain, three putative NlpC/P60 cell wall hydrolases were biochemically and structurally characterized. These enzymes all have γ-d-Glu-A2pm (A2pm is diaminopimelic acid) cysteine amidase (ordl-endopeptidase) activities but with different substrate specificities. One enzyme is a cell wall lysin that cleaves peptidoglycan (PG), while the other two are cell wall recycling enzymes that only cleave stem peptides with an N-terminall-Ala. Their crystal structures revealed a highly conserved structure consisting of two SH3b domains and a C-terminal NlpC/P60 catalytic domain, despite very low sequence identity. Interestingly, loops from the first SH3b domain dock into the ends of the active site groove of the catalytic domain, remodel the substrate binding site, and modulate substrate specificity. Two amino acid differences at the domain interface alter the substrate binding specificity in favor of stem peptides in recycling enzymes, whereas the SH3b domain may extend the peptidoglycan binding surface in the cell wall lysins. Remarkably, the cell wall lysin can be converted into a recycling enzyme with a single mutation.

    IMPORTANCEPeptidoglycan is a meshlike polymer that envelops the bacterial plasma membrane and bestows structural integrity. Cell wall lysins and recycling enzymes are part of a set of lytic enzymes that target covalent bonds connecting the amino acid and amino sugar building blocks of the PG network. These hydrolases are involved in processes such as cell growth and division, autolysis, invasion, and PG turnover and recycling. To avoid cleavage of unintended substrates, these enzymes have very selective substrate specificities. Our biochemical and structural

  2. Nuclear envelope-localized EGF family protein amphiregulin activates breast cancer cell migration in an EGF-like domain independent manner

    International Nuclear Information System (INIS)

    Highlights: ► Nuclear envelope-localized proAREG activates cancer cell migration via its cytoplasmic domain. ► The induction of cell migration does not require the EGF-like domain or EGR function. ► Nuclear envelope-localized proAREG suppresses breast cancer cell growth without EGFR function. ► This study revealed a novel function mediated by the intracellular domain of proAREG. -- Abstract: Amphiregulin (AREG), an EGF family protein, is synthesized as a type I transmembrane precursor (proAREG) and expressed on the cell surface with an extracellular EGF-like domain and an intracellular short cytoplasmic tail. The ectodomain shedding yields a soluble EGF receptor ligand (soluble AREG) which binds to EGF receptor (EGFR) and concomitantly induces migration of unshed proAREG from the plasma membrane to the nuclear envelope (NE). AREG is known to play a potential role in breast cancer and has been intensively investigated as an EGF receptor ligand, while the function of the NE-localized proAREG remains unknown. In this study we used a truncated mutant that mimics NE-localized proAREG without shedding stimuli to discriminate between the functions of NE-localized and plasma membrane-localized proAREG and demonstrate that NE-localized proAREG activates breast cancer cell migration, but suppresses cell growth. Moreover, the present study shows that induction of cell migration by NE-localized proAREG does not require the extracellular growth factor domain or EGF receptor function. Collectively these data demonstrate a novel function mediated by the intracellular domain of proAREG and suggest a significant role for NE-localized proAREG in driving human breast cancer progression.

  3. Room temperature electrocompetent bacterial cells improve DNA transformation and recombineering efficiency

    OpenAIRE

    Qiang Tu; Jia Yin; Jun Fu; Jennifer Herrmann; Yuezhong Li; Yulong Yin; Francis Stewart, A.; Rolf Müller; Youming Zhang

    2016-01-01

    Bacterial competent cells are essential for cloning, construction of DNA libraries, and mutagenesis in every molecular biology laboratory. Among various transformation methods, electroporation is found to own the best transformation efficiency. Previous electroporation methods are based on washing and electroporating the bacterial cells in ice-cold condition that make them fragile and prone to death. Here we present simple temperature shift based methods that improve DNA transformation and re...

  4. Revealing fosfomycin primary effect on Staphylococcus aureus transcriptome: modulation of cell envelope biosynthesis and phosphoenolpyruvate induced starvation

    Directory of Open Access Journals (Sweden)

    Gruden Kristina

    2010-06-01

    Full Text Available Abstract Background Staphylococcus aureus is a highly adaptable human pathogen and there is a constant search for effective antibiotics. Fosfomycin is a potent irreversible inhibitor of MurA, an enolpyruvyl transferase that uses phosphoenolpyruvate as substrate. The goal of this study was to identify the pathways and processes primarily affected by fosfomycin at the genome-wide transcriptome level to aid development of new drugs. Results S. aureus ATCC 29213 cells were treated with sub-MIC concentrations of fosfomycin and harvested at 10, 20 and 40 minutes after treatment. S. aureus GeneChip statistical data analysis was complemented by gene set enrichment analysis. A visualization tool for mapping gene expression data into biological pathways was developed in order to identify the metabolic processes affected by fosfomycin. We have shown that the number of significantly differentially expressed genes in treated cultures increased with time and with increasing fosfomycin concentration. The target pathway - peptidoglycan biosynthesis - was upregulated following fosfomycin treatment. Modulation of transport processes, cofactor biosynthesis, energy metabolism and nucleic acid biosynthesis was also observed. Conclusions Several pathways and genes downregulated by fosfomycin have been identified, in contrast to previously described cell wall active antibiotics, and was explained by starvation response induced by phosphoenolpyruvate accumulation. Transcriptomic profiling, in combination with meta-analysis, has been shown to be a valuable tool in determining bacterial response to a specific antibiotic.

  5. Plectasin, a Fungal Defensin, Targets the Bacterial Cell Wall Precursor Lipid II

    DEFF Research Database (Denmark)

    Schneider, Tanja; Kruse, Thomas; Wimmer, Reinhard;

    2010-01-01

    plectasin, a fungal defensin, acts by directly binding the bacterial cell-wall precursor Lipid II. A wide range of genetic and biochemical approaches identify cell-wall biosynthesis as the pathway targeted by plectasin. In vitro assays for cell-wall synthesis identified Lipid II as the specific cellular...

  6. Toxicity of a polymer-graphene oxide composite against bacterial planktonic cells, biofilms, and mammalian cells

    Science.gov (United States)

    Mejías Carpio, Isis E.; Santos, Catherine M.; Wei, Xin; Rodrigues, Debora F.

    2012-07-01

    It is critical to develop highly effective antimicrobial agents that are not harmful to humans and do not present adverse effects on the environment. Although antimicrobial studies of graphene-based nanomaterials are still quite limited, some researchers have paid particular attention to such nanocomposites as promising candidates for the next generation of antimicrobial agents. The polyvinyl-N-carbazole (PVK)-graphene oxide (GO) nanocomposite (PVK-GO), which contains only 3 wt% of GO well-dispersed in a 97 wt% PVK matrix, presents excellent antibacterial properties without significant cytotoxicity to mammalian cells. The high polymer content in this nanocomposite makes future large-scale material manufacturing possible in a high-yield process of adiabatic bulk polymerization. In this study, the toxicity of PVK-GO was assessed with planktonic microbial cells, biofilms, and NIH 3T3 fibroblast cells. The antibacterial effects were evaluated against two Gram-negative bacteria: Escherichia coli and Cupriavidus metallidurans; and two Gram-positive bacteria: Bacillus subtilis and Rhodococcus opacus. The results show that the PVK-GO nanocomposite presents higher antimicrobial effects than the pristine GO. The effectiveness of the PVK-GO in solution was demonstrated as the nanocomposite ``encapsulated'' the bacterial cells, which led to reduced microbial metabolic activity and cell death. The fact that the PVK-GO did not present significant cytotoxicity to fibroblast cells offers a great opportunity for potential applications in important biomedical and industrial fields.It is critical to develop highly effective antimicrobial agents that are not harmful to humans and do not present adverse effects on the environment. Although antimicrobial studies of graphene-based nanomaterials are still quite limited, some researchers have paid particular attention to such nanocomposites as promising candidates for the next generation of antimicrobial agents. The polyvinyl

  7. Hepatitis B synthetic immunogen comprised of nucleocapsid T-cell sites and an envelope B-cell epitope.

    OpenAIRE

    Milich, D R; Hughes, J L; A. McLachlan; Thornton, G B; Moriarty, A.

    1988-01-01

    Previous studies located T-cell recognition of the nucleocapsid of the hepatitis B virus (HBcAg) to residues 120-140 in mice bearing the H-2s or H-2b haplotypes. Herein, we demonstrate that B10.S (H-2s) and B10 (H-2b) H-2 congenic strains recognize distinct T-cell sites within the p120-140 (a synthetic peptide corresponding to residues 120-140 of HBcAg) sequence defined by p120-131 and p129-140, respectively. Peptide p120-131 stimulates B10.S HBcAg-primed T cells, and reciprocally p120-131-pr...

  8. IN VITRO INDUCTION OF BLOOD MONONUCLEAR CELL PROLIFERATION BY ENDOGENOUS RETROVIRAL HERV-Eλ4-1 ENVELOPE PEPTIDE IN PATIENTS WITH MULTIPLE SCLEROSIS

    Directory of Open Access Journals (Sweden)

    A. A. Smagin

    2014-08-01

    Full Text Available A comparative in vitro study of blood mononuclear cells from multiple sclerosis patients and healthy donors was performed, in order to evaluate proliferative response to a retroviral antigen, aiming to determine immunomodulatory properties of synthetic oligopeptide homologous to a highly conserved human endogenous retrovirus HERV-Eλ4-1 envelope protein. It was revealed that this oligopeptide is able to stimulate the in vitro spontaneous and mitogen-induced proliferation of blood mononuclear cells from either donor and multiple sclerosis patients. Intensity of this oligopeptide-induced stimulatory effect depends on the protein concentration, and on initial level of blood immunocompetent cells proliferation. Hence, the endogenous retrovirus HERV-Eλ4-1 envelope region protein is able to increase functional activity of immunocompetent cells from human blood, that suggesting its immunostimulatory properties. It is possible that the mitogenic effects of this protein upon immunocompetent cells of multiple sclerosis patients represent a potential mechanism of retroviral involvement into pathogenesis of the disorder.

  9. Identification of individual biofilm-forming bacterial cells using Raman tweezers

    Science.gov (United States)

    Samek, Ota; Bernatová, Silvie; Ježek, Jan; Šiler, Martin; Šerý, Mojmir; Krzyžánek, Vladislav; Hrubanová, Kamila; Zemánek, Pavel; Holá, Veronika; Růžička, Filip

    2015-05-01

    A method for in vitro identification of individual bacterial cells is presented. The method is based on a combination of optical tweezers for spatial trapping of individual bacterial cells and Raman microspectroscopy for acquisition of spectral "Raman fingerprints" obtained from the trapped cell. Here, Raman spectra were taken from the biofilm-forming cells without the influence of an extracellular matrix and were compared with biofilm-negative cells. Results of principal component analyses of Raman spectra enabled us to distinguish between the two strains of Staphylococcus epidermidis. Thus, we propose that Raman tweezers can become the technique of choice for a clearer understanding of the processes involved in bacterial biofilms which constitute a highly privileged way of life for bacteria, protected from the external environment.

  10. Cell order in bacterial swarms arises from reversals of moving direction

    Science.gov (United States)

    Wu, Yilin; Jiang, Yi; Kaiser, Dale; Alber, Mark

    2010-03-01

    Bacterial swarms are a beautiful example of the emergent behavior of systems of self-propelled rods. In swarming rod-shaped bacteria cells move smoothly even though they are packed together in high density. Experimental evidence shows that long-distance signaling is not required for bacterial swarming. It naturally raises the question how a swarm develops its order. Using a biomechanical model, we show here that regular periodic reversals of gliding direction in general systems of self-propelled rod shaped bacteria can lead to the extensive ordering of cells. We also show that an optimal reversal period and an optimal cell shape exist for producing such order. Given the observations of reversing behavior in several bacterial species,we suggest that the capacity to swarm depends less on the motility engine employed by individual cells, but more on the behavioral algorithm that enhances the flow of densely packed cells near the swarming edge.

  11. Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

    International Nuclear Information System (INIS)

    Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants

  12. Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Roehrig, John T., E-mail: jtr1@cdc.gov [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Butrapet, Siritorn; Liss, Nathan M. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Bennett, Susan L. [Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States); Luy, Betty E.; Childers, Thomas; Boroughs, Karen L.; Stovall, Janae L.; Calvert, Amanda E. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States); Blair, Carol D. [Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States); Huang, Claire Y.-H. [Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States)

    2013-07-05

    Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants.

  13. Transcriptional activity around bacterial cell death reveals molecular biomarkers for cell viability

    Directory of Open Access Journals (Sweden)

    Schuren Frank H

    2008-12-01

    Full Text Available Abstract Background In bacteriology, the ability to grow in selective media and to form colonies on nutrient agar plates is routinely used as a retrospective criterion for the detection of living bacteria. However, the utilization of indicators for bacterial viability-such as the presence of specific transcripts or membrane integrity-would overcome bias introduced by cultivation and reduces the time span of analysis from initiation to read out. Therefore, we investigated the correlation between transcriptional activity, membrane integrity and cultivation-based viability in the Gram-positive model bacterium Bacillus subtilis. Results We present microbiological, cytological and molecular analyses of the physiological response to lethal heat stress under accurately defined conditions through systematic sampling of bacteria from a single culture exposed to gradually increasing temperatures. We identified a coherent transcriptional program including known heat shock responses as well as the rapid expression of a small number of sporulation and competence genes, the latter only known to be active in the stationary growth phase. Conclusion The observed coordinated gene expression continued even after cell death, in other words after all bacteria permanently lost their ability to reproduce. Transcription of a very limited number of genes correlated with cell viability under the applied killing regime. The transcripts of the expressed genes in living bacteria – but silent in dead bacteria-include those of essential genes encoding chaperones of the protein folding machinery and can serve as molecular biomarkers for bacterial cell viability.

  14. Molecular Architecture of the Bacterial Flagellar Motor in Cells

    OpenAIRE

    Zhao, Xiaowei; Norris, Steven J; Liu, Jun

    2014-01-01

    The flagellum is one of the most sophisticated self-assembling molecular machines in bacteria. Powered by the proton-motive force, the flagellum rapidly rotates in either a clockwise or counterclockwise direction, which ultimately controls bacterial motility and behavior. Escherichia coli and Salmonella enterica have served as important model systems for extensive genetic, biochemical, and structural analysis of the flagellum, providing unparalleled insights into its structure, function, and ...

  15. Enhanced Toxic Metal Accumulation in Engineered Bacterial Cells Expressing Arabidopsis thaliana Phytochelatin Synthase

    OpenAIRE

    Sauge-Merle, Sandrine; Cuiné, Stéphan; Carrier, Patrick; Lecomte-Pradines, Catherine; Luu, Doan-Trung; Peltier, Gilles

    2003-01-01

    Phytochelatins (PCs) are metal-binding cysteine-rich peptides, enzymatically synthesized in plants and yeasts from glutathione in response to heavy metal stress by PC synthase (EC 2.3.2.15). In an attempt to increase the ability of bacterial cells to accumulate heavy metals, the Arabidopsis thaliana gene encoding PC synthase (AtPCS) was expressed in Escherichia coli. A marked accumulation of PCs was observed in vivo together with a decrease in the glutathione cellular content. When bacterial ...

  16. Type 1 pilus-mediated bacterial invasion of bladder epithelial cells

    OpenAIRE

    Martinez, Juan J.; Mulvey, Matthew A.; Schilling, Joel D.; Pinkner, Jerome S.; Hultgren, Scott J.

    2000-01-01

    Most strains of uropathogenic Escherichia coli (UPEC) encode filamentous adhesive organelles called type 1 pili. We have determined that the type 1 pilus adhesin, FimH, mediates not only bacterial adherence, but also invasion of human bladder epithelial cells. In contrast, adherence mediated by another pilus adhesin, PapG, did not initiate bacterial internalization. FimH-mediated invasion required localized host actin reorganization, phosphoinositide 3-kinase (PI 3-kinase) activation and host...

  17. TLR4-dependent hepcidin expression by myeloid cells in response to bacterial pathogens

    OpenAIRE

    Peyssonnaux, Carole; Zinkernagel, Annelies S.; Datta, Vivekanand; Lauth, Xavier; Johnson, Randall S; Nizet, Victor

    2006-01-01

    Hepcidin is an antimicrobial peptide secreted by the liver during inflammation that plays a central role in mammalian iron homeostasis. Here we demonstrate the endogenous expression of hepcidin by macrophages and neutrophils in vitro and in vivo. These myeloid cell types produced hepcidin in response to bacterial pathogens in a toll-like receptor 4 (TLR4)-dependent fashion. Conversely, bacterial stimulation of macrophages triggered a TLR4-dependent reduction in the iron exporter ferroportin. ...

  18. Single-molecule investigations of the stringent response machinery in living bacterial cells

    OpenAIRE

    English, Brian P.; Hauryliuk, Vasili; Sanamrad, Arash; Tankov, Stoyan; Dekker, Nynke H.; Elf, Johan

    2011-01-01

    The RelA-mediated stringent response is at the heart of bacterial adaptation to starvation and stress, playing a major role in the bacterial cell cycle and virulence. RelA integrates several environmental cues and synthesizes the alarmone ppGpp, which globally reprograms transcription, translation, and replication. We have developed and implemented novel single-molecule tracking methodology to characterize the intracellular catalytic cycle of RelA. Our single-molecule experiments show that Re...

  19. A simple and novel modification of comet assay for determination of bacteriophage mediated bacterial cell lysis.

    Science.gov (United States)

    Khairnar, Krishna; Sanmukh, Swapnil; Chandekar, Rajshree; Paunikar, Waman

    2014-07-01

    The comet assay is the widely used method for in vitro toxicity testing which is also an alternative to the use of animal models for in vivo testing. Since, its inception in 1984 by Ostling and Johansson, it is being modified frequently for a wide range of application. In spite of its wide applicability, unfortunately there is no report of its application in bacteriophages research. In this study, a novel application of comet assay for the detection of bacteriophage mediated bacterial cell lysis was described. The conventional methods in bacteriophage research for studying bacterial lysis by bacteriophages are plaque assay method. It is time consuming, laborious and costly. The lytic activity of bacteriophage devours the bacterial cell which results in the release of bacterial genomic material that gets detected by ethidium bromide staining method by the comet assay protocol. The objective of this study was to compare efficacy of comet assay with different assay used to study phage mediated bacterial lysis. The assay was performed on culture isolates (N=80 studies), modified comet assay appear to have relatively higher sensitivity and specificity than other assay. The results of the study showed that the application of comet assay can be an economical, time saving and less laborious alternative to conventional plaque assay for the detection of bacteriophage mediated bacterial cell lysis. PMID:24681053

  20. Selective Removal of DNA from Dead Cells of Mixed Bacterial Communities by Use of Ethidium Monoazide

    OpenAIRE

    Nocker, Andreas; Camper, Anne K.

    2006-01-01

    The distinction between viable and dead bacterial cells poses a major challenge in microbial diagnostics. Due to the persistence of DNA in the environment after cells have lost viability, DNA-based quantification methods overestimate the number of viable cells in mixed populations or even lead to false-positive results in the absence of viable cells. On the other hand, RNA-based diagnostic methods, which circumvent this problem, are technically demanding and suffer from some drawbacks. A prom...

  1. Bacterial ‘Cell’ Phones: Do cell phones carry potential pathogens?

    OpenAIRE

    Kiran Chawla; Chiranjay Mukhopadhayay; Bimala Gurung; Priya Bhate; Indira Bairy

    2009-01-01

    Cell phones are important companions for professionals especially health care workers (HCWs) for better communication in hospital. The present study compared the nature of the growth of potentially pathogenic bacterial flora on cell phones in hospital and community. 75% cell phones from both the categories grew at least one potentially pathogenic organism. Cell phones from HCWs grew significantly more potential pathogens like MRSA (20%), Acinetobacter species (5%), Pseudomonas species (2.5%) ...

  2. Regulation of the immune response to bacterial lipopolysaccharide by adherent cells.

    OpenAIRE

    Citron, M O; Michael, J G

    1981-01-01

    Immune response to bacterial lipopolysaccharide is usually short lived, but it often reappears without additional stimulus in a cyclic fashion. Activated adherent cells, presumably macrophages, were found to have a role in the reduction of the immune response to Escherichia coli O127 lipopolysaccharide. The suppressive activity of the adherent cells was abrogated before renewal of the responsiveness.

  3. Microspectrometric insights on the uptake of antibiotics at the single bacterial cell level

    Science.gov (United States)

    Cinquin, Bertrand; Maigre, Laure; Pinet, Elizabeth; Chevalier, Jacqueline; Stavenger, Robert A.; Mills, Scott; Réfrégiers, Matthieu; Pagès, Jean-Marie

    2015-01-01

    Bacterial multidrug resistance is a significant health issue. A key challenge, particularly in Gram-negative antibacterial research, is to better understand membrane permeation of antibiotics in clinically relevant bacterial pathogens. Passing through the membrane barrier to reach the required concentration inside the bacterium is a pivotal step for most antibacterials. Spectrometric methodology has been developed to detect drugs inside bacteria and recent studies have focused on bacterial cell imaging. Ultimately, we seek to use this method to identify pharmacophoric groups which improve penetration, and therefore accumulation, of small-molecule antibiotics inside bacteria. We developed a method to quantify the time scale of antibiotic accumulation in living bacterial cells. Tunable ultraviolet excitation provided by DISCO beamline (synchrotron Soleil) combined with microscopy allows spectroscopic analysis of the antibiotic signal in individual bacterial cells. Robust controls and measurement of the crosstalk between fluorescence channels can provide real time quantification of drug. This technique represents a new method to assay drug translocation inside the cell and therefore incorporate rational drug design to impact antibiotic uptake. PMID:26656111

  4. Probing living bacterial adhesion by single cell force spectroscopy using atomic force microscopy

    DEFF Research Database (Denmark)

    Zeng, Guanghong; Ogaki, Ryosuke; Regina, Viduthalai R.; Müller, Torsten; Meyer, Rikke Louise

    (ethylene glycol) (PEG) coatings on titanium. We investigate the ability of a high density poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) coating to resist bacterial adhesion and biofilm formation from three clinically relevant bacteria: Pseudomonas aeruginosa, Staphylococcus aureus and Staphylococcus...... cultures. The high density PLL-g-PEG coatings completely resisted bacterial colonization, whereas conventional coatings couldn’t resist colonization by S. epidermidis. The unique ability of S. epidermidis to colonize conventional PLL-g-PEG coatings was investigated by looking into the composition of S......Bacteria initiate attachment to the surfaces with the aid of different extracellular polymers. To quantitatively study how these polymers mediate bacterial adhesion and possibly their interactions, it is essential to go down to single cell level, with in mind that cell-to-cell variation should be...

  5. Therapeutic envelope vaccination in combination with antiretroviral therapy temporarily rescues SIV-specific CD4⁺ T-cell-dependent natural killer cell effector responses in chronically infected rhesus macaques.

    Science.gov (United States)

    Vargas-Inchaustegui, Diego A; Xiao, Peng; Demberg, Thorsten; Pal, Ranajit; Robert-Guroff, Marjorie

    2015-06-01

    Natural killer (NK) cells are essential components of the immune system, and due to their rapid response potential, can have a great impact during early anti-viral immune responses. We have previously shown that interleukin-2-dependent NK and CD4(+) T-cell co-operative immune responses exist in long-term simian immunodeficiency virus (SIV) -infected controlling macaques and can be rescued in SIV-infected non-controlling macaques by a short course of antiretroviral therapy (ART). Given that co-operative responses may play an important role in disease prevention and therapeutic treatment, in the present study we sought to determine if these responses can be enhanced in chronically SIV-infected macaques by vaccination with a single-dose of envelope protein given during ART. To this end, we treated 14 chronically SIV-infected macaques with ART for 11 weeks and gave 10 of these macaques a single intramuscular dose of SIV gp120 at week 9 of treatment. ART significantly decreased plasma and mucosal viral loads, increased the numbers of circulating CD4(+) T cells in all macaques, and increased T-cell-dependent envelope- and gag-specific interferon-γ and tumour necrosis factor-α production by circulatory CD56(+) NK cells. The therapeutic envelope immunization resulted in higher envelope-specific responses compared with those in macaques that received ART only. Functional T-cell responses restored by ART and therapeutic Env immunization were correlated with transiently reduced plasma viraemia levels following ART release. Collectively our results indicate that SIV-specific T-cell-dependent NK cell responses can be efficiently rescued by ART in chronically SIV-infected macaques and that therapeutic immunization may be beneficial in previously vaccinated individuals. PMID:25626488

  6. Search for MicroRNAs Expressed by Intracellular Bacterial Pathogens in Infected Mammalian Cells

    Science.gov (United States)

    Furuse, Yuki; Finethy, Ryan; Saka, Hector A.; Xet-Mull, Ana M.; Sisk, Dana M.; Smith, Kristen L. Jurcic; Lee, Sunhee; Coers, Jörn; Valdivia, Raphael H.; Tobin, David M.; Cullen, Bryan R.

    2014-01-01

    MicroRNAs are expressed by all multicellular organisms and play a critical role as post-transcriptional regulators of gene expression. Moreover, different microRNA species are known to influence the progression of a range of different diseases, including cancer and microbial infections. A number of different human viruses also encode microRNAs that can attenuate cellular innate immune responses and promote viral replication, and a fungal pathogen that infects plants has recently been shown to express microRNAs in infected cells that repress host cell immune responses and promote fungal pathogenesis. Here, we have used deep sequencing of total expressed small RNAs, as well as small RNAs associated with the cellular RNA-induced silencing complex RISC, to search for microRNAs that are potentially expressed by intracellular bacterial pathogens and translocated into infected animal cells. In the case of Legionella and Chlamydia and the two mycobacterial species M. smegmatis and M. tuberculosis, we failed to detect any bacterial small RNAs that had the characteristics expected for authentic microRNAs, although large numbers of small RNAs of bacterial origin could be recovered. However, a third mycobacterial species, M. marinum, did express an ∼23-nt small RNA that was bound by RISC and derived from an RNA stem-loop with the characteristics expected for a pre-microRNA. While intracellular expression of this candidate bacterial microRNA was too low to effectively repress target mRNA species in infected cultured cells in vitro, artificial overexpression of this potential bacterial pre-microRNA did result in the efficient repression of a target mRNA. This bacterial small RNA therefore represents the first candidate microRNA of bacterial origin. PMID:25184567

  7. Search for microRNAs expressed by intracellular bacterial pathogens in infected mammalian cells.

    Directory of Open Access Journals (Sweden)

    Yuki Furuse

    Full Text Available MicroRNAs are expressed by all multicellular organisms and play a critical role as post-transcriptional regulators of gene expression. Moreover, different microRNA species are known to influence the progression of a range of different diseases, including cancer and microbial infections. A number of different human viruses also encode microRNAs that can attenuate cellular innate immune responses and promote viral replication, and a fungal pathogen that infects plants has recently been shown to express microRNAs in infected cells that repress host cell immune responses and promote fungal pathogenesis. Here, we have used deep sequencing of total expressed small RNAs, as well as small RNAs associated with the cellular RNA-induced silencing complex RISC, to search for microRNAs that are potentially expressed by intracellular bacterial pathogens and translocated into infected animal cells. In the case of Legionella and Chlamydia and the two mycobacterial species M. smegmatis and M. tuberculosis, we failed to detect any bacterial small RNAs that had the characteristics expected for authentic microRNAs, although large numbers of small RNAs of bacterial origin could be recovered. However, a third mycobacterial species, M. marinum, did express an ∼ 23-nt small RNA that was bound by RISC and derived from an RNA stem-loop with the characteristics expected for a pre-microRNA. While intracellular expression of this candidate bacterial microRNA was too low to effectively repress target mRNA species in infected cultured cells in vitro, artificial overexpression of this potential bacterial pre-microRNA did result in the efficient repression of a target mRNA. This bacterial small RNA therefore represents the first candidate microRNA of bacterial origin.

  8. Influence of Disulfide-Stabilized Structure on the Specificity of Helper T-Cell and Antibody Responses to HIV Envelope Glycoprotein gp120▿ †

    Science.gov (United States)

    Mirano-Bascos, Denise; Steede, N. Kalaya; Robinson, James E.; Landry, Samuel J.

    2010-01-01

    CD4+ helper T cells specific for human immunodeficiency virus type 1 (HIV-1) are associated with control of viremia. Nevertheless, vaccines have had limited effectiveness thus far, in part because sequence variability and other structural features of the HIV envelope glycoprotein deflect the immune response. Previous studies indicated that CD4+ T-cell epitope dominance is controlled by antigen three-dimensional structure through its influence on antigen processing and presentation. In this work, three disulfide bonds in the outer domain of gp120 were individually deleted in order to destabilize the local three-dimensional structure and enhance the presentation of nearby weakly immunogenic epitopes. However, upon immunization of groups of BALB/c mice, the CD4+ T-cell response was broadly reduced for all three variants, and distinct epitope profiles emerged. For one variant, antibody titers were sharply increased, and the antibody exhibited significant CD4-blocking activity. PMID:20089653

  9. Yellow fever virus envelope protein expressed in insect cells is capable of syncytium formation in lepidopteran cells and could be used for immunodetection of YFV in human sera

    OpenAIRE

    Nagata Tatsuya; Degallier Nicolas; Chaib Antônio JM; Galasso Tatiane GCM; Barros Maria CES; Ribeiro Bergmann M

    2011-01-01

    Abstract Background Yellow fever is an haemorrhagic disease caused by a virus that belongs to the genus Flavivirus (Flaviviridae family) and is transmitted by mosquitoes. Among the viral proteins, the envelope protein (E) is the most studied one, due to its high antigenic potencial. Baculovirus are one of the most popular and efficient eukaryotic expression system. In this study a recombinant baculovirus (vSynYFE) containing the envelope gene (env) of the 17D vaccine strain of yellow fever vi...

  10. Solving the mysteries of the bacterial cell – application of novel techniques in fluorescence microscopy

    Directory of Open Access Journals (Sweden)

    Magdalena Donczew

    2011-01-01

    Full Text Available We have reviewed how the development of fluorescent markers, triggered by the discovery of green fluorescence protein and its other color variants leading to the establishment of methods for studies of protein interactions with application of fluorescent proteins, affected the view of bacterial cell organization. Application of the new microscopic methods allowed localization of proteins and chromosomal regions, and observation of their migration in real time. These studies revealed the spatial organization of bacterial cells which includes specific subcellular localization of proteins, the presence of dynamic cytoskeletal structures, orchestrated and active segregation of chromosomes, and spatiotemporal gene regulation.

  11. Sulphation of N-linked oligosaccharides of vesicular stomatitis and influenza virus envelope glycoproteins: host cell specificity, subcellular localization and identification of substituted saccharides.

    Science.gov (United States)

    Karaivanova, V K; Spiro, R G

    1998-02-01

    The presence of sulphate groups on various saccharide residues of N-linked carbohydrate units has now been observed in a number of glycoproteins. To explore the cell specificity of this post-translational modification, we evaluated sulphate incorporation into virus envelope glycoproteins by a variety of cells, since it is believed that assembly of their N-linked oligosaccharides is to a large extent dependent on the enzymic machinery of the host. Employing the vesicular stomatitis virus (VSV) envelope glycoprotein (G protein) as a model, we noted that the addition of [35S]sulphate substituents into its complex carbohydrate units occurred in Madin-Darby canine kidney (MDCK), Madin-Darby bovine kidney, LLC-PK1 and BHK-21 cell lines but was not detectable in BRL 3A, BW5147.3, Chinese hamster ovary, HepG2, NRK-49F, IEC-18, PtK1 or 3T3 cells. The sulphate groups were exclusively located on C-3 of galactose [Gal(3-SO4)] and/or C-6 of N-acetylglucosamine [GlcNAc(6-SO4)] residues in the N-acetyllactosamine sequence of the branch chains. Moreover, we observed that the pronounced host-cell-dependence of the terminal galactose sulphation was reflected by the 3'-phosphoadenosine 5'-phosphosulphate:Gal-3-O-sulphotransferase activity assayed in vitro. Comparative studies carried out on the haemagglutinin of the influenza virus envelope formed by MDCK and LLC-PK1 cells indicated that sulphate in this glycoprotein was confined to its complex N-linked oligosaccharides where it occurred as Gal(3-SO4) and GlcNAc(6-SO4) on peripheral chains as well as on the mannose-substituted N-acetylglucosamine of the core. Since sulphation in both internal and peripheral locations of the virus glycoproteins was found to be arrested by the alpha1-->2 mannosidase inhibitor, kifunensine, as well as by the intracellular migration block imposed by brefeldin A, it was concluded that this modification is a late biosynthetic event which most likely takes place in the trans-Golgi network. PMID:9445377

  12. Bacterial cell-cell communication in the host via RRNPP peptide-binding regulators

    Directory of Open Access Journals (Sweden)

    David ePerez-Pascual

    2016-05-01

    Full Text Available Human microbiomes are composed of complex and dense bacterial consortia. In these environments, bacteria are able to react quickly to change by coordinating their gene expression at the population level via small signaling molecules. In Gram-positive bacteria, cell-cell communication is mostly mediated by peptides that are released into the extracellular environment. Cell-cell communication based on these peptides is especially widespread in the group Firmicutes, in which they regulate a wide array of biological processes, including functions related to host-microbe interactions. Among the different agents of communication, the RRNPP family of cytoplasmic transcriptional regulators, together with their cognate re-internalized signaling peptides, represents a group of emerging importance. RRNPP members that have been studied so far are found mainly in species of bacilli, streptococci, and enterococci. These bacteria are characterized as both human commensal and pathogenic, and share different niches in the human body with other microorganisms. The goal of this mini-review is to present the current state of research on the biological relevance of RRNPP mechanisms in the context of the host, highlighting their specific roles in commensalism or virulence.

  13. Effect of cell physicochemical characteristics and motility on bacterial transport in groundwater

    Science.gov (United States)

    Becker, M.W.; Collins, S.A.; Metge, D.W.; Harvey, R.W.; Shapiro, A.M.

    2004-01-01

    The influence of physicochemical characteristics and motility on bacterial transport in groundwater were examined in flow-through columns. Four strains of bacteria isolated from a crystalline rock groundwater system were investigated, with carboxylate-modified and amidine-modified latex microspheres and bromide as reference tracers. The bacterial isolates included a gram-positive rod (ML1), a gram-negative motile rod (ML2), a nonmotile mutant of ML2 (ML2m), and a gram-positive coccoid (ML3). Experiments were repeated at two flow velocities, in a glass column packed with glass beads, and in another packed with iron-oxyhydroxide coated glass beads. Bacteria breakthrough curves were interpreted using a transport equation that incorporates a sorption model from microscopic observation of bacterial deposition in flow-cell experiments. The model predicts that bacterial desorption rate will decrease exponentially with the amount of time the cell is attached to the solid surface. Desorption kinetics appeared to influence transport at the lower flow rate, but were not discernable at the higher flow rate. Iron-oxyhydroxide coatings had a lower-than-expected effect on bacterial breakthrough and no effect on the microsphere recovery in the column experiments. Cell wall type and shape also had minor effects on breakthrough. Motility tended to increase the adsorption rate, and decrease the desorption rate. The transport model predicts that at field scale, desorption rate kinetics may be important to the prediction of bacteria transport rates. ?? 2003 Elsevier B.V. All rights reserved.

  14. Relationship between Milk Microbiota, Bacterial Load, Macronutrients, and Human Cells during Lactation.

    Science.gov (United States)

    Boix-Amorós, Alba; Collado, Maria C; Mira, Alex

    2016-01-01

    Human breast milk is considered the optimal nutrition for infants, providing essential nutrients and a broad range of bioactive compounds, as well as its own microbiota. However, the interaction among those components and the biological role of milk microorganisms is still uncovered. Thus, our aim was to identify the relationships between milk microbiota composition, bacterial load, macronutrients, and human cells during lactation. Bacterial load was estimated in milk samples from a total of 21 healthy mothers through lactation time by bacteria-specific qPCR targeted to the single-copy gene fusA. Milk microbiome composition and diversity was estimated by 16S-pyrosequencing and the structure of these bacteria in the fluid was studied by flow cytometry, qPCR, and microscopy. Fat, protein, lactose, and dry extract of milk as well as the number of somatic cells were also analyzed. We observed that milk bacterial communities were generally complex, and showed individual-specific profiles. Milk microbiota was dominated by Staphylococcus, Pseudomonas, Streptococcus, and Acinetobacter. Staphylococcus aureus was not detected in any of these samples from healthy mothers. There was high variability in composition and number of bacteria per milliliter among mothers and in some cases even within mothers at different time points. The median bacterial load was 10(6) bacterial cells/ml through time, higher than those numbers reported by 16S gene PCR and culture methods. Furthermore, milk bacteria were present in a free-living, "planktonic" state, but also in equal proportion associated to human immune cells. There was no correlation between bacterial load and the amount of immune cells in milk, strengthening the idea that milk bacteria are not sensed as an infection by the immune system. PMID:27148183

  15. Relationship between Milk Microbiota, Bacterial Load, Macronutrients, and Human Cells during Lactation

    Science.gov (United States)

    Boix-Amorós, Alba; Collado, Maria C.; Mira, Alex

    2016-01-01

    Human breast milk is considered the optimal nutrition for infants, providing essential nutrients and a broad range of bioactive compounds, as well as its own microbiota. However, the interaction among those components and the biological role of milk microorganisms is still uncovered. Thus, our aim was to identify the relationships between milk microbiota composition, bacterial load, macronutrients, and human cells during lactation. Bacterial load was estimated in milk samples from a total of 21 healthy mothers through lactation time by bacteria-specific qPCR targeted to the single-copy gene fusA. Milk microbiome composition and diversity was estimated by 16S-pyrosequencing and the structure of these bacteria in the fluid was studied by flow cytometry, qPCR, and microscopy. Fat, protein, lactose, and dry extract of milk as well as the number of somatic cells were also analyzed. We observed that milk bacterial communities were generally complex, and showed individual-specific profiles. Milk microbiota was dominated by Staphylococcus, Pseudomonas, Streptococcus, and Acinetobacter. Staphylococcus aureus was not detected in any of these samples from healthy mothers. There was high variability in composition and number of bacteria per milliliter among mothers and in some cases even within mothers at different time points. The median bacterial load was 106 bacterial cells/ml through time, higher than those numbers reported by 16S gene PCR and culture methods. Furthermore, milk bacteria were present in a free-living, “planktonic” state, but also in equal proportion associated to human immune cells. There was no correlation between bacterial load and the amount of immune cells in milk, strengthening the idea that milk bacteria are not sensed as an infection by the immune system.

  16. Cooperation between Monocyte-Derived Cells and Lymphoid Cells in the Acute Response to a Bacterial Lung Pathogen.

    Directory of Open Access Journals (Sweden)

    Andrew S Brown

    2016-06-01

    Full Text Available Legionella pneumophila is the causative agent of Legionnaires' disease, a potentially fatal lung infection. Alveolar macrophages support intracellular replication of L. pneumophila, however the contributions of other immune cell types to bacterial killing during infection are unclear. Here, we used recently described methods to characterise the major inflammatory cells in lung after acute respiratory infection of mice with L. pneumophila. We observed that the numbers of alveolar macrophages rapidly decreased after infection coincident with a rapid infiltration of the lung by monocyte-derived cells (MC, which, together with neutrophils, became the dominant inflammatory cells associated with the bacteria. Using mice in which the ability of MC to infiltrate tissues is impaired it was found that MC were required for bacterial clearance and were the major source of IL12. IL12 was needed to induce IFNγ production by lymphoid cells including NK cells, memory T cells, NKT cells and γδ T cells. Memory T cells that produced IFNγ appeared to be circulating effector/memory T cells that infiltrated the lung after infection. IFNγ production by memory T cells was stimulated in an antigen-independent fashion and could effectively clear bacteria from the lung indicating that memory T cells are an important contributor to innate bacterial defence. We also determined that a major function of IFNγ was to stimulate bactericidal activity of MC. On the other hand, neutrophils did not require IFNγ to kill bacteria and alveolar macrophages remained poorly bactericidal even in the presence of IFNγ. This work has revealed a cooperative innate immune circuit between lymphoid cells and MC that combats acute L. pneumophila infection and defines a specific role for IFNγ in anti-bacterial immunity.

  17. Cell motility and antibiotic tolerance of bacterial swarms

    Science.gov (United States)

    Zuo, Wenlong

    Many bacteria species can move across moist surfaces in a coordinated manner known as swarming. It is reported that swarm cells show higher tolerance to a wide variety of antibiotics than planktonic cells. We used the model bacterium E. coli to study how motility affects the antibiotic tolerance of swarm cells. Our results provide new insights for the control of pathogenic invasion via regulating cell motility. Mailing address: Room 306 Science Centre North Block, The Chinese University of Hong Kong, Shatin, N.T. Hong Kong SAR. Phone: +852-3943-6354. Fax: +852-2603-5204. E-mail: zwlong@live.com.

  18. Fluorescence imaging for bacterial cell biology: from localization to dynamics, from ensembles to single molecules.

    Science.gov (United States)

    Yao, Zhizhong; Carballido-López, Rut

    2014-01-01

    Fluorescent proteins and developments in superresolution (nanoscopy) and single-molecule techniques bring high sensitivity, speed, and one order of magnitude gain in spatial resolution to live-cell imaging. These technologies have only recently been applied to prokaryotic cell biology, revealing the exquisite subcellular organization of bacterial cells. Here, we review the parallel evolution of fluorescence microscopy methods and their application to bacteria, mainly drawing examples from visualizing actin-like MreB proteins in the model bacterium Bacillus subtilis. We describe the basic principles of nanoscopy and conventional techniques and their advantages and limitations to help microbiologists choose the most suitable technique for their biological question. Looking ahead, multidimensional live-cell nanoscopy combined with computational image analysis tools, systems biology approaches, and mathematical modeling will provide movie-like, mechanistic, and quantitative description of molecular events in bacterial cells. PMID:25002084

  19. Heterotrophic free-living and particle-bound bacterial cell size in the river Cauvery and its downstream tributaries

    Indian Academy of Sciences (India)

    T S Harsha; Sadanand M Yamakanamardi; M Mahadevaswamy

    2007-03-01

    This is the first comprehensive study on planktonic heterotrophic bacterial cell size in the river Cauvery and its important tributaries in Karnataka State, India. The initial hypothesis that the mean cell size of planktonic heterotrophic bacteria in the four tributaries are markedly different from each other and also from that in the main river Cauvery was rejected, because all five watercourses showed similar planktonic heterotrophic bacterial cell size. Examination of the correlation between mean heterotrophic bacterial cell size and environmental variables showed four correlations in the river Arkavathy and two in the river Shimsha. Regression analysis revealed that 18% of the variation in mean heterotrophic free-living bacterial cell size was due to biological oxygen demand (BOD) in the river Arkavathy, 11% due to surface water velocity (SWV) in the river Cauvery and 11% due to temperature in the river Kapila. Heterotrophic particle-bound bacterial cell size variation was 28% due to chloride and BOD in the river Arkavathy, 11% due to conductivity in the river Kapila and 8% due to calcium in the river Cauvery. This type of relationship between heterotrophic bacterial cell size and environmental variables suggests that, though the mean heterotrophic bacterial cell size was similar in all the five water courses, different sets of environmental variables apparently control the heterotrophic bacterial cell size in the various water bodies studied in this investigation. The possible cause for this environmental (bottom–up) control is discussed.

  20. Contribution of bacterial cell nitrogen to soil humic fractions

    International Nuclear Information System (INIS)

    Living cells of Serratia marcescens, uniformly labelled with 15N, were added to samples of maple (Acer saccharum) and black spruce (Picea mariana) forest soils. After different periods of incubation from zero time to 100 days, the soils were subjected to alkali-acid and phenol extraction to provide humic acid, fulvic acid, humin and 'humoprotein' fractions. Significant amounts of the cell nitrogen were recovered in the humic and fulvic acids immediately after addition. After incubation, less cell nitrogen appeared in the humic acid and more in the fulvic acid. The amount of cell nitrogen recovered in the humin fraction increased with incubation. Roughly 5 to 10 per cent of the added cell nitrogen was found as amino acid nitrogen from humoprotein in a phenol extract of the humic acid. The data are consistent with the occurrence of co-precipitation of biologically labile biomass nitrogen compounds with humic polymers during the alkaline extraction procedure involved in the humic-fulvic fractionation. (orig.)

  1. Enhanced metalloadsorption of bacterial cells displaying poly-His peptides

    Energy Technology Data Exchange (ETDEWEB)

    Sousa, C.; Cebolla, A.; Lorenzo, V. de [CSIC, Madrid (Spain)

    1996-08-01

    The properties of Escherichia coli cells, acquired by cell surface presentation of one or two hexahistidine (His) clusters carried by the outer membrane LamB protein, have been examined. Strains producing LamB hybrids with the His chains accumulated greater than 11-fold more Cd{sup 2} than E. coli cells expressing the protein without the His insert. Furthermore, the hexa-His chains on the cell surface caused cells to adhere reversibly to a Ni{sup 2+}-containing solid matrix in a metal-dependent fashion. Thus, expression of poly-His peptides enables bacteria to act as a metalloaffinity adsorbent. These results open up the possibility for biosorption of heavy ions using engineered microorganisms. 32 refs., 3 figs.

  2. Label-free isolation and deposition of single bacterial cells from heterogeneous samples for clonal culturing.

    Science.gov (United States)

    Riba, J; Gleichmann, T; Zimmermann, S; Zengerle, R; Koltay, P

    2016-01-01

    The isolation and analysis of single prokaryotic cells down to 1 μm and less in size poses a special challenge and requires micro-engineered devices to handle volumes in the picoliter to nanoliter range. Here, an advanced Single-Cell Printer (SCP) was applied for automated and label-free isolation and deposition of bacterial cells encapsulated in 35 pl droplets by inkjet-like printing. To achieve this, dispenser chips to generate micro droplets have been fabricated with nozzles 20 μm in size. Further, the magnification of the optical system used for cell detection was increased. Redesign of the optical path allows for collision-free addressing of any flat substrate since no compartment protrudes below the nozzle of the dispenser chip anymore. The improved system allows for deterministic isolation of individual bacterial cells. A single-cell printing efficiency of 93% was obtained as shown by printing fluorescent labeled E. coli. A 96-well plate filled with growth medium is inoculated with single bacteria cells on average within about 8 min. Finally, individual bacterial cells from a heterogeneous sample of E. coli and E. faecalis were isolated for clonal culturing directly on agar plates in user-defined array geometry. PMID:27596612

  3. Isolation of corneocyte envelopes from porcine epidermis.

    Science.gov (United States)

    Swartzendruber, D C; Kitko, D J; Wertz, P W; Madison, K C; Downing, D T

    1988-01-01

    Sheets of porcine stratum corneum were dispersed into individual corneocytes after 4 h in a solution consisting of 8 mM N,N-dimethyldodecylamine oxide and 2 mM sodium dodecylsulfate in phosphate-buffered isotonic saline, at 45 degrees C. With continued detergent treatment and moderate sonication, most of the cells lost their keratin contents and were then separated from the remaining intact cells by centrifugation in cesium chloride solution of density 1.280. Electron microscopy showed that the cell envelopes retained both the crosslinked protein envelope and its attached lipid envelope. The dry weight of envelopes was approximately 7% of the estimated dry weight of the original stratum corneum, while the corneocytes surviving intact also amounted to 7% of the starting weight. Mild alkaline hydrolysis of the corneocyte envelopes allowed the extraction of hydroxyceramides amounting to 10% of the dry weight of the envelopes. The procedure therefore provides isolated corneocyte envelopes suitable for studying both the protein and lipid components of this compound sheath. PMID:3207369

  4. Do bacterial cell numbers follow a theoretical Poisson distribution? Comparison of experimentally obtained numbers of single cells with random number generation via computer simulation.

    Science.gov (United States)

    Koyama, Kento; Hokunan, Hidekazu; Hasegawa, Mayumi; Kawamura, Shuso; Koseki, Shigenobu

    2016-12-01

    We investigated a bacterial sample preparation procedure for single-cell studies. In the present study, we examined whether single bacterial cells obtained via 10-fold dilution followed a theoretical Poisson distribution. Four serotypes of Salmonella enterica, three serotypes of enterohaemorrhagic Escherichia coli and one serotype of Listeria monocytogenes were used as sample bacteria. An inoculum of each serotype was prepared via a 10-fold dilution series to obtain bacterial cell counts with mean values of one or two. To determine whether the experimentally obtained bacterial cell counts follow a theoretical Poisson distribution, a likelihood ratio test between the experimentally obtained cell counts and Poisson distribution which parameter estimated by maximum likelihood estimation (MLE) was conducted. The bacterial cell counts of each serotype sufficiently followed a Poisson distribution. Furthermore, to examine the validity of the parameters of Poisson distribution from experimentally obtained bacterial cell counts, we compared these with the parameters of a Poisson distribution that were estimated using random number generation via computer simulation. The Poisson distribution parameters experimentally obtained from bacterial cell counts were within the range of the parameters estimated using a computer simulation. These results demonstrate that the bacterial cell counts of each serotype obtained via 10-fold dilution followed a Poisson distribution. The fact that the frequency of bacterial cell counts follows a Poisson distribution at low number would be applied to some single-cell studies with a few bacterial cells. In particular, the procedure presented in this study enables us to develop an inactivation model at the single-cell level that can estimate the variability of survival bacterial numbers during the bacterial death process. PMID:27554145

  5. Classification of Lactococcus lactis cell envelope proteinase based on gene sequencing, peptides formed after hydrolysis of milk, and computer modeling

    DEFF Research Database (Denmark)

    Børsting, Mette Winther; Qvist, K.B.; Brockmann, E.; Vindeløv, J.; Pedersen, T.L.; Vogensen, Finn Kvist; Ardö, Ylva Margareta

    2015-01-01

    Lactococcus lactis strains depend on a proteolytic system for growth in milk to release essential AA from casein. The cleavage specificities of the cell envelope proteinase (CEP) can vary between strains and environments and whether the enzyme is released or bound to the cell wall. Thirty-eight Lc....... lactis strains were grouped according to their CEP AA sequences and according to identified peptides after hydrolysis of milk. Finally, AA positions in the substrate binding region were suggested by the use of a new CEP template based on Streptococcus C5a CEP. Aligning the CEP AA sequences of 38 strains...... of Lc. lactis showed that 21 strains, which were previously classified as group d, could be subdivided into 3 groups. Independently, similar subgroupings were found based on comparison of the Lc. lactis CEP AA sequences and based on normalized quantity of identified peptides released from αS1-casein...

  6. Streptomyces: A Screening Tool for Bacterial Cell Division Inhibitors

    Science.gov (United States)

    Jani, Charul; Tocheva, Elitza I.; McAuley, Scott; Craney, Arryn; Jensen, Grant J.; Nodwell, Justin

    2016-01-01

    Cell division is essential for spore formation but not for viability in the filamentous streptomycetes bacteria. Failure to complete cell division instead blocks spore formation, a phenotype that can be visualized by the absence of gray (in Streptomyces coelicolor) and green (in Streptomyces venezuelae) spore-associated pigmentation. Despite the lack of essentiality, the streptomycetes divisome is similar to that of other prokaryotes. Therefore, the chemical inhibitors of sporulation in model streptomycetes may interfere with the cell division in rod-shaped bacteria as well. To test this, we investigated 196 compounds that inhibit sporulation in S. coelicolor. We show that 19 of these compounds cause filamentous growth in Bacillus subtilis, consistent with impaired cell division. One of the compounds is a DNA-damaging agent and inhibits cell division by activating the SOS response. The remaining 18 act independently of known stress responses and may therefore act on the divisome or on divisome positioning and stability. Three of the compounds (Fil-1, Fil-2, and Fil-3) confer distinct cell division defects on B. subtilis. They also block B. subtilis sporulation, which is mechanistically unrelated to the sporulation pathway of streptomycetes but is also dependent on the divisome. We discuss ways in which these differing phenotypes can be used in screens for cell division inhibitors. PMID:25256667

  7. The HIV-1 envelope protein gp120 is captured and displayed for B cell recognition by SIGN-R1(+) lymph node macrophages.

    Science.gov (United States)

    Park, Chung; Arthos, James; Cicala, Claudia; Kehrl, John H

    2015-01-01

    The HIV-1 envelope protein gp120 is both the target of neutralizing antibodies and a major focus of vaccine efforts; however how it is delivered to B cells to elicit an antibody response is unknown. Here, we show that following local gp120 injection lymph node (LN) SIGN-R1(+) sinus macrophages located in interfollicular pockets and underlying SIGN-R1(+) macrophages form a cellular network that rapidly captures gp120 from the afferent lymph. In contrast, two other antigens, phycoerythrin and hen egg lysozyme, were not captured by these cells. Intravital imaging of mouse LNs revealed persistent, but transient interactions between gp120 bearing interfollicular network cells and both trafficking and LN follicle resident gp120 specific B cells. The gp120 specific, but not the control B cells repetitively extracted gp120 from the network cells. Our findings reveal a specialized LN antigen delivery system poised to deliver gp120 and likely other pathogen derived glycoproteins to B cells. PMID:26258881

  8. Effects of bacterial cells and two types of extracellular polymers on bioclogging of sand columns

    Science.gov (United States)

    Xia, Lu; Zheng, Xilai; Shao, Haibing; Xin, Jia; Sun, Zhaoyue; Wang, Leyun

    2016-04-01

    Microbially induced reductions in the saturated hydraulic conductivity, Ks, of natural porous media, conventionally called bioclogging, occurs frequently in natural and engineered subsurface systems. Bioclogging can affect artificial groundwater recharge, in situ bioremediation of contaminated aquifers, or permeable reactive barriers. In this study, we designed a series of percolation experiments to simulate the growth and metabolism of bacteria in sand columns. The experimental results showed that the bacterial cell amount gradually increased to a maximum of 8.91 log10 CFU/g sand at 144 h during the bioclogging process, followed by a decrease to 7.89 log10 CFU/g sand until 336 h. The same variation pattern was found for the concentration of tightly bound extracellular polymeric substances (TB-EPS), which had a peak value of 220.76 μg/g sand at 144 h. In the same experiments, the concentration of loosely bound extracellular polymeric substances (LB-EPS) increased sharply from 54.45 to 575.57 μg/g sand in 192 h, followed by a slight decline to 505.04 μg/g sand. The increase of the bacterial cell amount along with the other two concentrations could reduce the Ks of porous media, but their relative contributions varied to a large degree during different percolation stages. At the beginning of the tests (e.g., 48 h before), bacterial cells were likely responsible for the Ks reduction of porous media because no increase was found for the other two concentrations. With the accumulation of cells and EPS production from 48 to 144 h, both were important for the reduction of Ks. However, in the late period of percolation tests from 144 to 192 h, LB-EPS was probably responsible for the further reduction of Ks, as the bacterial cell amount and TB-EPS concentration decreased. Quantitative contributions of bacterial cell amount and the two types of extracellular polymers to Ks reductions were also evaluated.

  9. Homeostatic interplay between bacterial cell-cell signaling and iron in virulence.

    Directory of Open Access Journals (Sweden)

    Ronen Hazan

    2010-03-01

    Full Text Available Pathogenic bacteria use interconnected multi-layered regulatory networks, such as quorum sensing (QS networks to sense and respond to environmental cues and external and internal bacterial cell signals, and thereby adapt to and exploit target hosts. Despite the many advances that have been made in understanding QS regulation, little is known regarding how these inputs are integrated and processed in the context of multi-layered QS regulatory networks. Here we report the examination of the Pseudomonas aeruginosa QS 4-hydroxy-2-alkylquinolines (HAQs MvfR regulatory network and determination of its interaction with the QS acyl-homoserine-lactone (AHL RhlR network. The aim of this work was to elucidate paradigmatically the complex relationships between multi-layered regulatory QS circuitries, their signaling molecules, and the environmental cues to which they respond. Our findings revealed positive and negative homeostatic regulatory loops that fine-tune the MvfR regulon via a multi-layered dependent homeostatic regulation of the cell-cell signaling molecules PQS and HHQ, and interplay between these molecules and iron. We discovered that the MvfR regulon component PqsE is a key mediator in orchestrating this homeostatic regulation, and in establishing a connection to the QS rhlR system in cooperation with RhlR. Our results show that P. aeruginosa modulates the intensity of its virulence response, at least in part, through this multi-layered interplay. Our findings underscore the importance of the homeostatic interplay that balances competition within and between QS systems via cell-cell signaling molecules and environmental cues in the control of virulence gene expression. Elucidation of the fine-tuning of this complex relationship offers novel insights into the regulation of these systems and may inform strategies designed to limit infections caused by P. aeruginosa and related human pathogens.

  10. Biosynthesis of a Fully Functional Cyclotide inside Living Bacterial Cells

    Energy Technology Data Exchange (ETDEWEB)

    Camarero, J A; Kimura, R H; Woo, Y; Cantor, J; Shekhtman, A

    2007-04-05

    The cyclotide MCoTI-II is a powerful trypsin inhibitor recently isolated from the seeds of Momordica cochinchinensis, a plant member of cucurbitaceae family. We report for the first time the in vivo biosynthesis of natively-folded MCoTI-II inside live E. coli cells. Our biomimetic approach involves the intracellular backbone cyclization of a linear cyclotide-intein fusion precursor mediated by a modified protein splicing domain. The cyclized peptide then spontaneously folds into its native conformation. The use of genetically engineered E. coli cells containing mutations in the glutathione and thioredoxin reductase genes considerably improves the production of folded MCoTI-II in vivo. Biochemical and structural characterization of the recombinant MCoTI-II confirmed its identity. Biosynthetic access to correctly-folded cyclotides allows the possibility of generating cell-based combinatorial libraries that can be screened inside living cells for their ability to modulate or inhibit cellular processes.

  11. Bacterial toxins fuel disease progression in cutaneous T-cell lymphoma

    DEFF Research Database (Denmark)

    Willerslev-Olsen, Andreas; Krejsgaard, Thorbjørn; Lindahl, Lise M;

    2013-01-01

    In patients with cutaneous T-cell lymphoma (CTCL) bacterial infections constitute a major clinical problem caused by compromised skin barrier and a progressive immunodeficiency. Indeed, the majority of patients with advanced disease die from infections with bacteria, e.g., Staphylococcus aureus...

  12. Soluble triggering receptor expressed on myeloid cells 1: a biomarker for bacterial meningitis

    NARCIS (Netherlands)

    R.M. Determann; M. Weisfelt; J. de Gans; A. van der Ende; M.J. Schultz; D. van de Beek

    2006-01-01

    Objective: To evaluate whether soluble triggering receptor expressed on myeloid cells 1 (sTREM-1) in CSF can serve as a biomarker for the presence of bacterial meningitis and outcome in patients with this disease. Design: Retrospective study of diagnostic accuracy. Setting and patients: CSF was coll

  13. Increased electrical output when a bacterial ABTS oxidizer is used in a microbial fuel cell

    Science.gov (United States)

    Microbial fuel cells (MFCs) are a technology that provides electrical energy from the microbial oxidation of organic compounds. Most MFCs use oxygen as the oxidant in the cathode chamber. The present study examined the formation in culture of an unidentified bacterial oxidant and investigated the ...

  14. Specific labeling of peptidoglycan precursors as a tool for bacterial cell wall studies

    NARCIS (Netherlands)

    van Dam, V.; Olrichs, N.K.; Breukink, E.J.

    2009-01-01

    Wall chart: The predominant component of the bacterial cell wall, peptidoglycan, consists of long alternating stretches of aminosugar subunits interlinked in a large three-dimensional network and is formed from precursors through several cytosolic and membrane-bound steps. The high tolerance of the

  15. The Role of Lipid Domains in Bacterial Cell Processes

    OpenAIRE

    Katarína Muchová; Imrich Barák

    2013-01-01

    Membranes are vital structures for cellular life forms. As thin, hydrophobic films, they provide a physical barrier separating the aqueous cytoplasm from the outside world or from the interiors of other cellular compartments. They maintain a selective permeability for the import and export of water-soluble compounds, enabling the living cell to maintain a stable chemical environment for biological processes. Cell membranes are primarily composed of two crucial substances, lipids and proteins....

  16. Spring constants and adhesive properties of native bacterial biofilm cells measured by atomic force microscopy.

    Science.gov (United States)

    Volle, C B; Ferguson, M A; Aidala, K E; Spain, E M; Núñez, M E

    2008-11-15

    Bacterial biofilms were imaged by atomic force microscopy (AFM), and their elasticity and adhesion to the AFM tip were determined from a series of tip extension and retraction cycles. Though the five bacterial strains studied included both Gram-negative and -positive bacteria and both environmental and laboratory strains, all formed simple biofilms on glass surfaces. Cellular spring constants, determined from the extension portion of the force cycle, varied between 0.16+/-0.01 and 0.41+/-0.01 N/m, where larger spring constants were measured for Gram-positive cells than for Gram-negative cells. The nonlinear regime in the extension curve depended upon the biomolecules on the cell surface: the extension curves for the smooth Gram-negative bacterial strains with the longest lipopolysaccharides on their surface had a larger nonlinear region than the rough bacterial strain with shorter lipopolysaccharides on the surface. Adhesive forces between the retracting silicon nitride tip and the cells varied between cell types in terms of the force components, the distance components, and the number of adhesion events. The Gram-negative cells' adhesion to the tip showed the longest distance components, sometimes more than 1 microm, whereas the shortest distance adhesion events were measured between the two Gram-positive cell types and the tip. Fixation of free-swimming planktonic cells by NHS and EDC perturbed both the elasticity and the adhesive properties of the cells. Here we consider the biochemical meaning of the measured physical properties of simple biofilms and implications to the colonization of surfaces in the first stages of biofilm formation. PMID:18815013

  17. In situ probing the interior of single bacterial cells at nanometer scale

    International Nuclear Information System (INIS)

    We report a novel approach to probe the interior of single bacterial cells at nanometre resolution by combining focused ion beam (FIB) and atomic force microscopy (AFM). After removing layers of pre-defined thickness in the order of 100 nm on the target bacterial cells with FIB milling, AFM of different modes can be employed to probe the cellular interior under both ambient and aqueous environments. Our initial investigations focused on the surface topology induced by FIB milling and the hydration effects on AFM measurements, followed by assessment of the sample protocols. With fine-tuning of the process parameters, in situ AFM probing beneath the bacterial cell wall was achieved for the first time. We further demonstrate the proposed method by performing a spatial mapping of intracellular elasticity and chemistry of the multi-drug resistant strain Klebsiella pneumoniae cells prior to and after it was exposed to the ‘last-line’ antibiotic polymyxin B. Our results revealed increased stiffness occurring in both surface and interior regions of the treated cells, suggesting loss of integrity of the outer membrane from polymyxin treatments. In addition, the hydrophobicity measurement using a functionalized AFM tip was able to highlight the evident hydrophobic portion of the cell such as the regions containing cell membrane. We expect that the proposed FIB–AFM platform will help in gaining deeper insights of bacteria–drug interactions to develop potential strategies for combating multi-drug resistance. (paper)

  18. Bacterial neuraminidase increases IL-8 production in lung epithelial cells via NF-κB-dependent pathway

    International Nuclear Information System (INIS)

    Bacterial neuraminidase, a sialic acid-degrading enzyme, is one of the virulent factors produced in pathogenic bacteria like as other bacterial components. However little is known about whether bacterial neuraminidase can initiate or modify a cellular response, such as cytokine production, in epithelial cells at infection and inflammation. We demonstrate here that bacterial neuraminidase, but not heat-inactivated neuraminidase, up-regulates expression of interleukin-8 (IL-8) mRNA and protein in lung epithelial A549 and NCI-H292 cells. We also show that bacterial neuraminidase significantly up-regulates IL-8 promoter activity as well as nuclear factor-kappaB (NF-κB) reporter activity. Moreover, inhibition of NF-κB signaling suppressed IL-8 mRNA expression induced by bacterial neuraminidase. Taken together, desialylation-induced IL-8 production in lung epithelial cells may play an important role in infection-associated inflammatory events.

  19. Note: An automated image analysis method for high-throughput classification of surface-bound bacterial cell motions.

    Science.gov (United States)

    Shen, Simon; Syal, Karan; Tao, Nongjian; Wang, Shaopeng

    2015-12-01

    We present a Single-Cell Motion Characterization System (SiCMoCS) to automatically extract bacterial cell morphological features from microscope images and use those features to automatically classify cell motion for rod shaped motile bacterial cells. In some imaging based studies, bacteria cells need to be attached to the surface for time-lapse observation of cellular processes such as cell membrane-protein interactions and membrane elasticity. These studies often generate large volumes of images. Extracting accurate bacterial cell morphology features from these images is critical for quantitative assessment. Using SiCMoCS, we demonstrated simultaneous and automated motion tracking and classification of hundreds of individual cells in an image sequence of several hundred frames. This is a significant improvement from traditional manual and semi-automated approaches to segmenting bacterial cells based on empirical thresholds, and a first attempt to automatically classify bacterial motion types for motile rod shaped bacterial cells, which enables rapid and quantitative analysis of various types of bacterial motion. PMID:26724085

  20. Note: An automated image analysis method for high-throughput classification of surface-bound bacterial cell motions

    Science.gov (United States)

    Shen, Simon; Syal, Karan; Tao, Nongjian; Wang, Shaopeng

    2015-12-01

    We present a Single-Cell Motion Characterization System (SiCMoCS) to automatically extract bacterial cell morphological features from microscope images and use those features to automatically classify cell motion for rod shaped motile bacterial cells. In some imaging based studies, bacteria cells need to be attached to the surface for time-lapse observation of cellular processes such as cell membrane-protein interactions and membrane elasticity. These studies often generate large volumes of images. Extracting accurate bacterial cell morphology features from these images is critical for quantitative assessment. Using SiCMoCS, we demonstrated simultaneous and automated motion tracking and classification of hundreds of individual cells in an image sequence of several hundred frames. This is a significant improvement from traditional manual and semi-automated approaches to segmenting bacterial cells based on empirical thresholds, and a first attempt to automatically classify bacterial motion types for motile rod shaped bacterial cells, which enables rapid and quantitative analysis of various types of bacterial motion.

  1. Storage envelopes or sleeves

    International Nuclear Information System (INIS)

    A storage envelope or sleeve particularly for processed X-ray films is described. It consists of front and back panels joined together at a hinge line and connected along the intermediate sides by connecting flaps. An inner pocket is formed from a third flap which is folded to lie against the inner face of the back panel. The panels may have additional score lines parallel to the closed sides of the envelope and the inner pocket so that the envelope and the inner pocket can accommodate bulky contents. The free edge of the pocket is inset from the open side of the envelope, and finger cut-outs may be provided to facilitate access to the contents of the envelope and the pocket. (author)

  2. Inclusion bodies, bacterial cells and compositions containing them and uses thereof

    OpenAIRE

    Veciana Miró, Jaume; Ratera Bastardas, Inmaculada; Díez Gil, César; Villaverde Corrales, Antonio Pedro; Vázquez Gómez, Esther; García Fruitós, Elena

    2008-01-01

    [EN] The present invention relates to an isolated inclusion body comprising a polypeptide, characterised in that such inclusion body is in particulate formo The present invention also refers to a bacterial cell comprising said inclusion body. The present invention additionally refers to a composition comprising said inclusion body and a eukaryotic cell. The present invention moreover refers to a composition comprising said inclusion body and animal or plant tissue. The present invent...

  3. Lipid-linked cell wall precursors regulate membrane association of bacterial actin MreB

    OpenAIRE

    Schirner, Kathrin; Eun, Ye-Jin; Dion, Mike; Luo, Yun; Helmann, John D.; Garner, Ethan C.; Walker, Suzanne

    2014-01-01

    Summary The bacterial actin homolog MreB, which is critical for rod shape determination, forms filaments that rotate around the cell width on the inner surface of the cytoplasmic membrane. What determines filament association with the membranes or with other cell wall elongation proteins is not known. Using specific chemical and genetic perturbations while following MreB filament motion, we find that MreB membrane association is an actively regulated process that depends on the presence of li...

  4. Spatial Patterning of Newly-Inserted Material during Bacterial Cell Growth

    Science.gov (United States)

    Ursell, Tristan

    2012-02-01

    In the life cycle of a bacterium, rudimentary microscopy demonstrates that cell growth and elongation are essential characteristics of cellular reproduction. The peptidoglycan cell wall is the main load-bearing structure that determines both cell shape and overall size. However, simple imaging of cellular growth gives no indication of the spatial patterning nor mechanism by which material is being incorporated into the pre-existing cell wall. We employ a combination of high-resolution pulse-chase fluorescence microscopy, 3D computational microscopy, and detailed mechanistic simulations to explore how spatial patterning results in uniform growth and maintenance of cell shape. We show that growth is happening in discrete bursts randomly distributed over the cell surface, with a well-defined mean size and average rate. We further use these techniques to explore the effects of division and cell wall disrupting antibiotics, like cephalexin and A22, respectively, on the patterning of cell wall growth in E. coli. Finally, we explore the spatial correlation between presence of the bacterial actin-like cytoskeletal protein, MreB, and local cell wall growth. Together these techniques form a powerful method for exploring the detailed dynamics and involvement of antibiotics and cell wall-associated proteins in bacterial cell growth.[4pt] In collaboration with Kerwyn Huang, Stanford University.

  5. A novel mechanism of bacterial toxin transfer within host blood cell-derived microvesicles.

    Directory of Open Access Journals (Sweden)

    Anne-lie Ståhl

    2015-02-01

    Full Text Available Shiga toxin (Stx is the main virulence factor of enterohemorrhagic Escherichia coli, which are non-invasive strains that can lead to hemolytic uremic syndrome (HUS, associated with renal failure and death. Although bacteremia does not occur, bacterial virulence factors gain access to the circulation and are thereafter presumed to cause target organ damage. Stx was previously shown to circulate bound to blood cells but the mechanism by which it would potentially transfer to target organ cells has not been elucidated. Here we show that blood cell-derived microvesicles, shed during HUS, contain Stx and are found within patient renal cortical cells. The finding was reproduced in mice infected with Stx-producing Escherichia coli exhibiting Stx-containing blood cell-derived microvesicles in the circulation that reached the kidney where they were transferred into glomerular and peritubular capillary endothelial cells and further through their basement membranes followed by podocytes and tubular epithelial cells, respectively. In vitro studies demonstrated that blood cell-derived microvesicles containing Stx undergo endocytosis in glomerular endothelial cells leading to cell death secondary to inhibited protein synthesis. This study demonstrates a novel virulence mechanism whereby bacterial toxin is transferred within host blood cell-derived microvesicles in which it may evade the host immune system.

  6. Metabolism of bacterial cells bound to solid carriers

    International Nuclear Information System (INIS)

    Escherichia coli 2260 cells bound to solid carriers (iodo-and bromoacetyl cellulose) were incubated in the Davis mineral medium and changes in respiratory activity, ATP level, 14C-valine and 14C-adenine incorporation and the number of bacteria able to multiply were investigated. As compared with suspended cells, no significant changes in the rate and capacity of the metabolic processes studied were recorded. Iodoacetyl cellulose had the smallest effect on the rates of nucleic acids and protein syntheses. (author). 3 figs., 2 tabs., 11 refs

  7. Cell wall mechanical properties as measured with bacterial thread made from Bacillus subtilis.

    OpenAIRE

    Mendelson, N H; Thwaites, J J

    1989-01-01

    Engineering approaches used in the study of textile fibers have been applied to the measurement of mechanical properties of bacterial cell walls by using the Bacillus subtilis bacterial thread system. Improved methods have been developed for the production of thread and for measuring its mechanical properties. The best specimens of thread produced from cultures of strain FJ7 grown in TB medium at 20 degrees C varied in diameter by a factor of 1.09 over a 30-mm thread length. The stress-strain...

  8. Heterologous Expression of Toxins from Bacterial Toxin-Antitoxin Systems in Eukaryotic Cells: Strategies and Applications.

    Science.gov (United States)

    Yeo, Chew Chieng; Abu Bakar, Fauziah; Chan, Wai Ting; Espinosa, Manuel; Harikrishna, Jennifer Ann

    2016-02-01

    Toxin-antitoxin (TA) systems are found in nearly all prokaryotic genomes and usually consist of a pair of co-transcribed genes, one of which encodes a stable toxin and the other, its cognate labile antitoxin. Certain environmental and physiological cues trigger the degradation of the antitoxin, causing activation of the toxin, leading either to the death or stasis of the host cell. TA systems have a variety of functions in the bacterial cell, including acting as mediators of programmed cell death, the induction of a dormant state known as persistence and the stable maintenance of plasmids and other mobile genetic elements. Some bacterial TA systems are functional when expressed in eukaryotic cells and this has led to several innovative applications, which are the subject of this review. Here, we look at how bacterial TA systems have been utilized for the genetic manipulation of yeasts and other eukaryotes, for the containment of genetically modified organisms, and for the engineering of high expression eukaryotic cell lines. We also examine how TA systems have been adopted as an important tool in developmental biology research for the ablation of specific cells and the potential for utility of TA systems in antiviral and anticancer gene therapies. PMID:26907343

  9. Heterologous Expression of Toxins from Bacterial Toxin-Antitoxin Systems in Eukaryotic Cells: Strategies and Applications

    Directory of Open Access Journals (Sweden)

    Chew Chieng Yeo

    2016-02-01

    Full Text Available Toxin-antitoxin (TA systems are found in nearly all prokaryotic genomes and usually consist of a pair of co-transcribed genes, one of which encodes a stable toxin and the other, its cognate labile antitoxin. Certain environmental and physiological cues trigger the degradation of the antitoxin, causing activation of the toxin, leading either to the death or stasis of the host cell. TA systems have a variety of functions in the bacterial cell, including acting as mediators of programmed cell death, the induction of a dormant state known as persistence and the stable maintenance of plasmids and other mobile genetic elements. Some bacterial TA systems are functional when expressed in eukaryotic cells and this has led to several innovative applications, which are the subject of this review. Here, we look at how bacterial TA systems have been utilized for the genetic manipulation of yeasts and other eukaryotes, for the containment of genetically modified organisms, and for the engineering of high expression eukaryotic cell lines. We also examine how TA systems have been adopted as an important tool in developmental biology research for the ablation of specific cells and the potential for utility of TA systems in antiviral and anticancer gene therapies.

  10. DNA-crosslinker cisplatin eradicates bacterial persister cells.

    Science.gov (United States)

    Chowdhury, Nityananda; Wood, Thammajun L; Martínez-Vázquez, Mariano; García-Contreras, Rodolfo; Wood, Thomas K

    2016-09-01

    For all bacteria, nearly every antimicrobial fails since a subpopulation of the bacteria enter a dormant state known as persistence, in which the antimicrobials are rendered ineffective due to the lack of metabolism. This tolerance to antibiotics makes microbial infections the leading cause of death worldwide and makes treating chronic infections, including those of wounds problematic. Here, we show that the FDA-approved anti-cancer drug cisplatin [cis-diamminodichloroplatinum(II)], which mainly forms intra-strand DNA crosslinks, eradicates Escherichia coli K-12 persister cells through a growth-independent mechanism. Additionally, cisplatin is more effective at killing Pseudomonas aeruginosa persister cells than mitomycin C, which forms inter-strand DNA crosslinks, and cisplatin eradicates the persister cells of several pathogens including enterohemorrhagic E. coli, Staphylococcus aureus, and P. aeruginosa. Cisplatin was also highly effective against clinical isolates of S. aureus and P. aeruginosa. Therefore, cisplatin has broad spectrum activity against persister cells. Biotechnol. Bioeng. 2016;113: 1984-1992. © 2016 Wiley Periodicals, Inc. PMID:26914280

  11. Electron microscopy study of antioxidant interaction with bacterial cells

    Science.gov (United States)

    Plotnikov, Oleg P.; Novikova, Olga V.; Konnov, Nikolai P.; Korsukov, Vladimir N.; Gunkin, Ivan F.; Volkov, Uryi P.

    2000-10-01

    To maintain native microorganisms genotype and phenotype features a lyophylization technique is widely used. However in this case cells are affected by influences of vacuum and low temperature that cause a part of the cells population to be destruction. Another factor reduced microorganisms vitality is formation of reactive oxygen forms that damage certain biological targets (such as DNA, membranes etc.) Recently to raise microorganism's resistance against adverse condition natural and synthetic antioxidants are used. Antioxidant- are antagonists of free radicals. Introduction of antioxidants in protective medium for lyophylization increase bacteria storage life about 2,0-4,8 fold in comparison with reference samples. In the article the main results of our investigation of antioxidants interaction with microorganism cells is described. As bacteria cells we use vaccine strain yersinia pestis EV, that were grown for 48 h at 28 degree(s)C on the Hottinger agar (pH 7,2). Antioxidants are inserted on the agar surface in specimen under test. To investigate a localization of antioxidants for electron microscopy investigation, thallium organic antioxidants were used. The thallium organic compounds have an antioxidant features if thallium is in low concentration (about 1(mu) g/ml). The localization of the thallium organic antioxidants on bacteria Y. pestis EV is visible in electron microscopy images, thallium being heavy metal with high electron density. The negatively stained bacteria and bacteria thin sections with thallium organic compounds were investigated by means of transmission electron microscopy. The localization of the thallium organic compounds is clearly visible in electron micrographs as small dark spots with size about 10-80nm. Probably mechanisms of interaction of antioxidants with bacteria cells are discussed.

  12. Induction of delayed-type hypersensitivity by the T cell line specific to bacterial peptidoglycans

    International Nuclear Information System (INIS)

    A T cell line specific for the chemically well-defined peptidoglycan of bacterial cell wall, disaccharide tetrapeptide, was established from Lewis rats immunized with the antigen covalently linked to the autologous rat serum albumin. The antigen specificity was examined with various analogues or derivatives of the peptidoglycan. The cell line was reactive to analogues with the COOH-terminal D-amino acid, but least reactive to those with L-amino acid as COOH terminus. Transferring of the T cell line into X-irradiated normal Lewis rats induced delayed-type hypersensitivity in an antigen specific manner

  13. Analysis of gene expression levels in individual bacterial cells without image segmentation

    International Nuclear Information System (INIS)

    Highlights: ► We present a method for extracting gene expression data from images of bacterial cells. ► The method does not employ cell segmentation and does not require high magnification. ► Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. ► We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

  14. Analysis of gene expression levels in individual bacterial cells without image segmentation

    Energy Technology Data Exchange (ETDEWEB)

    Kwak, In Hae; Son, Minjun [Physics Department, University of Florida, P.O. Box 118440, Gainesville, FL 32611-8440 (United States); Hagen, Stephen J., E-mail: sjhagen@ufl.edu [Physics Department, University of Florida, P.O. Box 118440, Gainesville, FL 32611-8440 (United States)

    2012-05-11

    Highlights: Black-Right-Pointing-Pointer We present a method for extracting gene expression data from images of bacterial cells. Black-Right-Pointing-Pointer The method does not employ cell segmentation and does not require high magnification. Black-Right-Pointing-Pointer Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. Black-Right-Pointing-Pointer We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

  15. Mutagenic effect of accelerated heavy ions on bacterial cells

    Science.gov (United States)

    Boreyko, A. V.; Krasavin, E. A.

    2011-11-01

    The heavy ion accelerators of the Joint Institute for Nuclear Research were used to study the regularities and mechanisms of formation of different types of mutations in prokaryote cells. The induction of direct (lac-, ton B-, col B) mutations for Esherichia coli cells and reverse his- → His+ mutations of Salmonella typhimurium, Bacillus subtilis cells under the action of radiation in a wide range of linear energy transfer (LET) was studied. The regularities of formation of gene and structural (tonB trp-) mutations for Esherichia coli bacteria under the action of accelerated heavy ions were studied. It was demonstrated that the rate of gene mutations as a function of the dose under the action of Γ rays and accelerated heavy ions is described by linear-quadratic functions. For structural mutations, linear "dose-effect" dependences are typical. The quadratic character of mutagenesis dose curves is determined by the "interaction" of two independent "hitting" events in the course of SOS repair of genetic structures. The conclusion made was that gene mutations under the action of accelerated heavy ions are induced by δ electron regions of charged particle tracks. The methods of SOS chromotest, SOS lux test, and λ prophage induction were used to study the regularities of SOS response of cells under the action of radiations in a wide LET range. The following proposition was substantiated: the molecular basis for formation of gene mutations are cluster single-strand DNA breaks, and that for structural mutations, double-strand DNA breaks. It was found out that the LET dependence of the relative biological efficiency of accelerated ions is described by curves with a local maximum. It was demonstrated that the biological efficiency of ionizing radiations with different physical characteristics on cells with different genotype, estimated by the lethal action, induction of gene and deletion mutations, precision excision of transposons, is determined by the specific

  16. Involvement of viral envelope GP2 in Ebola virus entry into cells expressing the macrophage galactose-type C-type lectin

    Energy Technology Data Exchange (ETDEWEB)

    Usami, Katsuaki [Laboratory of Cancer Biology and Molecular Immunology, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo 113-0033 (Japan); Matsuno, Keita; Igarashi, Manabu [Department of Global Epidemiology, Hokkaido University Research Center for Zoonosis Control, Sapporo 001-0020 (Japan); Denda-Nagai, Kaori [Laboratory of Cancer Biology and Molecular Immunology, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo 113-0033 (Japan); Takada, Ayato [Department of Global Epidemiology, Hokkaido University Research Center for Zoonosis Control, Sapporo 001-0020 (Japan); Irimura, Tatsuro, E-mail: irimura@mol.f.u-tokyo.ac.jp [Laboratory of Cancer Biology and Molecular Immunology, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo 113-0033 (Japan)

    2011-04-01

    Highlights: {yields} Ebola virus infection is mediated by binding to and fusion with the target cells. {yields} Structural feature of the viral glycoprotein determines the infectivity. {yields} Surface C-type lectin, MGL, of macrophages and dendritic cells mediate the infection. {yields} GP2, one of glycoprotein subunits, plays an essential role in MGL-mediated infection. {yields} There is a critical amino acid residue involved in high infectivity. -- Abstract: Ebola virus (EBOV) infection is initiated by the interaction of the viral surface envelope glycoprotein (GP) with the binding sites on target cells. Differences in the mortality among different species of the Ebola viruses, i.e., Zaire ebolavirus (ZEBOV) and Reston ebolavirus (REBOV), correspond to the in vitro infectivity of the pseudo-typed virus constructed with the GPs in cells expressing macrophage galactose-type calcium-type lectin (MGL/CD301). Through mutagenesis of GP2, the transmembrane-anchored subunit of GP, we found that residues 502-527 of the GP2 sequence determined the different infectivity between VSV-ZEBOV GP and -REBOV GP in MGL/CD301-expressing cells and a histidine residue at position 516 of ZEBOV GP2 appeared essential in the differential infectivity. These findings may provide a clue to clarify a molecular basis of different pathogenicity among EBOV species.

  17. Involvement of viral envelope GP2 in Ebola virus entry into cells expressing the macrophage galactose-type C-type lectin

    International Nuclear Information System (INIS)

    Highlights: → Ebola virus infection is mediated by binding to and fusion with the target cells. → Structural feature of the viral glycoprotein determines the infectivity. → Surface C-type lectin, MGL, of macrophages and dendritic cells mediate the infection. → GP2, one of glycoprotein subunits, plays an essential role in MGL-mediated infection. → There is a critical amino acid residue involved in high infectivity. -- Abstract: Ebola virus (EBOV) infection is initiated by the interaction of the viral surface envelope glycoprotein (GP) with the binding sites on target cells. Differences in the mortality among different species of the Ebola viruses, i.e., Zaire ebolavirus (ZEBOV) and Reston ebolavirus (REBOV), correspond to the in vitro infectivity of the pseudo-typed virus constructed with the GPs in cells expressing macrophage galactose-type calcium-type lectin (MGL/CD301). Through mutagenesis of GP2, the transmembrane-anchored subunit of GP, we found that residues 502-527 of the GP2 sequence determined the different infectivity between VSV-ZEBOV GP and -REBOV GP in MGL/CD301-expressing cells and a histidine residue at position 516 of ZEBOV GP2 appeared essential in the differential infectivity. These findings may provide a clue to clarify a molecular basis of different pathogenicity among EBOV species.

  18. Comparison of bacterial cells and amine-functionalized abiotic surfaces as support for Pd nanoparticle synthesis

    DEFF Research Database (Denmark)

    De Corte, Simon; Bechstein, Stefanie; Lokanathan, Arcot R.;

    2013-01-01

    An increasing demand for catalytic Pd nanoparticles has motivated the search for sustainable production methods. An innovative approach uses bacterial cells as support material for synthesizing Pd nanoparticles by reduction of Pd(II) with e.g. hydrogen or formate. Nevertheless, drawbacks of...... on these surfaces was higher than for Pd particles formed on Shewanella oneidensis cells. Smaller Pd nanoparticles generally have better catalytic properties, and previous studies have shown that the particle size can be lowered by increasing the amount of support material used during Pd particle...... materials were visualized by transmission electron microscopy, and their activity was evaluated by catalysis of p-nitrophenol reduction. Surfaces functionalized with 3-aminopropyltriethoxysilane and chitosan were interesting alternatives to bacterial cells, as the catalytic activity of Pd particles formed...

  19. Origin of diderm (Gram-negative) bacteria: antibiotic selection pressure rather than endosymbiosis likely led to the evolution of bacterial cells with two membranes

    OpenAIRE

    Gupta, Radhey S.

    2011-01-01

    The prokaryotic organisms can be divided into two main groups depending upon whether their cell envelopes contain one membrane (monoderms) or two membranes (diderms). It is important to understand how these and other variations that are observed in the cell envelopes of prokaryotic organisms have originated. In 2009, James Lake proposed that cells with two membranes (primarily Gram-negative bacteria) originated from an ancient endosymbiotic event involving an Actinobacteria and a Clostridia (...

  20. Bacterial glycosidases for the production of universal red blood cells

    DEFF Research Database (Denmark)

    Liu, Qiyong P; Sulzenbacher, Gerlind; Yuan, Huaiping;

    2007-01-01

    Enzymatic removal of blood group ABO antigens to develop universal red blood cells (RBCs) was a pioneering vision originally proposed more than 25 years ago. Although the feasibility of this approach was demonstrated in clinical trials for group B RBCs, a major obstacle in translating this...... alpha-N-acetylgalactosaminidase family reveals an unusual catalytic mechanism involving NAD+. The enzymatic conversion processes we describe hold promise for achieving the goal of producing universal RBCs, which would improve the blood supply while enhancing the safety of clinical transfusions....

  1. Nuclear Envelope, Nuclear Lamina, and Inherited Disease

    OpenAIRE

    Worman, Howard; Courvalin, Jean-Claude

    2005-01-01

    The nuclear envelope is composed of the nuclear membranes, nuclear lamina, and nuclear pore complexes. In recent years, mutations in nuclear-envelope proteins have been shown to cause a surprisingly wide array of inherited diseases. While the mutant proteins are generally expressed in most or all differentiated somatic cells, many mutations cause fairly tissue-specific disorders. Perhaps the most dramatic case is that of mutations in A-type lamins, intermediate filament proteins associated wi...

  2. Envelopes of Commutative Rings

    Institute of Scientific and Technical Information of China (English)

    Rafael PARRA; Manuel SAOR(I)N

    2012-01-01

    Given a significative class F of commutative rings,we study the precise conditions under which a commutative ring R has an F-envelope.A full answer is obtained when.F is the class of fields,semisimple commutative rings or integral domains.When F is the class of Noetherian rings,we give a full answer when the Krull dimension of R is zero and when the envelope is required to be epimorphic.The general problem is reduced to identifying the class of non-Noetherian rings having a monomorphic Noetherian envelope,which we conjecture is the empty class.

  3. Attachment and invasion of Neisseria meningitidis to host cells is related to surface hydrophobicity, bacterial cell size and capsule.

    Directory of Open Access Journals (Sweden)

    Stephanie N Bartley

    Full Text Available We compared exemplar strains from two hypervirulent clonal complexes, strain NMB-CDC from ST-8/11 cc and strain MC58 from ST-32/269 cc, in host cell attachment and invasion. Strain NMB-CDC attached to and invaded host cells at a significantly greater frequency than strain MC58. Type IV pili retained the primary role for initial attachment to host cells for both isolates regardless of pilin class and glycosylation pattern. In strain MC58, the serogroup B capsule was the major inhibitory determinant affecting both bacterial attachment to and invasion of host cells. Removal of terminal sialylation of lipooligosaccharide (LOS in the presence of capsule did not influence rates of attachment or invasion for strain MC58. However, removal of either serogroup B capsule or LOS sialylation in strain NMB-CDC increased bacterial attachment to host cells to the same extent. Although the level of inhibition of attachment by capsule was different between these strains, the regulation of the capsule synthesis locus by the two-component response regulator MisR, and the level of surface capsule determined by flow cytometry were not significantly different. However, the diplococci of strain NMB-CDC were shown to have a 1.89-fold greater surface area than strain MC58 by flow cytometry. It was proposed that the increase in surface area without changing the amount of anchored glycolipid capsule in the outer membrane would result in a sparser capsule and increase surface hydrophobicity. Strain NMB-CDC was shown to be more hydrophobic than strain MC58 using hydrophobicity interaction chromatography and microbial adhesion-to-solvents assays. In conclusion, improved levels of adherence of strain NMB-CDC to cell lines was associated with increased bacterial cell surface and surface hydrophobicity. This study shows that there is diversity in bacterial cell surface area and surface hydrophobicity within N. meningitidis which influence steps in meningococcal pathogenesis.

  4. Predominance of single bacterial cells in composting bioaerosols

    Science.gov (United States)

    Galès, Amandine; Bru-Adan, Valérie; Godon, Jean-Jacques; Delabre, Karine; Catala, Philippe; Ponthieux, Arnaud; Chevallier, Michel; Birot, Emmanuel; Steyer, Jean-Philippe; Wéry, Nathalie

    2015-04-01

    Bioaerosols emitted from composting plants have become an issue because of their potential harmful impact on public or workers' health. Accurate knowledge of the particle-size distribution in bioaerosols emitted from open-air composting facilities during operational activity is a requirement for improved modeling of air dispersal. In order to investigate the aerodynamic diameter of bacteria in composting bioaerosols this study used an Electrical Low Pressure Impactor for sampling and quantitative real-time PCR for quantification. Quantitative PCR results show that the size of bacteria peaked between 0.95 μm and 2.4 μm and that the geometric mean diameter of the bacteria was 1.3 μm. In addition, total microbial cells were counted by flow cytometry and revealed that these qPCR results corresponded to single whole bacteria. Finally, the enumeration of cultivable thermophilic microorganisms allowed us to set the upper size limit for fragments at an aerodynamic diameter of ∼0.3 μm. Particle-size distributions of microbial groups previously used to monitor composting bioaerosols were also investigated. In collected the bioaerosols, the aerodynamic diameter of the actinomycetes Saccharopolyspora rectivirgula-and-relatives and also of the fungus Aspergillus fumigatus, appeared to be consistent with a majority of individual cells. Together, this study provides the first culture-independent data on particle-size distribution of composting bioaerosols and reveals that airborne single bacteria were emitted predominantly from open-air composting facilities.

  5. Lipid II: a central component in bacterial cell wall synthesis and a target for antibiotics.

    Science.gov (United States)

    de Kruijff, Ben; van Dam, Vincent; Breukink, Eefjan

    2008-01-01

    The bacterial cell wall is mainly composed of peptidoglycan, which is a three-dimensional network of long aminosugar strands located on the exterior of the cytoplasmic membrane. These strands consist of alternating MurNAc and GlcNAc units and are interlinked to each other via peptide moieties that are attached to the MurNAc residues. Peptidoglycan subunits are assembled on the cytoplasmic side of the bacterial membrane on a polyisoprenoid anchor and one of the key components in the synthesis of peptidoglycan is Lipid II. Being essential for bacterial cell survival, it forms an attractive target for antibacterial compounds such as vancomycin and several lantibiotics. Lipid II consists of one GlcNAc-MurNAc-pentapeptide subunit linked to a polyiosoprenoid anchor 11 subunits long via a pyrophosphate linker. This review focuses on this special molecule and addresses three questions. First, why are special lipid carriers as polyprenols used in the assembly of peptidoglycan? Secondly, how is Lipid II translocated across the bacterial cytoplasmic membrane? And finally, how is Lipid II used as a receptor for lantibiotics to kill bacteria? PMID:19008088

  6. PROCALCITONIN AS A BIOMARKER OF BACTERIAL INFECTION IN SICKLE CELL VASO-OCCLUSIVE CRISIS.

    Directory of Open Access Journals (Sweden)

    Dilip Kumar Patel

    2014-02-01

    Full Text Available Bacterial infection is an important trigger of vaso-occlusive crisis (VOC in sickle cell anaemia (SCA. SCA Patients with VOC have signs of inflammation and it is difficult to diagnose bacterial infection in them. This study was undertaken to evaluate serum procalcitonin (PCT as a biomarker of bacterial infection in acute sickle cell vaso-occlusive crisis. Hundred SCA patients were studied at Sickle Cell Clinic and Molecular Biology Laboratory, V.S.S. Medical College, Burla, Odisha, India. SCA was diagnosed by haemoglobin electrophoresis, HPLC and molecular analysis. Patients were divided into 3categories namely Category-A (VOC/ACS with fever but without evidence of bacterial infection-66 patients; Category-B (VOC with fever and documentedbacterial infection-24 patients; and Category-C (Patients in steady statewithout VOC/ACS or fever-10 patients. Investigations like complete blood count, C-reactive protein estimation and PCT measurement was done in all the cases. There was no significant difference in total leucocytes count and C-reactiveprotein values between category A and B. In category A the PCT level was 0.5ng/mL with 87.5% of cases having >2ng/mL. In category C, PCT value was 2ng/mL is indicative of bacterial infection necessitating antimicrobial therapy. Patients with indeterminate PCT value of0.5-2ng/mL, need a repeat PCT estimation or an empirical antibiotic therapyawaiting the availability of microbiological report as deemed necessary.

  7. Viability of adhered bacterial cells: tracking MinD protein oscillations

    Science.gov (United States)

    Barrett, Matt; Colville, Keegan; Schultz-Nielsen, Chris; Jericho, Manfred; Dutcher, John

    2010-03-01

    To study bacterial cells using atomic force microscopy, it is necessary to immobilize the cells on a substrate. Because bacterial cells and common substrates such as glass and mica have a net negative charge, positively charged polymers such as poly-L-lysine (PLL) and polyethyleneimine (PEI) are commonly used as adhesion layers. However, the use of adhesion polymers could stress the cell and even render it inviable. Viable E. coli cells use oscillations of Min proteins along the axis of the rod-shaped cells to ensure accurate cell division. By tagging MinD proteins with GFP, oscillations can be observed using fluorescence microscopy. For a healthy cell in an ideal environment, the oscillation period is measured to be ˜40 s. Prior experiments have shown that PLL increases the oscillation period significantly (up to 80%). In the present study, we have used epifluorescence and total internal reflection fluorescence (TIRF) to track MinD protein oscillations in E. coli bacteria adhered to a variety of positively charged polymers on mica as a function of polymer surface coverage.

  8. A STUDY ON BACTERIAL CONTAMINATION OF CELL PHONES OF HEALTH CARE WORKERS (HCW) AT KIMS HOSPITAL, AMALAPURAM

    OpenAIRE

    Padmaja; Nageswara Rao

    2014-01-01

    A random 100 samples of cell phones of HCWs workers and Doctors working in various wards, OPDs and laboratories, Blood Bank, Causality, ICU of KIMS Hospital were subjected to bacterial analysis by conventional methods to know about the Bacterial contamination of the cell phones from July 2011 to December 2011. The commonest organism isolated from the contaminated cell phone is MRSA - 20%, followed by MSSA - 5%, CONS - 10 %, Micrococcus - 15 %. The obvious observation is no...

  9. Characterization of the pattern of alphas1- and beta-casein breakdown and release of a bioactive peptide by a cell envelope proteinase from Lactobacillus delbrueckii subsp. lactis CRL 581.

    Science.gov (United States)

    Hebert, Elvira María; Mamone, Gianfranco; Picariello, Gianluca; Raya, Raúl R; Savoy, Graciela; Ferranti, Pasquale; Addeo, Francesco

    2008-06-01

    The cell envelope-associated proteinases (CEPs) of the lactobacilli have key roles in bacterial nutrition and contribute to the development of the organoleptic properties of fermented milk products as well, as they can release bioactive health-beneficial peptides from milk proteins. The influence of the peptide supply, carbohydrate source, and osmolites on the CEP activity of the cheese starter Lactobacillus delbrueckii subsp. lactis CRL 581 was investigated. The CEP activity levels were controlled by the peptide content of the growth medium. The maximum activity was observed in a basal minimal defined medium, whereas in the presence of Casitone, Casamino Acids, or yeast extract, the synthesis of CEP was inhibited 99-, 70-, and 68-fold, respectively. The addition of specific di- or tripeptides containing branched-chain amino acids, such as leucylleucine, prolylleucine, leucylglycylglycine, or leucylproline, to the growth medium negatively affected CEP activity, whereas dipeptides without branched-chain amino acids had no effect on the enzyme's production. The carbon source and osmolites did not affect CEP activity. The CEP of L. delbrueckii subsp. lactis CRL 581 exhibited a mixed-type CEP(I/III) variant caseinolytic specificity. Mass-spectrometric screening of the main peptide peaks isolated by reverse-phase high-pressure liquid chromatography allowed the identification of 33 and 32 peptides in the alpha(s1)- and beta-casein hydrolysates, respectively. By characterizing the peptide sequence in these hydrolysates, a pattern of alpha(s1)- and beta-casein breakdown was defined and is reported herein, this being the first report for a CEP of L. delbrueckii subsp. lactis. In this pattern, a series of potentially bioactive peptides (antihypertensive and phosphopeptides) which are encrypted within the precursor protein could be visualized. PMID:18424544

  10. Single-cell-based sensors and synchrotron FTIR spectroscopy: a hybrid system towards bacterial detection.

    Science.gov (United States)

    Veiseh, Mandana; Veiseh, Omid; Martin, Michael C; Bertozzi, Carolyn; Zhang, Miqin

    2007-09-30

    Microarrays of single macrophage cell-based sensors were developed and demonstrated for potential real-time bacterium detection by synchrotron FTIR microscopy. The cells were patterned on gold electrodes of silicon oxide substrates by a surface engineering technique, in which the gold electrodes were immobilized with fibronectin to mediate cell adhesion and the silicon oxide background was passivated with polyethylene glycol (PEG) to resist protein adsorption and cell adhesion. Cell morphology and IR spectra of single, double, and triple cells on gold electrodes exposed to lipopolysaccharide (LPS) of different concentrations were compared to reveal the detection capability of this cell-based sensing platform. The single-cell-based system was found to generate the most significant and consistent IR spectrum shifts upon exposure to LPS, thus providing the highest detection sensitivity. Changes in cell morphology and IR shifts upon cell exposure to LPS were found to be dependent on the LPS concentration and exposure time, which established a method for the identification of LPS concentration and infected cell population. Possibility of using this single-cell system with conventional IR spectroscopy as well as its limitation was investigated by comparing IR spectra of single-cell arrays with gold electrode surface areas of 25, 100, and 400 microm2 using both synchrotron and conventional FTIR spectromicroscopes. This cell-based platform may potentially provide real-time, label-free, and rapid bacterial detection, and allow for high-throughput statistical analyses, and portability. PMID:17560777

  11. Convergent development of anodic bacterial communities in microbial fuel cells.

    KAUST Repository

    Yates, Matthew D

    2012-05-10

    Microbial fuel cells (MFCs) are often inoculated from a single wastewater source. The extent that the inoculum affects community development or power production is unknown. The stable anodic microbial communities in MFCs were examined using three inocula: a wastewater treatment plant sample known to produce consistent power densities, a second wastewater treatment plant sample, and an anaerobic bog sediment. The bog-inoculated MFCs initially produced higher power densities than the wastewater-inoculated MFCs, but after 20 cycles all MFCs on average converged to similar voltages (470±20 mV) and maximum power densities (590±170 mW m(-2)). The power output from replicate bog-inoculated MFCs was not significantly different, but one wastewater-inoculated MFC (UAJA3 (UAJA, University Area Joint Authority Wastewater Treatment Plant)) produced substantially less power. Denaturing gradient gel electrophoresis profiling showed a stable exoelectrogenic biofilm community in all samples after 11 cycles. After 16 cycles the predominance of Geobacter spp. in anode communities was identified using 16S rRNA gene clone libraries (58±10%), fluorescent in-situ hybridization (FISH) (63±6%) and pyrosequencing (81±4%). While the clone library analysis for the underperforming UAJA3 had a significantly lower percentage of Geobacter spp. sequences (36%), suggesting that a predominance of this microbe was needed for convergent power densities, the lower percentage of this species was not verified by FISH or pyrosequencing analyses. These results show that the predominance of Geobacter spp. in acetate-fed systems was consistent with good MFC performance and independent of the inoculum source.

  12. Repeatability of differential goat bulk milk culture and associations with somatic cell count, total bacterial count, and standard plate count

    NARCIS (Netherlands)

    Koop, G.; Dik, N.; Nielen, M.; Lipman, L.J.A.

    2010-01-01

    The aims of this study were to assess how different bacterial groups in bulk milk are related to bulk milk somatic cell count (SCC), bulk milk total bacterial count (TBC), and bulk milk standard plate count (SPC) and to measure the repeatability of bulk milk culturing. On 53 Dutch dairy goat farms,

  13. Room temperature electrocompetent bacterial cells improve DNA transformation and recombineering efficiency.

    Science.gov (United States)

    Tu, Qiang; Yin, Jia; Fu, Jun; Herrmann, Jennifer; Li, Yuezhong; Yin, Yulong; Stewart, A Francis; Müller, Rolf; Zhang, Youming

    2016-01-01

    Bacterial competent cells are essential for cloning, construction of DNA libraries, and mutagenesis in every molecular biology laboratory. Among various transformation methods, electroporation is found to own the best transformation efficiency. Previous electroporation methods are based on washing and electroporating the bacterial cells in ice-cold condition that make them fragile and prone to death. Here we present simple temperature shift based methods that improve DNA transformation and recombineering efficiency in E. coli and several other gram-negative bacteria thereby economizing time and cost. Increased transformation efficiency of large DNA molecules is a significant advantage that might facilitate the cloning of large fragments from genomic DNA preparations and metagenomics samples. PMID:27095488

  14. A Communal Bacterial Adhesin Anchors Biofilm and Bystander Cells to Surfaces

    OpenAIRE

    Absalon, Cedric; Van Dellen, Katrina; Paula I. Watnick

    2011-01-01

    Author Summary The bacterial multilayer biofilm consists of matrix-enclosed cells attached to each other to form large aggregates. The base of these aggregates may be attached to a living or non-living surface. The biofilm matrix most often contains at least one exopolysaccharide component and may also contain protein and DNA. While much is known about the exopolysaccharide component of the Gram-negative biofilm matrix, little is known about the function of biofilm matrix proteins. We hypothe...

  15. Mitomycin resistance in mammalian cells expressing the bacterial mitomycin C resistance protein MCRA

    OpenAIRE

    Belcourt, Michael F.; Penketh, Philip G.; Hodnick, William F.; Johnson, David A.; David H Sherman; Rockwell, Sara; Sartorelli, Alan C.

    1999-01-01

    The mitomycin C-resistance gene, mcrA, of Streptomyces lavendulae produces MCRA, a protein that protects this microorganism from its own antibiotic, the antitumor drug mitomycin C. Expression of the bacterial mcrA gene in mammalian Chinese hamster ovary cells causes profound resistance to mitomycin C and to its structurally related analog porfiromycin under aerobic conditions but produces little change in drug sensitivity under hypoxia. The mitomycins are prodrugs that are enzymatically reduc...

  16. Testing an agent-based model of bacterial cell motility: How nutrient concentration affects speed distribution

    OpenAIRE

    Garcia, Victor; Birbaumer, Mirko; Schweitzer, Frank

    2011-01-01

    We revisit a recently proposed agent-based model of active biological motion and compare its predictions with own experimental findings for the speed distribution of bacterial cells, \\emph{Salmonella typhimurium}. Agents move according to a stochastic dynamics and use energy stored in an internal depot for metabolism and active motion. We discuss different assumptions of how the conversion from internal to kinetic energy $d(v)$ may depend on the actual speed, to conclude that $d_{2}v^{\\xi}$ w...

  17. Quantitative analysis of initial adhesion of bacterial vaginosis anaerobes in ME-180 cells

    OpenAIRE

    Machado, António; Salgueiro, Débora; Harwich, Michael; Jefferson, Kimberly K; Cerca, Nuno

    2013-01-01

    Bacterial vaginosis (BV) is the most common vaginal disorder in women of reproductive age [1]. Despite decades of research, the etiology of BV still remains elusive. It is well established that adhesion to host cells or tissues is a necessary early step in the establishment of infection [2]. Since it is often considered a polymicrobial condition, it has been proposed that some bacteria have a preponderant role as early colonizers, while others have an impact later in the development of a mu...

  18. Radiosensitization of hypoxic bacterial cells and animal tumours by membrane active drugs and hyperthermia

    International Nuclear Information System (INIS)

    The present report deals with the results on phenothiazine derivatives such as promethazine (PMZ), trimeprazine (TMZ), trifluoperazine (TFP) and prochlorperazine (PCP) and their comparison with that of chlorpromazine (CPZ). Their efficiency in combination with hyperthermia, radiation and other anti-cancer drugs in treating murine tumors has also been presented herein. In addition, results on bacterial cells dealing with their mechanistic aspects are also included. (author). 57 refs., 27 figures, 13 tables

  19. Quantifying bacterial adhesion on antifouling polymer brushes via single-cell force spectroscopy

    Czech Academy of Sciences Publication Activity Database

    Rodriguez-Emmenegger, Cesar; Janel, S.; de los Santos Pereira, Andres; Bruns, M.; Lafont, F.

    2015-01-01

    Roč. 6, č. 31 (2015), s. 5740-5751. ISSN 1759-9954 R&D Projects: GA ČR(CZ) GJ15-09368Y; GA MŠk(CZ) ED1.1.00/02.0109 Grant ostatní: OPPK(XE) CZ.2.16/3.1.00/21545 Institutional support: RVO:61389013 Keywords : antifouling polymer brushes * single-cell force spectroscopy * bacterial adhesion Subject RIV: BO - Biophysics Impact factor: 5.520, year: 2014

  20. Biosynthesis of Bacterial Cellulose/Carboxylic Multi-Walled Carbon Nanotubes for Enzymatic Biofuel Cell Application

    OpenAIRE

    Pengfei Lv; Quan Feng; Qingqing Wang; Guohui Li; Dawei Li; Qufu Wei

    2016-01-01

    Novel nanocomposites comprised of bacterial cellulose (BC) with carboxylic multi-walled carbon nanotubes (c-MWCNTs) incorporated into the BC matrix were prepared through a simple method of biosynthesis. The biocathode and bioanode for the enzyme biological fuel cell (EBFC) were prepared using BC/c-MWCNTs composite injected by laccase (Lac) and glucose oxidase (GOD) with the aid of glutaraldehyde (GA) crosslinking. Biosynthesis of BC/c-MWCNTs composite was characterized by digital photos, scan...

  1. Instrumental analysis of bacterial cells using vibrational and emission Moessbauer spectroscopic techniques

    International Nuclear Information System (INIS)

    In biosciences and biotechnology, the expanding application of physicochemical approaches using modern instrumental techniques is an efficient strategy to obtain valuable and often unique information at the molecular level. In this work, we applied a combination of vibrational (Fourier transform infrared (FTIR), FT-Raman) spectroscopic techniques, useful in overall structural and compositional analysis of bacterial cells of the rhizobacterium Azospirillum brasilense, with 57Co emission Moessbauer spectroscopy (EMS) used for sensitive monitoring of metal binding and further transformations in live bacterial cells. The information obtained, together with ICP-MS analyses for metals taken up by the bacteria, is useful in analysing the impact of the environmental conditions (heavy metal stress) on the bacterial metabolism and some differences in the heavy metal stress-induced behaviour of non-endophytic (Sp7) and facultatively endophytic (Sp245) strains. The results show that, while both strains Sp7 and Sp245 take up noticeable and comparable amounts of heavy metals from the medium (0.12 and 0.13 mg Co, 0.48 and 0.44 mg Cu or 4.2 and 2.1 mg Zn per gram of dry biomass, respectively, at a metal concentration of 0.2 mM in the medium), their metabolic responses differ essentially. Whereas for strain Sp7 the FTIR measurements showed significant accumulation of polyhydroxyalkanoates as storage materials involved in stress endurance, strain Sp245 did not show any major changes in cellular composition. Nevertheless, EMS measurements showed rapid binding of cobalt(II) by live bacterial cells (chemically similar to metal binding by dead bacteria) and its further transformation in the live cells within an hour

  2. Instrumental analysis of bacterial cells using vibrational and emission Moessbauer spectroscopic techniques

    Energy Technology Data Exchange (ETDEWEB)

    Kamnev, Alexander A. [Laboratory of Biochemistry of Plant-Bacterial Symbioses, Institute of Biochemistry and Physiology of Plants and Microorganisms, Russian Academy of Sciences, 410049 Saratov (Russian Federation)]. E-mail: aakamnev@ibppm.sgu.ru; Tugarova, Anna V. [Laboratory of Biochemistry of Plant-Bacterial Symbioses, Institute of Biochemistry and Physiology of Plants and Microorganisms, Russian Academy of Sciences, 410049 Saratov (Russian Federation); Antonyuk, Lyudmila P. [Laboratory of Biochemistry of Plant-Bacterial Symbioses, Institute of Biochemistry and Physiology of Plants and Microorganisms, Russian Academy of Sciences, 410049 Saratov (Russian Federation); Tarantilis, Petros A. [Laboratory of Chemistry, Department of Science, Agricultural University of Athens, 11855 Athens (Greece); Kulikov, Leonid A. [Laboratory of Nuclear Chemistry Techniques, Department of Radiochemistry, Faculty of Chemistry, M.V. Lomonosov Moscow State University, 119992 Moscow (Russian Federation); Perfiliev, Yurii D. [Laboratory of Nuclear Chemistry Techniques, Department of Radiochemistry, Faculty of Chemistry, M.V. Lomonosov Moscow State University, 119992 Moscow (Russian Federation); Polissiou, Moschos G. [Laboratory of Chemistry, Department of Science, Agricultural University of Athens, 11855 Athens (Greece); Gardiner, Philip H.E. [Division of Chemistry, School of Science and Mathematics, Sheffield Hallam University, Sheffield S1 1WB (United Kingdom)

    2006-07-28

    In biosciences and biotechnology, the expanding application of physicochemical approaches using modern instrumental techniques is an efficient strategy to obtain valuable and often unique information at the molecular level. In this work, we applied a combination of vibrational (Fourier transform infrared (FTIR), FT-Raman) spectroscopic techniques, useful in overall structural and compositional analysis of bacterial cells of the rhizobacterium Azospirillum brasilense, with {sup 57}Co emission Moessbauer spectroscopy (EMS) used for sensitive monitoring of metal binding and further transformations in live bacterial cells. The information obtained, together with ICP-MS analyses for metals taken up by the bacteria, is useful in analysing the impact of the environmental conditions (heavy metal stress) on the bacterial metabolism and some differences in the heavy metal stress-induced behaviour of non-endophytic (Sp7) and facultatively endophytic (Sp245) strains. The results show that, while both strains Sp7 and Sp245 take up noticeable and comparable amounts of heavy metals from the medium (0.12 and 0.13 mg Co, 0.48 and 0.44 mg Cu or 4.2 and 2.1 mg Zn per gram of dry biomass, respectively, at a metal concentration of 0.2 mM in the medium), their metabolic responses differ essentially. Whereas for strain Sp7 the FTIR measurements showed significant accumulation of polyhydroxyalkanoates as storage materials involved in stress endurance, strain Sp245 did not show any major changes in cellular composition. Nevertheless, EMS measurements showed rapid binding of cobalt(II) by live bacterial cells (chemically similar to metal binding by dead bacteria) and its further transformation in the live cells within an hour.

  3. Bacterial regulatory networks—from self-organizing molecules to cell shape and patterns in bacterial communities

    OpenAIRE

    Hengge, Regine; Sourjik, Victor

    2013-01-01

    The ESF–EMBO Conference on ‘Bacterial Networks' was held in March, 2013. It brought together molecular microbiologists, bacteral systems biologists and synthetic biologists to discuss the architecture, function and dynamics of regulatory networks in bacteria.

  4. Biomechanical Loading Modulates Proinflammatory and Bone Resorptive Mediators in Bacterial-Stimulated PDL Cells

    Directory of Open Access Journals (Sweden)

    Andressa Vilas Boas Nogueira

    2014-01-01

    Full Text Available The present study aimed to evaluate in vitro whether biomechanical loading modulates proinflammatory and bone remodeling mediators production by periodontal ligament (PDL cells in the presence of bacterial challenge. Cells were seeded on BioFlex culture plates and exposed to Fusobacterium nucleatum ATCC 25586 and/or cyclic tensile strain (CTS of low (CTSL and high (CTSH magnitudes for 1 and 3 days. Synthesis of cyclooxygenase-2 (COX2 and prostaglandin E2 (PGE2 was evaluated by ELISA. Gene expression and protein secretion of osteoprotegerin (OPG and receptor activator of nuclear factor kappa-B ligand (RANKL were evaluated by quantitative RT-PCR and ELISA, respectively. F. nucleatum increased the production of COX2 and PGE2, which was further increased by CTS. F. nucleatum-induced increase of PGE2 synthesis was significantly (P<0.05 increased when CTSH was applied at 1 and 3 days. In addition, CTSH inhibited the F. nucleatum-induced upregulation of OPG at 1 and 3 days, thereby increasing the RANKL/OPG ratio. OPG and RANKL mRNA results correlated with the protein results. In summary, our findings provide original evidence that CTS can enhance bacterial-induced syntheses of molecules associated with inflammation and bone resorption by PDL cells. Therefore, biomechanical, such as orthodontic or occlusal, loading may enhance the bacterial-induced inflammation and destruction in periodontitis.

  5. Mineralization of Iron Oxyhydroxides in the Presence and in the Absence of Bacterial Cells

    Science.gov (United States)

    Châtellier, X.; West, M.; Rose, J.; Fortin, D.; Leppard, G. G.; Ferris, G.

    2001-12-01

    Because of their small size, iron oxides have a large surface area per unit weight ratio and are believed to play an important role as an adsorbing phase in lake sediments for various molecules, including potentially dangerous ones like heavy metals. They have been observed to form in close association with bacterial cells, by oxidation of ferrous ions. It is thus important to determine whether the presence of the bacterial cells can affect the mineralogy and the mesoscopic structure of the Fe-oxides particles, as well as their reactivity towards heavy metals. We synthesized in the lab nanoparticles of Fe-oxides by oxidation of ferrous ions. This was done in the presence and in the absence of various bacterial strains (Escherichia coli, Bacillus subtilis, Pseudomonas Aeruginosa and Bacillus licheniformis) and of inorganic ligands (sulfate, phosphate, silicate). The Fe-oxides particles were then observed by TEM on thin sections and on whole mounts. The chemical composition was estimated by wet chemistry and by EDS. The mineralogy was determined by XRD, SAED and EXAFS. Surface area was investigated by BET. And adsorption of cadmium was also measured at various pH. We observed that the size and the morphology of the particles as well as their mesoscopic spatial organization can be affected by the presence of the cells, whereas the mineralogy is controlled by the chemistry of the solution. The adsorption isotherms of cadmium on the various Fe-oxides will be discussed at the light of these observations.

  6. Priming B cell-mediated anti-HIV envelope responses by vaccination allows for the long-term control of infection in macaques exposed to a R5-tropic SHIV

    International Nuclear Information System (INIS)

    The potential of vaccine-elicited anti-HIV envelope antibodies to control HIV-infection was evaluated by immunizing macaques with the HIV envelope protein and transiently depleting them of their CD8+ cells before intravenous challenge with the pathogenic CCR5-tropic SIV/HIV chimeric virus, SHIVSF162P4. Although sterilizing immunity was not achieved, all vaccinated animals effectively controlled infection and remained free of disease for the duration of observation (over 3 years). In contrast, during the same period, the control animals progressed to disease. Both the vaccinees and the controls developed robust cell-mediated antiviral and neutralizing antibody responses following infection. A comparative analysis of these responses suggests that the more effective long-term control of infection by the vaccinated animals is due to the more rapid development of anti-HIV envelope antibodies. These studies suggest that priming by vaccination of B cell anti-HIV envelope responses maybe crucial for the long-term control of HIV infection

  7. Temporal expression of bacterial proteins instructs host CD4 T cell expansion and Th17 development.

    Directory of Open Access Journals (Sweden)

    Seung-Joo Lee

    2012-01-01

    Full Text Available Pathogens can substantially alter gene expression within an infected host depending on metabolic or virulence requirements in different tissues, however, the effect of these alterations on host immunity are unclear. Here we visualized multiple CD4 T cell responses to temporally expressed proteins in Salmonella-infected mice. Flagellin-specific CD4 T cells expanded and contracted early, differentiated into Th1 and Th17 lineages, and were enriched in mucosal tissues after oral infection. In contrast, CD4 T cells responding to Salmonella Type-III Secretion System (TTSS effectors steadily accumulated until bacterial clearance was achieved, primarily differentiated into Th1 cells, and were predominantly detected in systemic tissues. Thus, pathogen regulation of antigen expression plays a major role in orchestrating the expansion, differentiation, and location of antigen-specific CD4 T cells in vivo.

  8. Explosive cell lysis as a mechanism for the biogenesis of bacterial membrane vesicles and biofilms.

    Science.gov (United States)

    Turnbull, Lynne; Toyofuku, Masanori; Hynen, Amelia L; Kurosawa, Masaharu; Pessi, Gabriella; Petty, Nicola K; Osvath, Sarah R; Cárcamo-Oyarce, Gerardo; Gloag, Erin S; Shimoni, Raz; Omasits, Ulrich; Ito, Satoshi; Yap, Xinhui; Monahan, Leigh G; Cavaliere, Rosalia; Ahrens, Christian H; Charles, Ian G; Nomura, Nobuhiko; Eberl, Leo; Whitchurch, Cynthia B

    2016-01-01

    Many bacteria produce extracellular and surface-associated components such as membrane vesicles (MVs), extracellular DNA and moonlighting cytosolic proteins for which the biogenesis and export pathways are not fully understood. Here we show that the explosive cell lysis of a sub-population of cells accounts for the liberation of cytosolic content in Pseudomonas aeruginosa biofilms. Super-resolution microscopy reveals that explosive cell lysis also produces shattered membrane fragments that rapidly form MVs. A prophage endolysin encoded within the R- and F-pyocin gene cluster is essential for explosive cell lysis. Endolysin-deficient mutants are defective in MV production and biofilm development, consistent with a crucial role in the biogenesis of MVs and liberation of extracellular DNA and other biofilm matrix components. Our findings reveal that explosive cell lysis, mediated through the activity of a cryptic prophage endolysin, acts as a mechanism for the production of bacterial MVs. PMID:27075392

  9. The Immunodominance Change and Protection of CD4+ T-Cell Responses Elicited by an Envelope Protein Domain III-Based Tetravalent Dengue Vaccine in Mice.

    Directory of Open Access Journals (Sweden)

    Hsin-Wei Chen

    Full Text Available Dengue is the leading cause of mosquito-borne viral infections and no vaccine is available now. Envelope protein domain III (ED3 is the major target for the binding of dengue virus neutralizing antibodies; however, the ED3-specifc T-cell response is less well understood. To investigate the T-cell responses to four serotypes of dengue virus (DENV-1 to 4, we immunized mice using either a tetravalent ED3-based DNA or protein vaccine, or combined both as a DNA prime-protein boost strategy (prime-boost. A significant serotype-dependent IFN-γ or IL-4 response was observed in mice immunized with either the DNA or protein vaccine. The IFN-γ response was dominant to DENV-1 to 3, whereas the IL-4 response was dominant to DENV-4. Although the similar IgG titers for the four serotypes were observed in mice immunized with the tetravalent vaccines, the neutralizing antibody titers varied and followed the order of 2 = 3>1>4. Interestingly, the lower IFN-γ response to DENV-4 is attributable to the immunodominance change between two CD4+ T-cell epitopes; one T-cell epitope located at E349-363 of DENV-1 to 3 was more immunogenic than the DENV-4 epitope E313-327. Despite DENV-4 specific IFN-γ responses were suppressed by immunodominance change, either DENV-4-specific IFN-γ or neutralizing antibody responses were still recalled after DENV-4 challenge and contributed to virus clearance. Immunization with the prime-boost elicited both IFN-γ and neutralizing antibody responses and provided better protection than either DNA or protein immunization. Our findings shed light on how ED3-based tetravalent dengue vaccines sharpen host CD4 T-cell responses and contribute to protection against dengue virus.

  10. Desquamated epithelial cells covered with a polymicrobial biofilm typical for bacterial vaginosis are present in randomly selected cryopreserved donor semen.

    Science.gov (United States)

    Swidsinski, Alexander; Dörffel, Yvonne; Loening-Baucke, Vera; Mendling, Werner; Verstraelen, Hans; Dieterle, Stefan; Schilling, Johannes

    2010-08-01

    We tested whether the bacterial biofilm typical for bacterial vaginosis (BV) can be found on desquamated epithelial cells in cryopreserved donor semen. Bacteria were detected with FISH. Bacterial biofilm, covering the epithelial layer in vaginal biopsies of 20 women with BV, was evaluated on desquamated epithelial cells found in the urine of these same women and their male partners (N=20) and compared with the bacterial biofilm found on desquamated epithelial cells in randomly selected cryopreserved semen samples (N=20). Urine from 20 healthy women of laboratory and clinic personnel and urine from their partners were used as controls. Desquamated epithelial cells covered with a polymicrobial Gardnerella biofilm were identified in urine samples from all women with BV and 13 of their male partners and in none of the female controls and their partners. Gardnerella biofilm, typical for BV, was found in the semen of three of the 20 donors. Donor semen might be a vector for BV. PMID:20497224

  11. Branched signal wiring of an essential bacterial cell-cycle phosphotransfer protein

    OpenAIRE

    Blair, Jimmy A.; Xu, Qingping; Childers, W. Seth; Mathews, Irimpan I.; Kern, Justin W.; Eckart, Michael; Deacon, Ashley M.; Shapiro, Lucy

    2013-01-01

    Vital to bacterial survival is the faithful propagation of cellular signals, and in Caulobacter crescentus ChpT is an essential mediator within the cell cycle circuit. ChpT functions as a histidine-containing phosphotransfer protein (HPt) that shuttles a phosphoryl group from the receiver domain of CckA, the upstream hybrid histidine kinase (HK), to one of two downstream response regulators (RRs)—CtrA or CpdR—that controls cell cycle progression. To understand how ChpT interacts with multiple...

  12. Laser capture microdissection of bacterial cells targeted by fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Schou, Kirstine Klitgaard; Mølbak, Lars; Jensen, Tim Kåre; Lindboe, Christian Frederik; Boye, Mette

    2005-01-01

    Direct cultivation-independent sequence retrieval of unidentified bacteria from histological tissue sections has been limited by the difficulty of selectively isolating specific bacteria from a complex environment. Here, a new DNA isolation approach is presented for prokaryotic cells. By this......RNA gene PCR was performed from the dissected microcolonies, and the subsequent DNA sequence analysis identified the dissected bacterial cells as belonging to the Brachyspira aalborgi cluster 1. The advantage of this technique is the ability to combine the histological recognition of the specific bacteria...

  13. Secretion of dengue virus envelope protein ectodomain from mammalian cells is dependent on domain II serotype and affects the immune response upon DNA vaccination.

    Science.gov (United States)

    Slon Campos, J L; Poggianella, M; Marchese, S; Bestagno, M; Burrone, O R

    2015-11-01

    Dengue virus (DENV) is currently among the most important human pathogens and affects millions of people throughout the tropical and subtropical regions of the world. Although it has been a World Health Organization priority for several years, there is still no efficient vaccine available to prevent infection. The envelope glycoprotein (E), exposed on the surface on infective viral particles, is the main target of neutralizing antibodies. For this reason it has been used as the antigen of choice for vaccine development efforts. Here we show a detailed analysis of factors involved in the expression, secretion and folding of E ectodomain from all four DENV serotypes in mammalian cells, and how this affects their ability to induce neutralizing antibody responses in DNA-vaccinated mice. Proper folding of E domain II (DII) is essential for efficient E ectodomain secretion, with DIII playing a significant role in stabilizing soluble dimers. We also show that the level of protein secreted from transfected cells determines the strength and efficiency of antibody responses in the context of DNA vaccination and should be considered a pivotal feature for the development of E-based DNA vaccines against DENV. PMID:26358704

  14. Conformation-specific antibodies targeting the trimer-of-hairpins motif of the human T-cell leukemia virus type 1 transmembrane glycoprotein recognize the viral envelope but fail to neutralize viral entry.

    Science.gov (United States)

    Mirsaliotis, Antonis; Nurkiyanova, Kulpash; Lamb, Daniel; Woof, Jenny M; Brighty, David W

    2007-06-01

    Human T-cell leukemia virus type 1 (HTLV-1) entry into cells is dependent upon the viral envelope glycoprotein-catalyzed fusion of the viral and cellular membranes. Following receptor activation of the envelope, the transmembrane glycoprotein (TM) is thought to undergo a series of fusogenic conformational transitions through a rod-like prehairpin intermediate to a compact trimer-of-hairpins structure. Importantly, synthetic peptides that interfere with the conformational changes of TM are potent inhibitors of membrane fusion and HTLV-1 entry, suggesting that TM is a valid target for antiviral therapy. To assess the utility of TM as a vaccine target and to explore further the function of TM in HTLV-1 pathogenesis, we have begun to examine the immunological properties of TM. Here we demonstrate that a recombinant trimer-of-hairpins form of the TM ectodomain is strongly immunogenic. Monoclonal antibodies raised against the TM immunogen specifically bind to trimeric forms of TM, including structures thought to be important for membrane fusion. Importantly, these antibodies recognize the envelope on virally infected cells but, surprisingly, fail to neutralize envelope-mediated membrane fusion or infection by pseudotyped viral particles. Our data imply that, even in the absence of overt membrane fusion, there are multiple forms of TM on virally infected cells and that some of these display fusion-associated structures. Finally, we demonstrate that many of the antibodies possess the ability to recruit complement to TM, suggesting that envelope-derived immunogens capable of eliciting a combination of neutralizing and complement-fixing antibodies would be of value as subunit vaccines for intervention in HTLV infections. PMID:17376912

  15. Study on lipid droplet dynamics in live cells and fluidity changes in model bacterial membranes using optical microscopy techniques

    OpenAIRE

    Wong, Christine Shiang Yee

    2014-01-01

    In this thesis optical microscopy techniques are used to consider aspects of viral and bacterial infections. In part 1, the physical effects of cytomegalovirus on lipid droplet dynamics in live cells are studied; in part 2, the effects of an antimicrobial peptide on the fluidity of model bacterial membranes are studied. The optical microscopy techniques used to study the effects of murine-cytomegalovirus (mCMV) on lipid droplets in live NIH/3T3 fibroblast cells in real-time are...

  16. Targeting Bacterial Cell Wall Peptidoglycan Synthesis by Inhibition of Glycosyltransferase Activity.

    Science.gov (United States)

    Mesleh, Michael F; Rajaratnam, Premraj; Conrad, Mary; Chandrasekaran, Vasu; Liu, Christopher M; Pandya, Bhaumik A; Hwang, You Seok; Rye, Peter T; Muldoon, Craig; Becker, Bernd; Zuegg, Johannes; Meutermans, Wim; Moy, Terence I

    2016-02-01

    Synthesis of bacterial cell wall peptidoglycan requires glycosyltransferase enzymes that transfer the disaccharide-peptide from lipid II onto the growing glycan chain. The polymerization of the glycan chain precedes cross-linking by penicillin-binding proteins and is essential for growth for key bacterial pathogens. As such, bacterial cell wall glycosyltransferases are an attractive target for antibiotic drug discovery. However, significant challenges to the development of inhibitors for these targets include the development of suitable assays and chemical matter that is suited to the nature of the binding site. We developed glycosyltransferase enzymatic activity and binding assays using the natural products moenomycin and vancomycin as model inhibitors. In addition, we designed a library of disaccharide compounds based on the minimum moenomycin fragment with peptidoglycan glycosyltransferase inhibitory activity and based on a more drug-like and synthetically versatile disaccharide building block. A subset of these disaccharide compounds bound and inhibited the glycosyltransferase enzymes, and these compounds could serve as chemical entry points for antibiotic development. PMID:26358369

  17. Crystal structure of MraY, an essential membrane enzyme for bacterial cell wall synthesis.

    Science.gov (United States)

    Chung, Ben C; Zhao, Jinshi; Gillespie, Robert A; Kwon, Do-Yeon; Guan, Ziqiang; Hong, Jiyong; Zhou, Pei; Lee, Seok-Yong

    2013-08-30

    MraY (phospho-MurNAc-pentapeptide translocase) is an integral membrane enzyme that catalyzes an essential step of bacterial cell wall biosynthesis: the transfer of the peptidoglycan precursor phospho-MurNAc-pentapeptide to the lipid carrier undecaprenyl phosphate. MraY has long been considered a promising target for the development of antibiotics, but the lack of a structure has hindered mechanistic understanding of this critical enzyme and the enzyme superfamily in general. The superfamily includes enzymes involved in bacterial lipopolysaccharide/teichoic acid formation and eukaryotic N-linked glycosylation, modifications that are central in many biological processes. We present the crystal structure of MraY from Aquifex aeolicus (MraYAA) at 3.3 Å resolution, which allows us to visualize the overall architecture, locate Mg(2+) within the active site, and provide a structural basis of catalysis for this class of enzyme. PMID:23990562

  18. Analysis of bone marrow stromal cell transferred bacterial β-galactosidase gene by PIXE

    International Nuclear Information System (INIS)

    PIXE, Particle Induced X-ray Emission, is a powerful, multi-elemental analysis method which has many distinguishing features and has been used in varies research fields. Recently the method of applying baby cyclotrons for nuclear medicine to PIXE has been developed. This enables us to study biomedical phenomena from the physical point of view. Mouse bone marrow stromal cells were transferred bacterial β-galactosidase gene (LacZ gene) by murine retroviral vectors. Analysis of the bone marrow stromal cells with the LacZ gene by PIXE revealed remarkable changes of intracellular trace elements compared with the normal control cells. These results indicate that gene transfer by retroviral vectors may bring about a dynamic change of intracellular circumstances of the target cell. (author)

  19. Disruption of Bacterial Cells by Photocatalysis of Montmorillonite Supported Titanium Dioxide

    Institute of Scientific and Technical Information of China (English)

    LEI Shaomin; GUO Gaoli; XIONG Bihua; GONG Wenqi; MEI Guangjun

    2009-01-01

    The photo-induced antibacterial capacity of montmorillonite supported titanium dioxide(TiO_2/Mmt for short)was evaluated by using Escherichia coli and Staphylococcus aureus as modal organisms.The bactericidal activity of TiO_2/Mmt was examined by cell viability assay under different illumination modes.Atomic force microscopy(AFM)and total organic carbon/Total nitrogen (TOC/TN)analyses were employed to investigate the mechanism of the photocatalytic bactericidal process qualitatively and quantitatively.The kinetic data show that TiO_2/Mmt has excellent antibac-terial performance,and about 99%of both bacteria cells are inactivated within 75 min illumination.The AFM images demonstrate that the bacterial cells are irreversibly decomposed and some cell components are dissolved.Therefore,the content and phase of carbon and nitrogen in the solution are changed after photocatalytic reaction.

  20. FRACTIONAL CRYSTALLIZATION FEED ENVELOPE

    International Nuclear Information System (INIS)

    Laboratory work was completed on a set of evaporation tests designed to establish a feed envelope for the fractional crystallization process. The feed envelope defines chemical concentration limits within which the process can be operated successfully. All 38 runs in the half-factorial design matrix were completed successfully, based on the qualitative definition of success. There is no feed composition likely to be derived from saltcake dissolution that would cause the fractional crystallization process to not meet acceptable performance requirements. However, some compositions clearly would provide more successful operation than other compositions

  1. A novel envelope mediated post entry restriction of murine leukaemia virus in human cells is Ref1/TRIM5α independent

    Directory of Open Access Journals (Sweden)

    McKnight Áine

    2010-10-01

    Full Text Available Abstract Background 'Intrinsic' resistance to retroviral infection was first recognised with the Friend virus susceptibility gene (Fv1, which determines susceptibility to murine leukaemia virus (MLV infection in different murine species. Similarly, the tripartite motif (TRIM family of proteins determine lentiviral restriction in a primate host-species specific manner. For example rhesus TRIM5α (rhTRIM5α can potently restrict HIV-1 infection while human TRIM5α (huTRIM5α only has a mild effect on SIVmac and HIV-1 infectivity (Lv1. Human TRIM5α is able to restrict MLV-N virus replication, but is ineffective against MLV-B or MLV-NB virus infection. Lv2 restriction of some HIV-2 viruses is seen in human cells. Like Lv1, Lv2 is a post-entry restriction factor, whose viral determinants have been mapped to the viral capsid (CA. Unlike Lv1, however, Lv2 is determined by envelope (Env in addition to CA. Here we present evidence of a novel Env determined post entry restriction to infection in human cells of pseudotyped MLV-B and MLV-NB cores. Results We generated retroviral vectors pseudotyped with various gamma and lentiviral Envs on MLV-B and -NB CAs containing a green fluorescent protein (GFP reporter. Flow cytometry was used to determine transduction efficiencies in NP2/CD4/CXCR4 (glioma cell line stably transduced with the HIV receptors and HeLa/CD4 cell lines. The HeLa/CD4 cell line restricted both MLV CAs in an Env dependent manner, compared to NP2/CD4/CXCR4 cells. Quantitative polymerase chain reaction (QT-PCR analysis of reverse transcription (RT transcripts demonstrates that this restriction occurs at a post entry and RT level. siRNA knockdown of huTRIM5α ruled out a direct role for this cellular component in mediating this restriction. We describe a previously unobserved Env determined restriction of MLV-B and MLV-NB CAs in HeLa/CD4 cells when pseudotyped with HIV-2 and RD114 Envs, but not gibbon ape leukaemia virus (GALV, HIV-1 or

  2. Solid-State NMR on bacterial cells: selective cell wall signal enhancement and resolution improvement using dynamic nuclear polarization

    International Nuclear Information System (INIS)

    Dynamic nuclear polarization (DNP) enhanced solid-state nuclear magnetic resonance (NMR) has recently emerged as a powerful technique for the study of material surfaces. In this study, we demonstrate its potential to investigate cell surface in intact cells. Using Bacillus subtilis bacterial cells as an example, it is shown that the polarizing agent 1-(TEMPO-4-oxy)-3-(TEMPO-4-amino)propan-2-ol (TOTAPOL) has a strong binding affinity to cell wall polymers (peptidoglycan). This particular interaction is thoroughly investigated with a systematic study on extracted cell wall materials, disrupted cells, and entire cells, which proved that TOTAPOL is mainly accumulating in the cell wall. This property is used on one hand to selectively enhance or suppress cell wall signals by controlling radical concentrations and on the other hand to improve spectral resolution by means of a difference spectrum. Comparing DNP-enhanced and conventional solid-state NMR, an absolute sensitivity ratio of 24 was obtained on the entire cell sample. This important increase in sensitivity together with the possibility of enhancing specifically cell wall signals and improving resolution really opens new avenues for the use of DNP-enhanced solid-state NMR as an on-cell investigation tool. (authors)

  3. Long acting β2-agonist and corticosteroid restore airway glandular cell function altered by bacterial supernatant

    Directory of Open Access Journals (Sweden)

    Nawrocki-Raby Béatrice

    2010-01-01

    Full Text Available Abstract Background Staphylococcus aureus releases virulence factors (VF that may impair the innate protective functions of airway cells. The aim of this study was to determine whether a long-acting β2 adrenergic receptor agonist (salmeterol hydroxynaphthoate, Sal combined with a corticosteroid (fluticasone propionate, FP was able to regulate ion content and cytokine expression by airway glandular cells after exposure to S. aureus supernatant. Methods A human airway glandular cell line was incubated with S. aureus supernatant for 1 h and then treated with the combination Sal/FP for 4 h. The expression of actin and CFTR proteins was analyzed by immunofluorescence. Videomicroscopy was used to evaluate chloride secretion and X-ray microanalysis to measure the intracellular ion and water content. The pro-inflammatory cytokine expression was assessed by RT-PCR and ELISA. Results When the cells were incubated with S. aureus supernatant and then with Sal/FP, the cellular localisation of CFTR was apical compared to the cytoplasmic localisation in cells incubated with S. aureus supernatant alone. The incubation of airway epithelial cells with S. aureus supernatant reduced by 66% the chloride efflux that was fully restored by Sal/FP treatment. We also observed that Sal/FP treatment induced the restoration of ion (Cl and S and water content within the intracellular secretory granules of airway glandular cells and reduced the bacterial supernatant-dependent increase of pro-inflammatory cytokines IL8 and TNFα. Conclusions Our results demonstrate that treatment with the combination of a corticosteroid and a long-acting β2 adrenergic receptor agonist after bacterial infection restores the airway glandular cell function. Abnormal mucus induced by defective ion transport during pulmonary infection could benefit from treatment with a combination of β2 adrenergic receptor agonist and glucocorticoid.

  4. Live cell imaging of SOS and prophage dynamics in isogenic bacterial populations.

    Science.gov (United States)

    Helfrich, Stefan; Pfeifer, Eugen; Krämer, Christina; Sachs, Christian Carsten; Wiechert, Wolfgang; Kohlheyer, Dietrich; Nöh, Katharina; Frunzke, Julia

    2015-11-01

    Almost all bacterial genomes contain DNA of viral origin, including functional prophages or degenerated phage elements. A frequent but often unnoted phenomenon is the spontaneous induction of prophage elements (SPI) even in the absence of an external stimulus. In this study, we have analyzed SPI of the large, degenerated prophage CGP3 (187 kbp), which is integrated into the genome of the Gram-positive Corynebacterium glutamicum ATCC 13032. Time-lapse fluorescence microscopy of fluorescent reporter strains grown in microfluidic chips revealed the sporadic induction of the SOS response as a prominent trigger of CGP3 SPI but also displayed a considerable fraction (∼30%) of RecA-independent SPI. Whereas approx. 20% of SOS-induced cells recovered from this stress and resumed growth, the spontaneous induction of CGP3 always led to a stop of growth and likely cell death. A carbon source starvation experiment clearly emphasized that SPI only occurs in actively proliferating cells, whereas sporadic SOS induction was still observed in resting cells. These data highlight the impact of sporadic DNA damage on the activity of prophage elements and provide a time-resolved, quantitative description of SPI as general phenomenon of bacterial populations. PMID:26235130

  5. Pretreatment of Epithelial Cells with Rifaximin Alters Bacterial Attachment and Internalization Profiles▿

    Science.gov (United States)

    Brown, Eric L.; Xue, Qiong; Jiang, Zhi-Dong; Xu, Yi; DuPont, Herbert L.

    2010-01-01

    Rifaximin is a poorly absorbed semisynthetic antibiotic derivative of rifampin licensed for use in the treatment of traveler's diarrhea. Rifaximin reduces the symptoms of enteric infection, often without pathogen eradication and with limited effects on intestinal flora. Epithelial cells (HEp-2 [laryngeal], HCT-8 [ileocecal], A549 [lung], and HeLa [cervical]) were pretreated with rifaximin (or control antibiotics) prior to the addition of enteroaggregative Escherichia coli (EAEC). EAEC adherence was significantly reduced following rifaximin pretreatment compared to pretreatment with rifampin or doxycycline for three of the four cell lines tested. The rifaximin-mediated changes to epithelial cells were explored further by testing the attachment and internalization of either Bacillus anthracis or Shigella sonnei into A549 or HeLa cells, respectively. The attachment and internalization of B. anthracis were significantly reduced following rifaximin pretreatment. In contrast, neither the attachment nor the internalization of S. sonnei was affected by rifaximin pretreatment of HeLa cells, suggesting that rifaximin-mediated modulation of host cell physiology affected bacteria utilizing distinct attachment/internalization mechanisms differently. In addition, rifaximin pretreatment of HEp-2 cells led to reduced concentrations of inflammatory cytokines from uninfected cells. The study provides evidence that rifaximin-mediated changes in epithelial cell physiology are associated with changes in bacterial attachment/internalization and reduced inflammatory cytokine release. PMID:19858255

  6. Nanoscale imaging of the growth and division of bacterial cells on planar substrates with the atomic force microscope

    Energy Technology Data Exchange (ETDEWEB)

    Van Der Hofstadt, M. [Institut de Bioenginyeria de Catalunya (IBEC), C/ Baldiri i Reixac 11-15, 08028 Barcelona (Spain); Hüttener, M.; Juárez, A. [Institut de Bioenginyeria de Catalunya (IBEC), C/ Baldiri i Reixac 11-15, 08028 Barcelona (Spain); Departament de Microbiologia, Universitat de Barcelona, Avinguda Diagonal 645, 08028 Barcelona (Spain); Gomila, G., E-mail: ggomila@ibecbarcelona.eu [Institut de Bioenginyeria de Catalunya (IBEC), C/ Baldiri i Reixac 11-15, 08028 Barcelona (Spain); Departament d' Electronica, Universitat de Barcelona, C/ Marti i Franqués 1, 08028 Barcelona (Spain)

    2015-07-15

    With the use of the atomic force microscope (AFM), the Nanomicrobiology field has advanced drastically. Due to the complexity of imaging living bacterial processes in their natural growing environments, improvements have come to a standstill. Here we show the in situ nanoscale imaging of the growth and division of single bacterial cells on planar substrates with the atomic force microscope. To achieve this, we minimized the lateral shear forces responsible for the detachment of weakly adsorbed bacteria on planar substrates with the use of the so called dynamic jumping mode with very soft cantilever probes. With this approach, gentle imaging conditions can be maintained for long periods of time, enabling the continuous imaging of the bacterial cell growth and division, even on planar substrates. Present results offer the possibility to observe living processes of untrapped bacteria weakly attached to planar substrates. - Highlights: • Gelatine coatings used to weakly attach bacterial cells onto planar substrates. • Use of the dynamic jumping mode as a non-perturbing bacterial imaging mode. • Nanoscale resolution imaging of unperturbed single living bacterial cells. • Growth and division of single bacteria cells on planar substrates observed.

  7. Characterization of the TolB-Pal trans-envelope complex from Xylella fastidiosa reveals a dynamic and coordinated protein expression profile during the biofilm development process.

    Science.gov (United States)

    Santos, Clelton A; Janissen, Richard; Toledo, Marcelo A S; Beloti, Lilian L; Azzoni, Adriano R; Cotta, Monica A; Souza, Anete P

    2015-10-01

    The intriguing roles of the bacterial Tol-Pal trans-envelope protein complex range from maintenance of cell envelope integrity to potential participation in the process of cell division. In this study, we report the characterization of the XfTolB and XfPal proteins of the Tol-Pal complex of Xylella fastidiosa. X. fastidiosa is a major plant pathogen that forms biofilms inside xylem vessels, triggering the development of diseases in important cultivable plants around the word. Based on functional complementation experiments in Escherichia coli tolB and pal mutant strains, we confirmed the role of xftolB and xfpal in outer membrane integrity. In addition, we observed a dynamic and coordinated protein expression profile during the X. fastidiosa biofilm development process. Using small-angle X-ray scattering (SAXS), the low-resolution structure of the isolated XfTolB-XfPal complex in solution was solved for the first time. Finally, the localization of the XfTolB and XfPal polar ends was visualized via immunofluorescence labeling in vivo during bacterial cell growth. Our results highlight the major role of the components of the cell envelope, particularly the TolB-Pal complex, during the different phases of bacterial biofilm development. PMID:26049080

  8. Lamin b1 polymorphism influences morphology of the nuclear envelope, cell cycle progression, and risk of neural tube defects in mice.

    Directory of Open Access Journals (Sweden)

    Sandra C P De Castro

    in maintaining integrity of the nuclear envelope and ensuring normal cell cycle progression.

  9. Total and Envelope Protein-Specific Antibody-Secreting Cell Response in Pediatric Dengue Is Highly Modulated by Age and Subsequent Infections.

    Science.gov (United States)

    Toro, Jessica F; Salgado, Doris M; Vega, Rocío; Rodríguez, Jairo A; Rodríguez, Luz-Stella; Angel, Juana; Franco, Manuel A; Greenberg, Harry B; Narváez, Carlos F

    2016-01-01

    The response of antibody-secreting cells (ASC) induced by dengue has only recently started to be characterized. We propose that young age and previous infections could be simple factors that affect this response. Here, we evaluated the primary and secondary responses of circulating ASC in infants (6-12 months old) and children (1-14 years old) infected with dengue showing different degrees of clinical severity. The ASC response was delayed and of lower magnitude in infants, compared with older children. In primary infection (PI), the total and envelope (E) protein-specific IgM ASC were dominant in infants but not in children, and a negative correlation was found between age and the number of IgM ASC (rho = -0.59, P = 0.03). However, infants with plasma dengue-specific IgG detectable in the acute phase developed an intense ASC response largely dominated by IgG and comparable to that of children with secondary infection (SI). IgM and IgG produced by ASC circulating in PI or SI were highly cross-reactive among the four serotypes. Dengue infection caused the disturbance of B cell subsets, particularly a decrease in the relative frequency of naïve B cells. Higher frequencies of total and E protein-specific IgM ASC in the infants and IgG in the children were associated with clinically severe forms of infection. Therefore, the ASC response induced by dengue is highly influenced by the age at which infection occurs and previous immune status, and its magnitude is a relevant element in the clinical outcome. These results are important in the search for correlates of protection and for determining the ideal age for vaccinating against dengue. PMID:27560782

  10. Envelope Gene of the Human Endogenous Retrovirus HERV-W Encodes a Functional Retrovirus Envelope

    OpenAIRE

    An, Dong Sung; Xie, Yi-ming; Chen, Irvin S. Y.

    2001-01-01

    A member of the human endogenous retrovirus (HERV) family termed HERV-W encodes a highly fusogenic membrane glycoprotein that appears to be expressed specifically in the placenta. It is unclear whether the glycoproteins of the HERVs can serve as functional retrovirus envelope proteins to confer infectivity on retrovirus particles. We found that the HERV-W envelope glycoprotein can form pseudotypes with human immunodeficiency virus type 1 virions and confers tropism for CD4-negative cells. Thu...

  11. Selective induction of cell-mediated immunity and protection of rhesus macaques from chronic SHIVKU2 infection by prophylactic vaccination with a conserved HIV-1 envelope peptide-cocktail

    International Nuclear Information System (INIS)

    Infection of Indian-origin rhesus macaques by the simian human immunodeficiency virus (SHIV) is considered to be a suitable preclinical model for directly testing efficacy of vaccine candidates based on the HIV-1 envelope. We used this model for prophylactic vaccination with a peptide-cocktail comprised of highly conserved HIV-1 envelope sequences immunogenic/antigenic in macaques and humans. Separate groups of macaques were immunized with the peptide-cocktail by intravenous and subcutaneous routes using autologous dendritic cells (DC) and Freund's adjuvant, respectively. The vaccine elicited antigen specific IFN-γ-producing cells and T-cell proliferation, but not HIV-neutralizing antibodies. The vaccinated animals also exhibited efficient cross-clade cytolytic activity against target cells expressing envelope proteins corresponding to HIV-1 strains representative of multiple clades that increased after intravenous challenge with pathogenic SHIVKU2. Virus-neutralizing antibodies were either undetectable or present only transiently at low levels in the control as well as vaccinated monkeys after infection. Significant control of plasma viremia leading to undetectable levels was achieved in majority of vaccinated monkeys compared to mock-vaccinated controls. Monkeys vaccinated with the peptide-cocktail using autologous DC, compared to Freund's adjuvant, and the mock-vaccinated animals, showed significantly higher IFN-γ production, higher levels of vaccine-specific IFN-γ producing CD4+ cells and significant control of plasma viremia. These results support DC-based vaccine delivery and the utility of the conserved HIV-1 envelope peptide-cocktail, capable of priming strong cell-mediated immunity, for potential inclusion in HIV vaccination strategies

  12. Improving protein delivery of fibroblast growth factor-2 from bacterial inclusion bodies used as cell culture substrates

    OpenAIRE

    Seras Franzoso, Joaquin; Peebo, Karl; Garcia Fruitós, Elena; Vázquez Gómez, Esther; Rinas, Ursula; Villaverde Corrales, Antonio

    2014-01-01

    Altres ajuts: We are indebted CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN, Spain) for funding our research on inclusion bodies. Bacterial inclusion bodies (IBs) have recently been used to generate biocompatible cell culture interfaces, with diverse effects on cultured cells such as cell adhesion enhancement, stimulation of cell growth or induction of mesenchymal stem cell differentiation. Additionally, novel applications of IBs as sustained protein delivery systems with...

  13. Elliptic stable envelope

    CERN Document Server

    Aganagic, Mina

    2016-01-01

    We construct stable envelopes in equivariant elliptic cohomology of Nakajima quiver varieties. In particular, this gives an elliptic generalization of the results of arXiv:1211.1287. We apply them to the computation of the monodromy of $q$-difference equations arising the enumerative K-theory of rational curves in Nakajima varieties, including the quantum Knizhnik-Zamolodchikov equations.

  14. Structure and operation of bacterial tripartite pumps.

    Science.gov (United States)

    Hinchliffe, Philip; Symmons, Martyn F; Hughes, Colin; Koronakis, Vassilis

    2013-01-01

    In bacteria such as Pseudomonas aeruginosa and Escherichia coli, tripartite membrane machineries, or pumps, determine the efflux of small noxious molecules, such as detergents, heavy metals, and antibiotics, and the export of large proteins including toxins. They are therefore influential in bacterial survival, particularly during infections caused by multidrug-resistant pathogens. In these tripartite pumps an inner membrane transporter, typically an ATPase or proton antiporter, binds and translocates export or efflux substrates. In cooperation with a periplasmic adaptor protein it recruits and opens a TolC family cell exit duct, which is anchored in the outer membrane and projects across the periplasmic space between inner and outer membranes. Assembled tripartite pumps thus span the entire bacterial cell envelope. We review the atomic structures of each of the three pump components and discuss how these have allowed high-resolution views of tripartite pump assembly, operation, and possible inhibition. PMID:23808339

  15. Comparative study of HOCl-inflicted damage to bacterial DNA ex vivo and within cells.

    Science.gov (United States)

    Suquet, Christine; Warren, Jeffrey J; Seth, Nimulrith; Hurst, James K

    2010-01-15

    The prospects for using bacterial DNA as an intrinsic probe for HOCl and secondary oxidants/chlorinating agents associated with it has been evaluated using both in vitro and in vivo studies. Single-strand and double-strand breaks occurred in bare plasmid DNA that had been exposed to high levels of HOCl, although these reactions were very inefficient compared to polynucleotide chain cleavage caused by the OH.-generating reagent, peroxynitrite. Plasmid nicking was not increased when intact Escherichia coli were exposed to HOCl; rather, the amount of recoverable plasmid diminished in a dose-dependent manner. At concentration levels of HOCl exceeding lethal doses, genomic bacterial DNA underwent extensive fragmentation and the amount of precipitable DNA-protein complexes increased several-fold. The 5-chlorocytosine content of plasmid and genomic DNA isolated from HOCl-exposed E. coli was also slightly elevated above controls, as measured by mass spectrometry of the deaminated product, 5-chlorouracil. However, the yields were not dose-dependent over the bactericidal concentration range. Genomic DNA recovered from E. coli that had been subjected to phagocytosis by human neutrophils occasionally showed small increases in 5-chlorocytosine content when compared to analogous cellular reactions where myeloperoxidase activity was inhibited by azide ion. Overall, the amount of isolable 5-chlorouracil from the HOCl-exposed bacterial cells was far less than the damage manifested in polynucleotide bond cleavage and cross-linking. PMID:19850004

  16. Staying in Shape: the Impact of Cell Shape on Bacterial Survival in Diverse Environments.

    Science.gov (United States)

    Yang, Desirée C; Blair, Kris M; Salama, Nina R

    2016-03-01

    Bacteria display an abundance of cellular forms and can change shape during their life cycle. Many plausible models regarding the functional significance of cell morphology have emerged. A greater understanding of the genetic programs underpinning morphological variation in diverse bacterial groups, combined with assays of bacteria under conditions that mimic their varied natural environments, from flowing freshwater streams to diverse human body sites, provides new opportunities to probe the functional significance of cell shape. Here we explore shape diversity among bacteria, at the levels of cell geometry, size, and surface appendages (both placement and number), as it relates to survival in diverse environments. Cell shape in most bacteria is determined by the cell wall. A major challenge in this field has been deconvoluting the effects of differences in the chemical properties of the cell wall and the resulting cell shape perturbations on observed fitness changes. Still, such studies have begun to reveal the selective pressures that drive the diverse forms (or cell wall compositions) observed in mammalian pathogens and bacteria more generally, including efficient adherence to biotic and abiotic surfaces, survival under low-nutrient or stressful conditions, evasion of mammalian complement deposition, efficient dispersal through mucous barriers and tissues, and efficient nutrient acquisition. PMID:26864431

  17. Real-time Bacterial Detection by Single Cell Based Sensors UsingSynchrotron FTIR Spectromicroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Veiseh, Mandana; Veiseh, Omid; Martin, Michael C.; Bertozzi,Carolyn; Zhang, Miqin

    2005-08-10

    Microarrays of single macrophage cell based sensors weredeveloped and demonstrated for real time bacterium detection bysynchrotron FTIR microscopy. The cells were patterned on gold-SiO2substrates via a surface engineering technique by which the goldelectrodes were immobilized with fibronectin to mediate cell adhesion andthe silicon oxide background were passivated with PEG to resist proteinadsorption and cell adhesion. Cellular morphology and IR spectra ofsingle, double, and triple cells on gold electrodes exposed tolipopolysaccharide (LPS) of different concentrations were compared toreveal the detection capabilities of these biosensors. The single-cellbased sensors were found to generate the most significant IR wave numbervariation and thus provide the highest detection sensitivity. Changes inmorphology and IR spectrum for single cells exposed to LPS were found tobe time- and concentration-dependent and correlated with each other verywell. FTIR spectra from single cell arrays of gold electrodes withsurface area of 25 mu-m2, 100 mu-m2, and 400 mu-m2 were acquired usingboth synchrotron and conventional FTIR spectromicroscopes to study thesensitivity of detection. The results indicated that the developedsingle-cell platform can be used with conventional FTIRspectromicroscopy. This technique provides real-time, label-free, andrapid bacterial detection, and may allow for statistic and highthroughput analyses, and portability.

  18. The role of T cell subsets and cytokines in the regulation of intracellular bacterial infection

    Directory of Open Access Journals (Sweden)

    Oliveira S.C.

    1998-01-01

    Full Text Available Cellular immune responses are a critical part of the host's defense against intracellular bacterial infections. Immunity to Brucella abortus crucially depends on antigen-specific T cell-mediated activation of macrophages, which are the major effectors of cell-mediated killing of this organism. T lymphocytes that proliferate in response to B. abortus were characterized for phenotype and cytokine activity. Human, murine, and bovine T lymphocytes exhibited a type 1 cytokine profile, suggesting an analogous immune response in these different hosts. In vivo protection afforded by a particular cell type is dependent on the antigen presented and the mechanism of antigen presentation. Studies using MHC class I and class II knockout mice infected with B. abortus have demonstrated that protective immunity to brucellosis is especially dependent on CD8+ T cells. To target MHC class I presentation we transfected ex vivo a murine macrophage cell line with B. abortus genes and adoptively transferred them to BALB/c mice. These transgenic macrophage clones induced partial protection in mice against experimental brucellosis. Knowing the cells required for protection, vaccines can be designed to activate the protective T cell subset. Lastly, as a new strategy for priming a specific class I-restricted T cell response in vivo, we used genetic immunization by particle bombardment-mediated gene transfer

  19. Immunization of rabbits with highly purified, soluble, trimeric human immunodeficiency virus type 1 envelope glycoprotein induces a vigorous B cell response and broadly cross-reactive neutralization.

    Directory of Open Access Journals (Sweden)

    Gerald V Quinnan

    Full Text Available Previously we described induction of cross-reactive HIV-1 neutralizing antibody responses in rabbits using a soluble HIV-1 gp140 envelope glycoprotein (Env in an adjuvant containing monophosphoryl lipid A (MPL and QS21 (AS02A. Here, we compared different forms of the same HIV-1 strain R2 Env for antigenic and biophysical characteristics, and in rabbits characterized the extent of B cell induction for specific antibody expression and secretion and neutralizing responses. The forms of this Env that were produced in and purified from stably transformed 293T cells included a primarily dimeric gp140, a trimeric gp140 appended to a GCN4 trimerization domain (gp140-GCN4, gp140-GCN4 with a 15 amino acid flexible linker between the gp120 and gp41 ectodomain (gp140-GCN4-L, also trimeric, and a gp140 with the flexible linker purified from cell culture supernatants as either dimer (gp140-L(D or monomer (gp140-L(M. Multimeric states of the Env proteins were assessed by native gel electrophoresis and analytical ultracentrifugation. The different forms of gp140 bound broadly cross-reactive neutralizing (BCN human monoclonal antibodies (mAbs similarly in ELISA and immunoprecipitation assays. All Envs bound CD4i mAbs in the presence and absence of sCD4, as reported for the R2 Env. Weak neutralization of some strains of HIV-1 was seen after two additional doses in AS02A. Rabbits that were given a seventh dose of gp140-GCN4-L developed BCN responses that were weak to moderate, similar to our previous report. The specificity of these responses did not appear similar to that of any of the known BCN human mAbs. Induction of spleen B cell and plasma cells producing immunoglobulins that bound trimeric gp140-GCN4-L was vigorous, based on ELISpot and flow cytometry analyses. The results demonstrate that highly purified gp140-GCN4-L trimer in adjuvant elicits BCN responses in rabbits accompanied by vigorous B cell induction.

  20. Phylogenetic and metagenomic analyses of substrate-dependent bacterial temporal dynamics in microbial fuel cells.

    Directory of Open Access Journals (Sweden)

    Husen Zhang

    Full Text Available Understanding the microbial community structure and genetic potential of anode biofilms is key to improve extracellular electron transfers in microbial fuel cells. We investigated effect of substrate and temporal dynamics of anodic biofilm communities using phylogenetic and metagenomic approaches in parallel with electrochemical characterizations. The startup non-steady state anodic bacterial structures were compared for a simple substrate, acetate, and for a complex substrate, landfill leachate, using a single-chamber air-cathode microbial fuel cell. Principal coordinate analysis showed that distinct community structures were formed with each substrate type. The bacterial diversity measured as Shannon index decreased with time in acetate cycles, and was restored with the introduction of leachate. The change of diversity was accompanied by an opposite trend in the relative abundance of Geobacter-affiliated phylotypes, which were acclimated to over 40% of total Bacteria at the end of acetate-fed conditions then declined in the leachate cycles. The transition from acetate to leachate caused a decrease in output power density from 243±13 mW/m2 to 140±11 mW/m2, accompanied by a decrease in Coulombic electron recovery from 18±3% to 9±3%. The leachate cycles selected protein-degrading phylotypes within phylum Synergistetes. Metagenomic shotgun sequencing showed that leachate-fed communities had higher cell motility genes including bacterial chemotaxis and flagellar assembly, and increased gene abundance related to metal resistance, antibiotic resistance, and quorum sensing. These differentially represented genes suggested an altered anodic biofilm community in response to additional substrates and stress from the complex landfill leachate.

  1. Emulsification efficiency of adsorbed chitosan for bacterial cells accumulation at the oil-water interface.

    Science.gov (United States)

    Archakunakorn, Somwit; Charoenrat, Nattapat; Khamsakhon, Somruethai; Pongtharangkul, Thunyarat; Wongkongkatep, Pravit; Suphantharika, Manop; Wongkongkatep, Jirarut

    2015-04-01

    The use of bacterial cell or biocatalyst for industrial synthetic chemistry is on the way of significant growth since the biocatalyst requires low energy input compared to the chemical synthesis and can be considered as a green technology. However, majority of natural bacterial cell surface is hydrophilic which allows poor access to the hydrophobic substrate or product. In this study, Escherichia coli (E. coli) as a representative of hydrophilic bacterial cells were accumulated at the oil-water interface after association with chitosan at a concentration range of 0.75-750 mg/L. After association with negatively charged E coli having a ζ potential of -19.9 mV, a neutralization of positively charged chitosan occurred as evidenced by an increase in the ζ potential value of the mixtures with increasing chitosan concentration up to +3.5 mV at 750 mg/L chitosan. Both emulsification index and droplet size analysis revealed that chitosan-E. coli system is an excellent emulsion stabilizer to date because the threshold concentration was as low as 7.5 mg/L or 0.00075% w/v. A dramatic increase in the surface hydrophobicity of the E. coli as evidenced by an increase in contact angle from 19 to 88° with increasing chitosan concentration from 0 to 750 mg/L, respectively, resulted in an increase in the stability of oil-in-water emulsions stabilized by chitosan-E. coli system. The emulsion was highly stable even the emulsification was performed under 20% salt condition, or temperature ranged between 20 and 50 °C. Emulsification was failed when the oil volume fraction was higher than 0.5, indicating that no phase inversion occurred. The basic investigation presented in this study is a crucial platform for its application in biocatalyst industry and bioremediation of oil spill. PMID:25341365

  2. Identification of Bacterial Cell Wall Lyases via Pseudo Amino Acid Composition.

    Science.gov (United States)

    Chen, Xin-Xin; Tang, Hua; Li, Wen-Chao; Wu, Hao; Chen, Wei; Ding, Hui; Lin, Hao

    2016-01-01

    Owing to the abuse of antibiotics, drug resistance of pathogenic bacteria becomes more and more serious. Therefore, it is interesting to develop a more reasonable way to solve this issue. Because they can destroy the bacterial cell structure and then kill the infectious bacterium, the bacterial cell wall lyases are suitable candidates of antibacteria sources. Thus, it is urgent to develop an accurate and efficient computational method to predict the lyases. Based on the consideration, in this paper, a set of objective and rigorous data was collected by searching through the Universal Protein Resource (the UniProt database), whereafter a feature selection technique based on the analysis of variance (ANOVA) was used to acquire optimal feature subset. Finally, the support vector machine (SVM) was used to perform prediction. The jackknife cross-validated results showed that the optimal average accuracy of 84.82% was achieved with the sensitivity of 76.47% and the specificity of 93.16%. For the convenience of other scholars, we built a free online server called Lypred. We believe that Lypred will become a practical tool for the research of cell wall lyases and development of antimicrobial agents. PMID:27437396

  3. Applications of whole-cell bacterial sensors in biotechnology and environmental science

    Energy Technology Data Exchange (ETDEWEB)

    Yagi, Kiyohito [Osaka Univ., Suita (Japan). Graduate School of Pharmaceutical Sciences

    2007-01-15

    Biosensors have major advantages over chemical or physical analyses with regard to specificity, sensitivity, and portability. Recently, many types of whole-cell bacterial biosensors have been developed using recombinant DNA technology. The bacteria are genetically engineered to respond to the presence of chemicals or physiological stresses by synthesizing a reporter protein, such as luciferase, {beta}-galactosidase, or green fluorescent protein. In addition to an overview of conventional biosensors, this minireview discusses a novel type of biosensor using a photosynthetic bacterium as the sensor strain and the crtA gene, which is responsible for carotenoid synthesis, as the reporter. Since bacteria possess a wide variety of stress-response mechanisms, including antioxidation, heat-shock responses, nutrient-starvation, and membrane-damage responses, DNA response elements for several stress-response proteins can be fused with various reporter genes to construct a versatile set of bacterial biosensors for a variety of analytes. Portable biosensors for on-site monitoring have been developed using a freeze-dried biosensing strain, and cell array biosensors have been designed for high-throughput analysis. Moreover, in the future, the use of single-cell biosensors will permit detailed analyses of samples. Signals from such sensors could be detected with digital imaging, epifluorescence microscopy, and/or flow cytometry. (orig.)

  4. Identification of Bacterial Cell Wall Lyases via Pseudo Amino Acid Composition

    Science.gov (United States)

    Tang, Hua; Li, Wen-Chao; Wu, Hao; Ding, Hui

    2016-01-01

    Owing to the abuse of antibiotics, drug resistance of pathogenic bacteria becomes more and more serious. Therefore, it is interesting to develop a more reasonable way to solve this issue. Because they can destroy the bacterial cell structure and then kill the infectious bacterium, the bacterial cell wall lyases are suitable candidates of antibacteria sources. Thus, it is urgent to develop an accurate and efficient computational method to predict the lyases. Based on the consideration, in this paper, a set of objective and rigorous data was collected by searching through the Universal Protein Resource (the UniProt database), whereafter a feature selection technique based on the analysis of variance (ANOVA) was used to acquire optimal feature subset. Finally, the support vector machine (SVM) was used to perform prediction. The jackknife cross-validated results showed that the optimal average accuracy of 84.82% was achieved with the sensitivity of 76.47% and the specificity of 93.16%. For the convenience of other scholars, we built a free online server called Lypred. We believe that Lypred will become a practical tool for the research of cell wall lyases and development of antimicrobial agents. PMID:27437396

  5. Selective removal of DNA from dead cells of mixed bacterial communities by use of ethidium monoazide.

    Science.gov (United States)

    Nocker, Andreas; Camper, Anne K

    2006-03-01

    The distinction between viable and dead bacterial cells poses a major challenge in microbial diagnostics. Due to the persistence of DNA in the environment after cells have lost viability, DNA-based quantification methods overestimate the number of viable cells in mixed populations or even lead to false-positive results in the absence of viable cells. On the other hand, RNA-based diagnostic methods, which circumvent this problem, are technically demanding and suffer from some drawbacks. A promising and easy-to-use alternative utilizing the DNA-intercalating dye ethidium monoazide bromide (EMA) was published recently. This chemical is known to penetrate only into "dead" cells with compromised cell membrane integrity. Subsequent photoinduced cross-linking was reported to inhibit PCR amplification of DNA from dead cells. We provide evidence here that in addition to inhibition of amplification, most of the DNA from dead cells is actually lost during the DNA extraction procedure, probably together with cell debris which goes into the pellet fraction. Exposure of bacteria to increasing stress and higher proportions of dead cells in defined populations led to increasing loss of genomic DNA. Experiments were performed using Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium as model pathogens and using real-time PCR for their quantification. Results showed that EMA treatment of mixed populations of these two species provides a valuable tool for selective removal of DNA of nonviable cells by using conventional extraction protocols. Furthermore, we provide evidence that prior to denaturing gradient gel electrophoresis, EMA treatment of a mature mixed-population drinking-water biofilm containing a substantial proportion of dead cells can result in community fingerprints dramatically different from those for an untreated biofilm. The interpretation of such fingerprints can have important implications in the field of microbial ecology. PMID:16517648

  6. Simultaneous selection of soil electroactive bacterial communities associated to anode and cathode in a two-chamber Microbial Fuel Cell

    Science.gov (United States)

    Chiellini, Carolina; Bacci, Giovanni; Fani, Renato; Mocali, Stefano

    2016-04-01

    Different bacteria have evolved strategies to transfer electrons over their cell surface to (or from) their extracellular environment. This electron transfer enables the use of these bacteria in bioelectrochemical systems (BES) such as Microbial Fuel Cells (MFCs). In MFC research the biological reactions at the cathode have long been a secondary point of interest. However, bacterial biocathodes in MFCs represent a potential advantage compared to traditional cathodes, for both their low costs and their low impact on the environment. The main challenge in biocathode set-up is represented by the selection of a bacterial community able to efficiently accept electrons from the electrode, starting from an environmental matrix. In this work, a constant voltage was supplied on a two-chamber MFC filled up with soil over three weeks in order to simultaneously select an electron donor bacterial biomass on the anode and an electron acceptor biomass on the cathode, starting from the same soil. Next Generation Sequencing (NGS) analysis was performed to characterize the bacterial community of the initial soil, in the anode, in the cathode and in the control chamber not supplied with any voltage. Results highlighted that both the MFC conditions and the voltage supply affected the soil bacterial communities, providing a selection of different bacterial groups preferentially associated to the anode (Betaproteobacteria, Bacilli and Clostridia) and to the cathode (Actinobacteria and Alphaproteobacteria). These results confirmed that several electroactive bacteria are naturally present within a top soil and, moreover, different soil bacterial genera could provide different electrical properties.

  7. Regulatory T cell suppressive potency dictates the balance between bacterial proliferation and clearance during persistent Salmonella infection.

    Directory of Open Access Journals (Sweden)

    Tanner M Johanns

    Full Text Available The pathogenesis of persistent infection is dictated by the balance between opposing immune activation and suppression signals. Herein, virulent Salmonella was used to explore the role and potential importance of Foxp3-expressing regulatory T cells in dictating the natural progression of persistent bacterial infection. Two distinct phases of persistent Salmonella infection are identified. In the first 3-4 weeks after infection, progressively increasing bacterial burden was associated with delayed effector T cell activation. Reciprocally, at later time points after infection, reductions in bacterial burden were associated with robust effector T cell activation. Using Foxp3(GFP reporter mice for ex vivo isolation of regulatory T cells, we demonstrate that the dichotomy in infection tempo between early and late time points is directly paralleled by drastic changes in Foxp3(+ Treg suppressive potency. In complementary experiments using Foxp3(DTR mice, the significance of these shifts in Treg suppressive potency on infection outcome was verified by enumerating the relative impacts of regulatory T cell ablation on bacterial burden and effector T cell activation at early and late time points during persistent Salmonella infection. Moreover, Treg expression of CTLA-4 directly paralleled changes in suppressive potency, and the relative effects of Treg ablation could be largely recapitulated by CTLA-4 in vivo blockade. Together, these results demonstrate that dynamic regulation of Treg suppressive potency dictates the course of persistent bacterial infection.

  8. Sequential evolution of bacterial morphology by co-option of a developmental regulator

    OpenAIRE

    Jiang, Chao; Brown, Pamela J.B.; Ducret, Adrien; Brun, Yves V.

    2014-01-01

    What mechanisms underlie the transitions responsible for the diverse shapes observed in the living world? While bacteria display a myriad of morphologies 1 , the mechanisms responsible for the evolution of bacterial cell shape are not understood. We investigated morphological diversity in a group of bacteria that synthesize an appendage-like extension of the cell envelope called the stalk 2,3 . The location and number of stalks varies among species, as exemplified by three distinct sub-cellul...

  9. Definition of a Bacterial Type IV Secretion Pathway for a DNA Substrate

    OpenAIRE

    Cascales, Eric; Christie, Peter J.

    2004-01-01

    Bacteria use conjugation systems, a subfamily of the type IV secretion systems, to transfer DNA to recipient cells. Despite 50 years of research, the architecture and mechanism of action of the channel mediating DNA transfer across the bacterial cell envelope remains obscure. By use of a sensitive, quantifiable assay termed transfer DNA immunoprecipitation (TrIP), we identify contacts between a DNA substrate (T-DNA) and 6 of 12 components of the VirB/D4 conjugation system of the phytopathogen...

  10. (Quasi-)Poisson enveloping algebras

    OpenAIRE

    Yang, Yan-Hong; Yuan YAO; Ye, Yu

    2010-01-01

    We introduce the quasi-Poisson enveloping algebra and Poisson enveloping algebra for a non-commutative Poisson algebra. We prove that for a non-commutative Poisson algebra, the category of quasi-Poisson modules is equivalent to the category of left modules over its quasi-Poisson enveloping algebra, and the category of Poisson modules is equivalent to the category of left modules over its Poisson enveloping algebra.

  11. Identification of human linear B-cell epitope sites on the envelope glycoproteins of Crimean-Congo haemorrhagic fever virus.

    Science.gov (United States)

    Goedhals, D; Paweska, J T; Burt, F J

    2015-05-01

    A peptide library was used to screen for regions containing potential linear B-cell epitope sites in the glycoproteins and nucleoprotein of Crimean-Congo haemorrhagic fever virus (CCHFV) in an enzyme-linked immunosorbent assay (ELISA). The library consisted of 156 peptides, spanning the nucleoprotein and mature GN and GC proteins in a 19-mer with 9-mer overlap format. Using pooled serum samples from convalescent patients to screen the library, six peptides were identified as potential epitope sites. Further testing of these six peptides with individual patient sera identified two of these peptides as probable epitope sites, with peptide G1451-1469 reacting to 13/15 and peptide G1613-1631 to 14/15 human sera. These peptides are situated on the GC protein at amino acid positions 1451-1469 (relative to CCHFV isolate SPU103/97) (TCTGCYACSSGISCKVRIH) and 1613-1631 (FMFGWRILFCFKCCRRTRG). Identified peptides may have application in ELISA for diagnostic or serosurveillance purposes. PMID:25185583

  12. Probing interaction of Gram-positive and Gram-negative bacterial cells with ZnO nanorods

    Energy Technology Data Exchange (ETDEWEB)

    Jain, Aanchal; Bhargava, Richa; Poddar, Pankaj, E-mail: p.poddar@ncl.res.in

    2013-04-01

    In the present work, the physiological effects of the ZnO nanorods on the Gram positive (Staphylococcus aureus and Bacillus subtilis) and Gram-negative (Escherichia coli and Aerobacter aerogenes) bacterial cells have been studied. The analysis of bacterial growth curves for various concentrations of ZnO nanorods indicates that Gram positive and Gram negative bacterial cells show inhibition at concentrations of ∼ 64 and ∼ 256 μg/mL respectively. The marked difference in susceptibility towards nanorods was also validated by spread plate and disk diffusion methods. In addition, the scanning electron micrographs show a clear damage to the cells via changed morphology of the cells from rod to coccoid etc. The confocal optical microscopy images of these cells also demonstrate the reduction in live cell count in the presence of ZnO nanorods. These, results clearly indicate that the antibacterial activity of ZnO nanorods is higher towards Gram positive bacterium than Gram negative bacterium which indicates that the structure of the cell wall might play a major role in the interaction with nanostructured materials and shows high sensitivity to the particle concentration. Highlights: ► Effect of ZnO nanorods on the growth cycles of four bacterial strains. ► A relation has been established between growth rate of bacteria and concentration. ► Serious damage in the morphology of bacterial cells in the presence of ZnO nanorods. ► Microscopic studies to see the time dependent effect on bacterial cells.

  13. Dynamics of Different Bacterial Communities Are Capable of Generating Sustainable Electricity from Microbial Fuel Cells with Organic Waste

    OpenAIRE

    Yamamoto, Shuji; Suzuki, Kei; Araki, Yoko; Mochihara, Hiroki; Hosokawa, Tetsuya; KUBOTA, Hiroko; Chiba, Yusuke; Rubaba, Owen; Tashiro, Yosuke; Futamata, Hiroyuki

    2014-01-01

    The relationship between the bacterial communities in anolyte and anode biofilms and the electrochemical properties of microbial fuel cells (MFCs) was investigated when a complex organic waste-decomposing solution was continuously supplied to MFCs as an electron donor. The current density increased gradually and was maintained at approximately 100 to 150 mA m−2. Polarization curve analyses revealed that the maximum power density was 7.4 W m−3 with an internal resistance of 110 Ω. Bacterial co...

  14. Yeast Cell Wall Extract Induces Disease Resistance against Bacterial and Fungal Pathogens in Arabidopsis thaliana and Brassica Crop

    OpenAIRE

    Narusaka, Mari; Minami, Taichi; Iwabuchi, Chikako; Hamasaki, Takashi; Takasaki, Satoko; Kawamura, Kimito; Narusaka, Yoshihiro

    2015-01-01

    Housaku Monogatari (HM) is a plant activator prepared from a yeast cell wall extract. We examined the efficacy of HM application and observed that HM treatment increased the resistance of Arabidopsis thaliana and Brassica rapa leaves to bacterial and fungal infections. HM reduced the severity of bacterial leaf spot and anthracnose on A. thaliana and Brassica crop leaves with protective effects. In addition, gene expression analysis of A. thaliana plants after treatment with HM indicated incre...

  15. Structural and functional studies of MinD ATPase: implications for the molecular recognition of the bacterial cell division apparatus

    OpenAIRE

    Hayashi, Ikuko; Oyama, Takuji; Morikawa, Kosuke

    2001-01-01

    Proper placement of the bacterial cell division site requires the site-specific inactivation of other potential division sites. In Escherichia coli, selection of the correct mid-cell site is mediated by the MinC, MinD and MinE proteins. To clarify the functional role of the bacterial cell division inhibitor MinD, which is a membrane-associated ATPase that works as an activator of MinC, we determined the crystal structure of a Pyrococcus furiosus MinD homologue complexed with a substrate analo...

  16. Involvement of the cell-specific pigment genes pks and sult in bacterial defense response of sea urchins Strongylocentrotus intermedius.

    Science.gov (United States)

    Kiselev, Konstantin V; Ageenko, Natalya V; Kurilenko, Valeria V

    2013-03-26

    Bacterial infections are one of the most important problems in mass aquaculture, causing the loss of millions of juvenile organisms. We isolated 22 bacterial strains from the cavity fluid of the sea urchin Strongylocentrotus pallidus and used phylogenetic analysis based on 16S rRNA gene sequences to separate the bacterial strains into 9 genera (Aliivibrio, Bizionia, Colwellia, Olleya, Paenibacillus, Photobacterium, Pseudoalteromonas, Shewanella, and Vibrio). Incubating Strongylocentrotus intermedius larvae with a strain from each of the 9 bacterial genera, we investigated the viability of the larvae, the amount of pigment cells, and the level of polyketide synthase (pks) and sulfotransferase (sult) gene expression. Results of the assay on sea urchin development showed that all bacterial strains, except Pseudoalteromonas and Bizionia, suppressed sea urchin development (resulting in retardation of the embryos' development with cellular disorders) and reduced cell viability. We found that pks expression in the sea urchin larvae after incubation with the bacteria of 9 tested genera was significantly increased, while the sult expression was increased only after the treatment with Pseudoalteromonas and Shewanella. Shikimic acid, which is known to activate the biosynthesis of naphthoquinone pigments, increased the tolerance of the sea urchin embryos to the bacteria. In conclusion, we show that the cell-specific pigment genes pks and sult are involved in the bacterial defense response of sea urchins. PMID:23548362

  17. Enteric Bacterial Invasion Of Intestinal Epithelial Cells In Vitro Is Dramatically Enhanced Using a Vertical Diffusion Chamber Model

    OpenAIRE

    Naz, Neveda; Mills, Dominic C.; Wren, Brendan W.; Dorrell, Nick

    2013-01-01

    The interactions of bacterial pathogens with host cells have been investigated extensively using in vitro cell culture methods. However as such cell culture assays are performed under aerobic conditions, these in vitro models may not accurately represent the in vivo environment in which the host-pathogen interactions take place. We have developed an in vitro model of infection that permits the coculture of bacteria and host cells under different medium and gas conditions. The Vertical Diffusi...

  18. ATP-mediated Erk1/2 activation stimulates bacterial capture by filopodia, which precedes Shigella invasion of epithelial cells.

    OpenAIRE

    Romero S; Grompone G; Carayol N; Mounier J; Guadagnini S; Prevost MC; Sansonetti PJ; Nhieu GT

    2011-01-01

    Shigella, the causative agent of bacillary dysentery in humans, invades epithelial cells, using a type III secretory system (T3SS) to inject bacterial effectors into host cells and remodel the actin cytoskeleton. ATP released through connexin hemichanels on the epithelial membrane stimulates Shigella invasion and dissemination in epithelial cells. Here, we show that prior to contact with the cell body, Shigella is captured by nanometer-thin micropodial extensions (NMEs) at a distance from the...

  19. Influence of dietary nucleotide restriction on bacterial sepsis and phagocytic cell function in mice.

    Science.gov (United States)

    Kulkarni, A D; Fanslow, W C; Drath, D B; Rudolph, F B; Van Buren, C T

    1986-02-01

    Although enzyme defects in purine metabolism have revealed the importance of these substrates to maintenance of a normal immune response, the role of exogenous nucleotides on the cells that mediate the host defense system has remained largely unexplored. Recent investigations have revealed that dietary nucleotides are vital to the maintenance of cell-mediated responses to antigen stimulation. To test the influence of dietary nucleotide deprivation on resistance to infection, Balb/c mice were maintained on chow, a nucleotide-free (NF) diet, or an NF diet repleted with adenine, uracil, or RNA. Mice on the NF diet suffered 100% mortality following intravenous challenge with Staphylococcus aureus, while chow-fed and RNA- or uracil-repleted mice demonstrated significantly greater resistance to this bacterial challenge. Macrophages from mice on the NF diet had decreased phagocytic activity as measured by uptake of radiolabeled bacteria compared with mice maintained on the NF diet supplemented with adenine, uracil, or RNA. No change in S aureus antibody response was noted on the various diets. Although the mechanism of this suppression of nonspecific immunity remains unclear, provision of nucleotides to defined diets appears vital to maintain host resistance to bacterial challenge. PMID:3947217

  20. Testing an agent-based model of bacterial cell motility: How nutrient concentration affects speed distribution

    Science.gov (United States)

    Garcia, V.; Birbaumer, M.; Schweitzer, F.

    2011-08-01

    We revisit a recently proposed agent-based model of active biological motion and compare its predictions with own experimental findings for the speed distribution of bacterial cells, Salmonella typhimurium. Agents move according to a stochastic dynamics and use energy stored in an internal depot for metabolism and active motion. We discuss different assumptions of how the conversion from internal to kinetic energy d( v) may depend on the actual speed, to conclude that d 2 v ξ with either ξ = 2 or 1 speed distribution of bacteria which were obtained in media of different nutrient concentration and at different times. We find that both hypotheses are in line with the experimental observations, with ξ between 1.67 and 2.0. Regarding the influence of a higher nutrient concentration, we conclude that the take-up of energy by bacterial cells is indeed increased. But this energy is not used to increase the speed, with 40 μm/s as the most probable value of the speed distribution, but is rather spend on metabolism and growth.

  1. Spatiotemporal development of the bacterial community in a tubular longitudinal microbial fuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jung Rae; Premier, Giuliano C. [Glamorgan Univ., Pontypridd (United Kingdom). Faculty of Advnaced Technology; Beecroft, Nelli J.; Avignone-Rossa, Claudio [Surrey Univ., Guildford (United Kingdom). Microbial Sciences; Varcoe, John R.; Slade, Robert C.T. [Surrey Univ., Guildford (United Kingdom). Chemical Sciences; Dinsdale, Richard M.; Guwy, Alan J. [Glamorgan Univ., Pontypridd (United Kingdom). Faculty of Health, Sport and Science; Thumser, Alfred [Surrey Univ., Guildford (United Kingdom). Biochemical Sciences

    2011-05-15

    The spatiotemporal development of a bacterial community in an exoelectrogenic biofilm was investigated in sucrose-fed longitudinal tubular microbial fuel cell reactors, consisting of two serially connected modules. The proportional changes in the microbial community composition were assessed by polymerase chain reaction-denaturing gradient gel electrophoresis (DGGE) and DNA sequencing in order to relate them to the performance and stability of the bioelectrochemical system. The reproducibility of duplicated reactors, evaluated by cluster analysis and Jaccard's coefficient, shows 80-90% similarity in species composition. Biofilm development through fed-batch start-up and subsequent stable continuous operation results in a population shift from {gamma}-Proteobacteria- and Bacteroidetes- to Firmicutes-dominated communities, with other diverse species present at much lower relative proportions. DGGE patterns were analysed by range-weighted richness (Rr) and Pareto-Lorenz evenness distribution curves to investigate the evolution of the bacterial community. The first modules shifted from dominance by species closely related to Bacteroides graminisolvens, Raoultella ornithinolytica and Klebsiella sp. BM21 at the start of continuous-mode operation to a community dominated by Paludibacter propionicigenes-, Lactococcus sp.-, Pantoea agglomerans- and Klebsiella oxytoca-related species with stable power generation (6.0 W/m{sup 3}) at day 97. Operational strategies that consider the dynamics of the population will provide useful parameters for evaluating system performance in the practical application of microbial fuel cells. (orig.)

  2. Thermal Activated Envelope

    DEFF Research Database (Denmark)

    Foged, Isak Worre; Pasold, Anke

    2015-01-01

    search procedure, the combination of materials and their bonding temperature is found in relation to the envelope effect on a thermal environment inside a defined space. This allows the designer to articulate dynamic composites with time-based thermal functionality, related to the material dynamics......, environmental dynamics and occupancy dynamics. Lastly, a physical prototype is created, which illustrates the physical expression of the bi-materials and the problems related to manufacturing of these composite structures.......The research studies the making of a responsive architectural envelope based on bi-materials. The bi-materials are organized according to a method that combines different isotropic metals and plastic into an active composite structure that reacts to temperature variations. Through an evolutionary...

  3. Thermal Responsive Envelope

    DEFF Research Database (Denmark)

    Foged, Isak Worre; Pasold, Anke

    2015-01-01

    includes the calculation of bending behaviour, the calculation of perceived temperatures inside the envelope and the evolutionary module, which in a design process advance the composite structure in relation to the thermal environment desired. The research presents the methods used and developed, the way...... composite layers and their relative layer lengths thereby embedding the merged material effect to create a responsive behavioural architectural envelope. Copper and polypropylene are used as base materials for the composite structure due to their high differences in thermal expansion, surface emissivity...... alterations, their respective durability and copper’s architectural (visual and transformative) aesthetic qualities. Through the use of an evolutionary solver, the composite structure of the elements are organised to find the bending behaviour specified by and for the thermal environments. The entire model...

  4. On isogeometric yield envelopes.

    OpenAIRE

    Coombs, W.M.

    2015-01-01

    In numerical analysis the failure of engineering materials is controlled through specifying yield envelopes (or surfaces) that bound the allowable stress in the material. Simple examples include the prismatic von Mises (circle) and Tresca (hexagon) yield surfaces. However, each surface is distinct and requires a specific equation describing the shape of the surface to be formulated in each case. These equations impact on the numerical implementation (specifically relating to st...

  5. Data envelopment analysis

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    This review introduces the history and present status of data envelopment analysis (DEA) research, particularly the evaluation process. And extensions of some DEA models are also described. It is pointed out that mathematics, economics and management science are the main forces in the DEA development, optimization provides the fundamental method for the DEA research, and the wide range of applications enforces the rapid development of DEA.

  6. INTERNAL MAIL ENVELOPES

    CERN Multimedia

    Mail Office

    2001-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration.

  7. INTERNAL MAIL ENVELOPES

    CERN Multimedia

    Mail Office

    2002-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration. Mail Office

  8. INTERNAL MAIL ENVELOPES

    CERN Multimedia

    Mail Office

    2002-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration.   Mail Office

  9. INTERNAL MAIL ENVELOPES

    CERN Multimedia

    Mail Office

    2002-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration.

  10. URGENT - Internal Mail Envelopes

    CERN Multimedia

    2007-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration. Mail Office

  11. INTERNAL CIRCULATION ENVELOPES

    CERN Multimedia

    Mail Office

    2001-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or a piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration.

  12. URGENT - Internal Mail Envelopes

    CERN Multimedia

    Mail Office

    2004-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unused stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration. Mail Office

  13. Internal mail envelopes

    CERN Multimedia

    2003-01-01

    Internal mail envelopes often finish up in large piles in certain offices, thus creating a shortage for other users of the mail service, who would be grateful if everyone with an unusual stock could deposit them in their mail box, after attaching them together with an elastic band or piece of string. The messengers will then collect them so that the Mail Office can put them back in circulation. Thank you for your understanding and collaboration. Mail Office

  14. Analysis of a Stochastic Model for Bacterial Growth and the Lognormality of the Cell-Size Distribution

    Science.gov (United States)

    Yamamoto, Ken; Wakita, Jun-ichi

    2016-07-01

    This paper theoretically analyzes a phenomenological stochastic model for bacterial growth. This model comprises cell division and the linear growth of cells, where growth rates and cell cycles are drawn from lognormal distributions. We find that the cell size is expressed as a sum of independent lognormal variables. We show numerically that the quality of the lognormal approximation greatly depends on the distributions of the growth rate and cell cycle. Furthermore, we show that actual parameters of the growth rate and cell cycle take values that give a good lognormal approximation; thus, the experimental cell-size distribution is in good agreement with a lognormal distribution.

  15. Analysis of a stochastic model for bacterial growth and the lognormality in the cell-size distribution

    CERN Document Server

    Yamamoto, Ken

    2016-01-01

    This paper theoretically analyzes a phenomenological stochastic model for bacterial growth. This model comprises cell divisions and linear growth of cells, where growth rates and cell cycles are drawn from lognormal distributions. We derive that the cell size is expressed as a sum of independent lognormal variables. We show numerically that the quality of the lognormal approximation greatly depends on the distributions of the growth rate and cell cycle. Furthermore, we show that actual parameters of the growth rate and cell cycle take values which give good lognormal approximation, so the experimental cell-size distribution is in good agreement with a lognormal distribution.

  16. Dynamics of different bacterial communities are capable of generating sustainable electricity from microbial fuel cells with organic waste.

    Science.gov (United States)

    Yamamoto, Shuji; Suzuki, Kei; Araki, Yoko; Mochihara, Hiroki; Hosokawa, Tetsuya; Kubota, Hiroko; Chiba, Yusuke; Rubaba, Owen; Tashiro, Yosuke; Futamata, Hiroyuki

    2014-01-01

    The relationship between the bacterial communities in anolyte and anode biofilms and the electrochemical properties of microbial fuel cells (MFCs) was investigated when a complex organic waste-decomposing solution was continuously supplied to MFCs as an electron donor. The current density increased gradually and was maintained at approximately 100 to 150 mA m(-2). Polarization curve analyses revealed that the maximum power density was 7.4 W m(-3) with an internal resistance of 110 Ω. Bacterial community structures in the organic waste-decomposing solution and MFCs differed from each other. Clonal analyses targeting 16S rRNA genes indicated that bacterial communities in the biofilms on MFCs developed to specific communities dominated by novel Geobacter. Multidimensional scaling analyses based on DGGE profiles revealed that bacterial communities in the organic waste-decomposing solution fluctuated and had no dynamic equilibrium. Bacterial communities on the anolyte in MFCs had a dynamic equilibrium with fluctuations, while those of the biofilm converged to the Geobacter-dominated structure. These bacterial community dynamics of MFCs differed from those of control-MFCs under open circuit conditions. These results suggested that bacterial communities in the anolyte and biofilm have a gentle symbiotic system through electron flow, which resulted in the advance of current density from complex organic waste. PMID:24789988

  17. Concentrating bacterial cells using a ratchet system: a lattice Monte Carlo simulation study

    Science.gov (United States)

    Tao, Yuguo; Slater, Gary

    2012-02-01

    Rectification of motile E. coli bacteria has been observed in the presence of funnel-like channels. We present a lattice Monte Carlo model which takes into account both the size and the mechanical and thermodynamic properties of autonomous bacterial cells. The motion of the cells is composed of alternating run and tumble periods. We show that the rectification effect of the funnels is strongly dependent upon the effective random walk step length of the run/tumble cycle as well as the size of the funnel's aperture. Our results agree with experimental observations, and also confirm some conclusions from a previous simulation model of point-like bacteria. We also explore series of funnels as a means to pump and concentrate cells. We observe deviations from theoretical predictions when the size of the cells is comparable to that of the aperture of the funnel. The current model can be extended to study cells with different shapes, e.g. cigar-shape bacteria.

  18. Cytotoxicity and genotoxicity of bacterial magnetosomes against human retinal pigment epithelium cells

    Science.gov (United States)

    Qi, Lei; Lv, Xiujuan; Zhang, Tongwei; Jia, Peina; Yan, Ruiying; Li, Shuli; Zou, Ruitao; Xue, Yuhua; Dai, Liming

    2016-06-01

    A variety of nanomaterials have been developed for ocular diseases. The ability of these nanomaterials to pass through the blood-ocular barrier and their biocompatibility are essential characteristics that must be considered. Bacterial magnetosomes (BMs) are a type of biogenic magnetic nanomaterials synthesized by magnetotactic bacteria. Due to their unique biomolecular membrane shell and narrow size distribution of approximately 30 nm, BMs can pass through the blood-brain barrier. The similarity of the blood-ocular barrier to the blood-brain barrier suggests that BMs have great potential as treatments for ocular diseases. In this work, BMs were isolated from magnetotactic bacteria and evaluated in various cytotoxicity and genotoxicity studies in human retinal pigment epithelium (ARPE-19) cells. The BMs entered ARPE-19 cells by endocytosis after a 6-h incubation and displayed much lower cytotoxicity than chemically synthesized magnetic nanoparticles (MNPs). MNPs exhibited significantly higher genotoxicity than BMs and promoted the expression of Bax (the programmed cell death acceleration protein) and the induction of greater cell necrosis. In BM-treated cells, apoptosis tended to be suppressed via increased expression of the Bcl-2 protein. In conclusion, BMs display excellent biocompatibility and potential for use in the treatment of ocular diseases.

  19. Colour removal from aqueous solutions of metal-complex azo dyes using bacterial cells of Shewanella strain J18 143.

    Science.gov (United States)

    Li, Tie; Guthrie, James Thomas

    2010-06-01

    The decoloration treatment of textile dye effluents through biodegradation, using bacterial cells, has been studied as a possible means of solving some of the problems that are associated with the pollution of water sources by colorants. In this paper, the use of whole bacterial cells of Shewanella J18 143 for the reduction of aqueous solutions of selected mono-azo, metal-complex dyes, namely Irgalan Grey GLN, Irgalan Black RBLN and Irgalan Blue 3GL, was investigated. The effects of temperature, pH and dye concentration on colour removal were also investigated and shown to be important. The operative conditions for the removal of colour were 30 degrees C, at pH 6.8, with a final dye concentration of 0.12 g/L in the colour reduction system. This study provides an extension to the application of Shewanella strain J18 143 bacterial cells in the decoloration of textile wastewaters. PMID:20167478

  20. Micro Corona Ionizer as an Ozone Source for Bacterial Cell Lysis

    Science.gov (United States)

    Lee, Eun-Hee; Lim, Hyun Jeong; Chua, Beelee; Son, Ahjeong

    2015-04-01

    DNA extraction is a critical process of DNA assays including polymerase chain reaction (PCR), microarrays, molecular cloning, and DNA hybridization which has been well established and can be implemented by commercial kits. DNA extraction involves cell lysis, precipitation, and purification through the combination of physical and chemical processes. Cell lysis is essential to high DNA recovery yield which can be achieved via a variety of physical, chemical, and enzymatic methods. However, these methods were originally developed for bioassays that were labor intensive, time consuming, and vulnerable to contamination and inhibition. Here, we proposed to employ a micro corona ionizer as an ozone source to lyse bacterial cells. Ozone has been well known and used as a disinfectant which allows cell lysis and DNA extraction. Previously, we have shown that a micro corona ionizer is capable of generating a significant amount of ozone. In this study, we employed the micro corona ionizer for the bacterial cell lysis which consists of a 50 μm diameter cantilever wire as the discharge cathode and a 50 μm thick copper foil as anode. Applied voltages varied from 1900 to 2200 V with corresponding corona currents from 16 to 28 μA. The resultant ozone (concentration > 0.14 ppm) generated from the micro corona ionizer was bubbled into the sample via a miniature pump. We demonstrated the cell lysis of Pseudomonas putida as the target bacterium using the micro corona ionizer. At a flow rate of 38 ml/min and applied corona voltage of 2000 V, 98.5 ± 0.2% lysis (normalized to sonication result) was achieved after 10 min. In comparison, untreated and air-treated samples showed normalized % lysis of 11.9 ± 2.4 and 36.1 ± 1.7%, respectively. We also showed that the cell lysis efficiency could be significantly increased by increasing the flow rate and the applied corona voltage. By comparing the experimental results for continuous and pulsed treatment, we verified that the percentage of

  1. Effects of zinc oxide nanoparticles on Kupffer cell phagosomal motility, bacterial clearance, and liver function

    Directory of Open Access Journals (Sweden)

    Watson CY

    2015-06-01

    Full Text Available Christa Y Watson, Ramon M Molina, Andressa Louzada, Kimberly M Murdaugh, Thomas C Donaghey, Joseph D BrainCenter for Nanotechnology and Nanotoxicology, Molecular and Integrative Physiological Sciences Program, Department of Environmental Health, Harvard T.H. Chan School of Public Health, Boston, MA, USABackground: Zinc oxide engineered nanoparticles (ZnO ENPs have potential as nanomedicines due to their inherent properties. Studies have described their pulmonary impact, but less is known about the consequences of ZnO ENP interactions with the liver. This study was designed to describe the effects of ZnO ENPs on the liver and Kupffer cells after intravenous (IV administration.Materials and methods: First, pharmacokinetic studies were conducted to determine the tissue distribution of neutron-activated 65ZnO ENPs post-IV injection in Wistar Han rats. Then, a noninvasive in vivo method to assess Kupffer cell phagosomal motility was employed using ferromagnetic iron particles and magnetometry. We also examined whether prior IV injection of ZnO ENPs altered Kupffer cell bactericidal activity on circulating Pseudomonas aeruginosa. Serum and liver tissues were collected to assess liver-injury biomarkers and histological changes, respectively.Results: We found that the liver was the major site of initial uptake of 65ZnO ENPs. There was a time-dependent decrease in tissue levels of 65Zn in all organs examined, reflecting particle dissolution. In vivo magnetometry showed a time-dependent and transient reduction in Kupffer cell phagosomal motility. Animals challenged with P. aeruginosa 24 hours post-ZnO ENP injection showed an initial (30 minutes delay in vascular bacterial clearance. However, by 4 hours, IV-injected bacteria were cleared from the blood, liver, spleen, lungs, and kidneys. Seven days post-ZnO ENP injection, creatine phosphokinase and aspartate aminotransferase levels in serum were significantly increased. Histological evidence of

  2. PREPARATION AND PURIFICATION OF DNA FROM BACTERIAL CELLS; CHARACTERIZATION OF PLASMID DNA

    Directory of Open Access Journals (Sweden)

    M Praveen, G Adarsh, T Ramesh, M Ramesh*

    2013-01-01

    Full Text Available The use of genetic material to deliver genes for therapeutic purposes has been practiced for many years. With the advancement in genetic engineering, foreign genes of industrial applications can be inserted into cloning vector for mass production in various host cells. Escherichia coli is an extremely important model organism in modern biological engineering, the suitable growth media is essential for the optimal expression of the genes in E. coli. The present study aims at isolation and purification of genomic DNA from E. coli, the characterization of pBR322 plasmid DNA. Bacterial culture conditions were optimized in shake – flask cultures based on optimal temperature, inoculum size and medium composition. Solutions and methods are disclosed for the effective, simple isolation of DNA from bacterial cells. High bioprocess recovery and product quality were primarily associated with the complete removal of total cellular RNA impurity. The process was demonstrated without the use of animal-derived RNase. High-molecular-weight (HMW RNA and other impurities were removed by selective precipitation using calcium chloride at an optimal concentration.The optimal conditions for the growth of Escherichia coli were shown maximum absorbance as 7.5 at 370C temperature, 1% inoculum size using TB medium composition. The purified genomic DNA had concentration as 73.5 µg/ml and purity 1.8. The 0.5M CaCl2 was optimal concentration for removal of RNA. The plasmid DNA pBR322 was confirmed by comparing the band to 4.36 Kb, purity of plasmid was 1.85 and it contains 96.8% of super coiled DNA. The contaminants like chromosomal DNA, RNA, host cell proteins and mycoplasma were absent in the plasmid DNA.

  3. Fluorescence-Activated Cell Sorting of Live Versus Dead Bacterial Cells and Spores

    Science.gov (United States)

    Bernardini, James N.; LaDuc, Myron T.; Diamond, Rochelle; Verceles, Josh

    2012-01-01

    This innovation is a coupled fluorescence-activated cell sorting (FACS) and fluorescent staining technology for purifying (removing cells from sampling matrices), separating (based on size, density, morphology, and live versus dead), and concentrating cells (spores, prokaryotic, eukaryotic) from an environmental sample.

  4. Antibiotic Discovery: Combatting Bacterial Resistance in Cells and in Biofilm Communities

    Directory of Open Access Journals (Sweden)

    Anahit Penesyan

    2015-03-01

    Full Text Available Bacterial resistance is a rapidly escalating threat to public health as our arsenal of effective antibiotics dwindles. Therefore, there is an urgent need for new antibiotics. Drug discovery has historically focused on bacteria growing in planktonic cultures. Many antibiotics were originally developed to target individual bacterial cells, being assessed in vitro against microorganisms in a planktonic mode of life. However, towards the end of the 20th century it became clear that many bacteria live as complex communities called biofilms in their natural habitat, and this includes habitats within a human host. The biofilm mode of life provides advantages to microorganisms, such as enhanced resistance towards environmental stresses, including antibiotic challenge. The community level resistance provided by biofilms is distinct from resistance mechanisms that operate at a cellular level, and cannot be overlooked in the development of novel strategies to combat infectious diseases. The review compares mechanisms of antibiotic resistance at cellular and community levels in the light of past and present antibiotic discovery efforts. Future perspectives on novel strategies for treatment of biofilm-related infectious diseases are explored.

  5. Bacterial porin disrupts mitochondrial membrane potential and sensitizes host cells to apoptosis.

    Directory of Open Access Journals (Sweden)

    Vera Kozjak-Pavlovic

    2009-10-01

    Full Text Available The bacterial PorB porin, an ATP-binding beta-barrel protein of pathogenic Neisseria gonorrhoeae, triggers host cell apoptosis by an unknown mechanism. PorB is targeted to and imported by host cell mitochondria, causing the breakdown of the mitochondrial membrane potential (DeltaPsi(m. Here, we show that PorB induces the condensation of the mitochondrial matrix and the loss of cristae structures, sensitizing cells to the induction of apoptosis via signaling pathways activated by BH3-only proteins. PorB is imported into mitochondria through the general translocase TOM but, unexpectedly, is not recognized by the SAM sorting machinery, usually required for the assembly of beta-barrel proteins in the mitochondrial outer membrane. PorB integrates into the mitochondrial inner membrane, leading to the breakdown of DeltaPsi(m. The PorB channel is regulated by nucleotides and an isogenic PorB mutant defective in ATP-binding failed to induce DeltaPsi(m loss and apoptosis, demonstrating that dissipation of DeltaPsi(m is a requirement for cell death caused by neisserial infection.

  6. Bacterial capsular polysaccharide prevents the onset of asthma through T-cell activation.

    Science.gov (United States)

    Johnson, Jenny L; Jones, Mark B; Cobb, Brian A

    2015-04-01

    Over the last four decades, increases in the incidence of immune-mediated diseases in the Western world have been linked to changes in microbial exposure. It is becoming increasingly clear that the normal microbiota in the gut can profoundly alter susceptibility to a wide range of diseases, such as asthma, in which immune homeostasis is disrupted, yet the mechanisms governing this microbial influence remains poorly defined. In this study, we show that gastrointestinal exposure to PSA, a capsular polysaccharide derived from the commensal bacterium Bacteroides fragilis, significantly limits susceptibility to the induction of experimental asthma. We report that direct treatment of mice with PSA generates protection from asthma, and this effect can be given to a naïve recipient by adoptive transfer of CD4(+) T cells from PSA-exposed mice. Remarkably, we found that these PSA-induced T cells are not canonical FoxP3(+) regulatory T cells, but that they potently inhibit both Th1 and Th2 models of asthma in an IL-10-dependent fashion. These findings reveal that bacterial polysaccharides link the microbiota with the peripheral immune system by activating CD4(+)Foxp3(-) T cells upon exposure in the gut, and they facilitate resistance to unnecessary inflammatory responses via the production of IL-10. PMID:25347992

  7. Cell resistant zwitterionic polyelectrolyte coating promotes bacterial attachment: an adhesion contradiction.

    Science.gov (United States)

    Martinez, Jessica S; Kelly, Kristopher D; Ghoussoub, Yara E; Delgado, Jose D; Keller Iii, Thomas C S; Schlenoff, Joseph B

    2016-04-22

    Polymers of various architectures with zwitterionic functionality have recently been shown to effectively suppress nonspecific fouling of surfaces by proteins and prokaryotic (bacteria) or eukaryotic (mammalian) cells as well as other microorganisms and environmental contaminants. In this work, zwitterionic copolymers were used to make thin coatings on substrates with the layer-by-layer method. Polyelectrolyte multilayers, PEMUs, were built with [poly(allylamine hydrochloride)], PAH, and copolymers of acrylic acid and either the AEDAPS zwitterionic group 3-[2-(acrylamido)-ethyldimethyl ammonio] propane sulfonate (PAA-co-AEDAPS), or benzophenone (PAABp). Benzophenone allowed the PEMU to be toughened by photocrosslinking post-deposition. The attachment of two mammalian cell lines, rat aortic smooth muscle (A7r5) and mouse fibroblasts (3T3), and the biofilm-forming Gram-negative bacteria Escherichia coli was studied on PEMUs terminated with PAA-co-AEDAPS. Consistent with earlier studies, it is shown that PAH/PAA-co-AEDAPS PEMUs resist the adhesion of mammalian cells, but, contrary to our initial hypothesis, are bacterial adhesive and significantly so after maximizing the surface presentation of PAA-co-AEDAPS. This unexpected contrast in the adhesive behavior of prokaryotic and eukaryotic cells is explained by differences in adhesion mechanisms as well as different responses to the topology and morphology of the multilayer surface. PMID:26872345

  8. Autonomous bioluminescent expression of the bacterial luciferase gene cassette (lux in a mammalian cell line.

    Directory of Open Access Journals (Sweden)

    Dan M Close

    Full Text Available The bacterial luciferase (lux gene cassette consists of five genes (luxCDABE whose protein products synergistically generate bioluminescent light signals exclusive of supplementary substrate additions or exogenous manipulations. Historically expressible only in prokaryotes, the lux operon was re-synthesized through a process of multi-bicistronic, codon-optimization to demonstrate for the first time self-directed bioluminescence emission in a mammalian HEK293 cell line in vitro and in vivo.Autonomous in vitro light production was shown to be 12-fold greater than the observable background associated with untransfected control cells. The availability of reduced riboflavin phosphate (FMNH(2 was identified as the limiting bioluminescence substrate in the mammalian cell environment even after the addition of a constitutively expressed flavin reductase gene (frp from Vibrio harveyi. FMNH(2 supplementation led to a 151-fold increase in bioluminescence in cells expressing mammalian codon-optimized luxCDE and frp genes. When injected subcutaneously into nude mice, in vivo optical imaging permitted near instantaneous light detection that persisted independently for the 60 min length of the assay with negligible background.The speed, longevity, and self-sufficiency of lux expression in the mammalian cellular environment provides a viable and powerful alternative for real-time target visualization not currently offered by existing bioluminescent and fluorescent imaging technologies.

  9. Cell proliferation, viability, and in vitro differentiation of equine mesenchymal stem cells seeded on bacterial cellulose hydrogel scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Favi, Pelagie M.; Benson, Roberto S. [Department of Materials Science and Engineering, College of Engineering, University of Tennessee, Knoxville, TN 37996 (United States); Neilsen, Nancy R. [Department of Biomedical and Diagnostic Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996 (United States); Hammonds, Ryan L. [Department of Materials Science and Engineering, College of Engineering, University of Tennessee, Knoxville, TN 37996 (United States); Bates, Cassandra C. [Department of Large Animal Clinical Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996 (United States); Stephens, Christopher P. [Department of Surgery, Graduate School of Medicine, University of Tennessee, Knoxville, TN 37996 (United States); Center for Materials Processing, University of Tennessee, Knoxville, TN 37996 (United States); Dhar, Madhu S., E-mail: mdhar@utk.edu [Department of Large Animal Clinical Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, TN 37996 (United States)

    2013-05-01

    The culture of multipotent mesenchymal stem cells on natural biopolymers holds great promise for treatments of connective tissue disorders such as osteoarthritis. The safety and performance of such therapies relies on the systematic in vitro evaluation of the developed stem cell-biomaterial constructs prior to in vivo implantation. This study evaluates bacterial cellulose (BC), a biocompatible natural polymer, as a scaffold for equine-derived bone marrow mesenchymal stem cells (EqMSCs) for application in bone and cartilage tissue engineering. An equine model was chosen due to similarities in size, load and types of joint injuries suffered by horses and humans. Lyophilized and critical point dried BC hydrogel scaffolds were characterized using scanning electron microscopy (SEM) to confirm nanostructure morphology which demonstrated that critical point drying induces fibre bundling unlike lyophilisation. EqMSCs positively expressed the undifferentiated pluripotent mesenchymal stem cell surface markers CD44 and CD90. The BC scaffolds were shown to be cytocompatible, supporting cellular adhesion and proliferation, and allowed for osteogenic and chondrogenic differentiation of EqMSCs. The cells seeded on the BC hydrogel were shown to be viable and metabolically active. These findings demonstrate that the combination of a BC hydrogel and EqMSCs are promising constructs for musculoskeletal tissue engineering applications. - Highlights: ► Critical point drying induces fibre bundling unlike lyophilisation. ► Cells positively expressed undifferentiated pluripotent stem cell markers. ► BCs were cytocompatible, supported cell adhesion, proliferation and differentiation ► Cells seeded on BC scaffolds were viable and metabolically active. ► Findings demonstrate that BC and EqMSCs are promising tissue engineered constructs.

  10. Cell proliferation, viability, and in vitro differentiation of equine mesenchymal stem cells seeded on bacterial cellulose hydrogel scaffolds

    International Nuclear Information System (INIS)

    The culture of multipotent mesenchymal stem cells on natural biopolymers holds great promise for treatments of connective tissue disorders such as osteoarthritis. The safety and performance of such therapies relies on the systematic in vitro evaluation of the developed stem cell-biomaterial constructs prior to in vivo implantation. This study evaluates bacterial cellulose (BC), a biocompatible natural polymer, as a scaffold for equine-derived bone marrow mesenchymal stem cells (EqMSCs) for application in bone and cartilage tissue engineering. An equine model was chosen due to similarities in size, load and types of joint injuries suffered by horses and humans. Lyophilized and critical point dried BC hydrogel scaffolds were characterized using scanning electron microscopy (SEM) to confirm nanostructure morphology which demonstrated that critical point drying induces fibre bundling unlike lyophilisation. EqMSCs positively expressed the undifferentiated pluripotent mesenchymal stem cell surface markers CD44 and CD90. The BC scaffolds were shown to be cytocompatible, supporting cellular adhesion and proliferation, and allowed for osteogenic and chondrogenic differentiation of EqMSCs. The cells seeded on the BC hydrogel were shown to be viable and metabolically active. These findings demonstrate that the combination of a BC hydrogel and EqMSCs are promising constructs for musculoskeletal tissue engineering applications. - Highlights: ► Critical point drying induces fibre bundling unlike lyophilisation. ► Cells positively expressed undifferentiated pluripotent stem cell markers. ► BCs were cytocompatible, supported cell adhesion, proliferation and differentiation ► Cells seeded on BC scaffolds were viable and metabolically active. ► Findings demonstrate that BC and EqMSCs are promising tissue engineered constructs

  11. Uncertain data envelopment analysis

    CERN Document Server

    Wen, Meilin

    2014-01-01

    This book is intended to present the milestones in the progression of uncertain Data envelopment analysis (DEA). Chapter 1 gives some basic introduction to uncertain theories, including probability theory, credibility theory, uncertainty theory and chance theory. Chapter 2 presents a comprehensive review and discussion of basic DEA models. The stochastic DEA is introduced in Chapter 3, in which the inputs and outputs are assumed to be random variables. To obtain the probability distribution of a random variable, a lot of samples are needed to apply the statistics inference approach. Chapter 4

  12. Coronatine inhibits stomatal closure and delays hypersensitive response cell death induced by nonhost bacterial pathogens

    Directory of Open Access Journals (Sweden)

    Seonghee Lee

    2013-02-01

    Full Text Available Pseudomonas syringae is the most widespread bacterial pathogen in plants. Several strains of P. syringae produce a phytotoxin, coronatine (COR, which acts as a jasmonic acid mimic and inhibits plant defense responses and contributes to disease symptom development. In this study, we found that COR inhibits early defense responses during nonhost disease resistance. Stomatal closure induced by a nonhost pathogen, P. syringae pv. tabaci, was disrupted by COR in tomato epidermal peels. In addition, nonhost HR cell death triggered by P. syringae pv. tabaci on tomato was remarkably delayed when COR was supplemented along with P. syringae pv. tabaci inoculation. Using isochorismate synthase (ICS-silenced tomato plants and transcript profiles of genes in SA- and JA-related defense pathways, we show that COR suppresses SA-mediated defense during nonhost resistance.

  13. Quantification of bioavailable chlortetracycline in pig feces using a bacterial whole-cell biosensor

    DEFF Research Database (Denmark)

    Hansen, L. H.; Aarestrup, Frank Møller; Sørensen, S. J.

    2002-01-01

    Bacterial whole-cell biosensors were used to measure the concentration of chlortetracycline (CTC) in the feces of pigs. In this study, the Escherichia coli biosensor used has a detection limit of 0.03 mg/kg CTC in pig feces. The tetracycline concentration was correlated with the appearance and...... maintenance of fecal coliform bacteria resistant to tetracycline. Initially, large quantities of water-extractable CTC were excreted from the pigs and measurable amounts were detected even at 30 days after treatment cessation. This led to a sharp rise in the number of tetracycline resistant coliform bacteria...... in the feces, to within the same order of magnitude as the total coliform count. The high level of tetracycline resistance was maintained in spite of the declining concentration of tetracycline. (C) 2002 Elsevier Science B.V. All rights reserved....

  14. Bacterial nanocellulose/Nafion composite membranes for low temperature polymer electrolyte fuel cells

    Science.gov (United States)

    Jiang, Gao-peng; Zhang, Jing; Qiao, Jin-li; Jiang, Yong-ming; Zarrin, Hadis; Chen, Zhongwei; Hong, Feng

    2015-01-01

    Novel nanocomposite membranes aimed for both proton-exchange membrane fuel cell (PEMFC) and direct methanol fuel cell (DMFC) are presented in this work. The membranes are based on blending bacterial nanocellulose pulp and Nafion (abbreviated as BxNy, where x and y indicates the mass ratio of bacterial cellulose to Nafion). The structure and properties of BxNy membranes are characterized by FTIR, SEM, TG, DMA and EIS, along with water uptake, swelling behavior and methanol permeability tests. It is found that the BxNy composite membranes with reinforced concrete-like structure show excellent mechanical and thermal stability regardless of annealing. The water uptake plus area and volume swelling ratios are all decreased compared to Nafion membranes. The proton conductivities of pristine and annealed B1N9 are 0.071 and 0.056 S cm-1, respectively, at 30 °C and 100% humidity. Specifically, annealed B1N1 exhibited the lowest methanol permeability of 7.21 × 10-7 cm2 s-1. Through the selectivity analysis, pristine and annealed B1N7 are selected to assemble the MEAs. The performances of annealed B1N7 in PEMFC and DMFC show the maximum power densities of 106 and 3.2 mW cm-2, respectively, which are much higher than those of pristine B1N7 at 25 °C. The performances of the pristine and annealed B1N7 reach a level as high as 21.1 and 20.4 mW cm-2 at 80 °C in DMFC, respectively.

  15. Decolorization of industrial synthetic dyes using engineered Pseudomonas putida cells with surface-immobilized bacterial laccase

    Directory of Open Access Journals (Sweden)

    Wang Wei

    2012-06-01

    Full Text Available Abstract Background Microbial laccases are highly useful in textile effluent dye biodegradation. However, the bioavailability of cellularly expressed or purified laccases in continuous operations is usually limited by mass transfer impediment or enzyme regeneration difficulty. Therefore, this study develops a regenerable bacterial surface-displaying system for industrial synthetic dye decolorization, and evaluates its effects on independent and continuous operations. Results A bacterial laccase (WlacD was engineered onto the cell surface of the solvent-tolerant bacterium Pseudomonas putida to construct a whole-cell biocatalyst. Ice nucleation protein (InaQ anchor was employed, and the ability of 1 to 3 tandemly aligned N-terminal repeats to direct WlacD display were compared. Immobilized WlacD was determined to be surface-displayed in functional form using Western blot analysis, immunofluorescence microscopy, flow cytometry, and whole-cell enzymatic activity assay. Engineered P. putida cells were then applied to decolorize the anthraquinone dye Acid Green (AG 25 and diazo-dye Acid Red (AR 18. The results showed that decolorization of both dyes is Cu2+- and mediator-independent, with an optimum temperature of 35°C and pH of 3.0, and can be stably performed across a temperature range of 15°C to 45°C. A high activity toward AG25 (1 g/l with relative decolorization values of 91.2% (3 h and 97.1% (18 h, as well as high activity to AR18 (1 g/l by 80.5% (3 h and 89.0% (18 h, was recorded. The engineered system exhibited a comparably high activity compared with those of separate dyes in a continuous three-round shake-flask decolorization of AG25/AR18 mixed dye (each 1 g/l. No significant decline in decolorization efficacy was noted during first two-rounds but reaction equilibriums were elongated, and the residual laccase activity eventually decreased to low levels. However, the decolorizing capacity of the system was easily retrieved

  16. Shaping T Cell – B Cell Collaboration in the Response to Human Immunodeficiency Virus Type 1 Envelope Glycoprotein gp120 by Peptide Priming

    OpenAIRE

    N Kalaya Steede; Rust, Blake J.; Hossain, Mohammad M.; Freytag, Lucy C.; Robinson, James E.; Landry, Samuel J.

    2013-01-01

    Prime-boost vaccination regimes have shown promise for obtaining protective immunity to HIV. Poorly understood mechanisms of cellular immunity could be responsible for improved humoral responses. Although CD4+ T-cell help promotes B-cell development, the relationship of CD4+ T-cell specificity to antibody specificity has not been systematically investigated. Here, protein and peptide-specific immune responses to HIV-1 gp120 were characterized in groups of ten mucosally immunized BALB/c mice. ...

  17. Quantification of the lateral detachment force for bacterial cells using atomic force microscope and centrifugation

    International Nuclear Information System (INIS)

    To determine the lateral detachment force for individual bacterial cells, a quantitative method using the contact mode of an atomic force microscope (AFM) was developed in this study. Three key factors for the proposed method, i.e. scan size, scan rate and cantilever choice, were evaluated and optimized. The scan size of 40x40 μm2 was optimal for capturing sufficient number of adhered cells in a microscopic field and provide adequate information for cell identification and detachment force measurement. The scan rate affected the measurement results significantly, and was optimized at 40 μm/s considering both force measurement accuracy and experimental efficiency. The hardness of applied cantilevers also influenced force determination. The proposed protocol for cantilever selection is to use those with the lowest spring constant first and then step up to a harder cantilever until all cells are detached. The lateral detachment force of Escherichia coli cells on polished stainless steel and a glass-slide coated with poly-L-lysine were measured as 0.763±0.167 and 0.639±0.136 nN, respectively. The results showed that the established method had good repeatability and sensitivity to various bacteria/substrata combinations. The detachment force quantified by AFM (0.639±0.136 nN) was comparable to that measured by the centrifugation method (1.12 nN). -- Research highlights: → A quantitative method via AFM is developed to measure the lateral detachment force of an attached cell. → The parameters of AFM operation for this method are optimized. → The tests using E. coli on different substrata show that the method has good repeatability and sensitivity. → The method could obtain reliable results that are comparable to those using the centrifugation approach.

  18. Nano-ranged low-energy ion-beam-induced DNA transfer in biological cells

    Science.gov (United States)

    Yu, L. D.; Wongkham, W.; Prakrajang, K.; Sangwijit, K.; Inthanon, K.; Thongkumkoon, P.; Wanichapichart, P.; Anuntalabhochai, S.

    2013-06-01

    Low-energy ion beams at a few tens of keV were demonstrated to be able to induce exogenous macromolecules to transfer into plant and bacterial cells. In the process, the ion beam with well controlled energy and fluence bombarded living cells to cause certain degree damage in the cell envelope in nanoscales to facilitate the macromolecules such as DNA to pass through the cell envelope and enter the cell. Consequently, the technique was applied for manipulating positive improvements in the biological species. This physical DNA transfer method was highly efficient and had less risk of side-effects compared with chemical and biological methods. For better understanding of mechanisms involved in the process, a systematic study on the mechanisms was carried out. Applications of the technique were also expanded from DNA transfer in plant and bacterial cells to DNA transfection in human cancer cells potentially for the stem cell therapy purpose. Low-energy nitrogen and argon ion beams that were applied in our experiments had ranges of 100 nm or less in the cell envelope membrane which was majorly composed of polymeric cellulose. The ion beam bombardment caused chain-scission dominant damage in the polymer and electrical property changes such as increase in the impedance in the envelope membrane. These nano-modifications of the cell envelope eventually enhanced the permeability of the envelope membrane to favor the DNA transfer. The paper reports details of our research in this direction.

  19. Nano-ranged low-energy ion-beam-induced DNA transfer in biological cells

    Energy Technology Data Exchange (ETDEWEB)

    Yu, L.D., E-mail: yuld@fnrf.science.cmu.ac.th [Thailand Center of Excellence in Physics, Commission on Higher Education, 328 Si Ayutthaya Road, Bangkok 10400 (Thailand); Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Wongkham, W. [Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Prakrajang, K. [Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Sangwijit, K.; Inthanon, K. [Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Thongkumkoon, P. [Thailand Center of Excellence in Physics, Commission on Higher Education, 328 Si Ayutthaya Road, Bangkok 10400 (Thailand); Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Wanichapichart, P. [Thailand Center of Excellence in Physics, Commission on Higher Education, 328 Si Ayutthaya Road, Bangkok 10400 (Thailand); Membrane Science and Technology Research Center, Department of Physics, Faculty of Science, Prince of Songkla University, Hat Yai, Songkla 90112 (Thailand); Anuntalabhochai, S. [Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand)

    2013-06-15

    Low-energy ion beams at a few tens of keV were demonstrated to be able to induce exogenous macromolecules to transfer into plant and bacterial cells. In the process, the ion beam with well controlled energy and fluence bombarded living cells to cause certain degree damage in the cell envelope in nanoscales to facilitate the macromolecules such as DNA to pass through the cell envelope and enter the cell. Consequently, the technique was applied for manipulating positive improvements in the biological species. This physical DNA transfer method was highly efficient and had less risk of side-effects compared with chemical and biological methods. For better understanding of mechanisms involved in the process, a systematic study on the mechanisms was carried out. Applications of the technique were also expanded from DNA transfer in plant and bacterial cells to DNA transfection in human cancer cells potentially for the stem cell therapy purpose. Low-energy nitrogen and argon ion beams that were applied in our experiments had ranges of 100 nm or less in the cell envelope membrane which was majorly composed of polymeric cellulose. The ion beam bombardment caused chain-scission dominant damage in the polymer and electrical property changes such as increase in the impedance in the envelope membrane. These nano-modifications of the cell envelope eventually enhanced the permeability of the envelope membrane to favor the DNA transfer. The paper reports details of our research in this direction.

  20. Nano-ranged low-energy ion-beam-induced DNA transfer in biological cells

    International Nuclear Information System (INIS)

    Low-energy ion beams at a few tens of keV were demonstrated to be able to induce exogenous macromolecules to transfer into plant and bacterial cells. In the process, the ion beam with well controlled energy and fluence bombarded living cells to cause certain degree damage in the cell envelope in nanoscales to facilitate the macromolecules such as DNA to pass through the cell envelope and enter the cell. Consequently, the technique was applied for manipulating positive improvements in the biological species. This physical DNA transfer method was highly efficient and had less risk of side-effects compared with chemical and biological methods. For better understanding of mechanisms involved in the process, a systematic study on the mechanisms was carried out. Applications of the technique were also expanded from DNA transfer in plant and bacterial cells to DNA transfection in human cancer cells potentially for the stem cell therapy purpose. Low-energy nitrogen and argon ion beams that were applied in our experiments had ranges of 100 nm or less in the cell envelope membrane which was majorly composed of polymeric cellulose. The ion beam bombardment caused chain-scission dominant damage in the polymer and electrical property changes such as increase in the impedance in the envelope membrane. These nano-modifications of the cell envelope eventually enhanced the permeability of the envelope membrane to favor the DNA transfer. The paper reports details of our research in this direction.

  1. Atomic Force Microscopy Measurements of the Mechanical Properties of Cell Walls on Living Bacterial Cells

    Science.gov (United States)

    Bailey, Richard; Mullin, Nic; Turner, Robert; Foster, Simon; Hobbs, Jamie

    2014-03-01

    Staphylococcus aureus is a major cause of infection in humans, including the Methicillin resistant strain, MRSA. However, very little is known about the mechanical properties of these cells. Our investigations use AFM to examine live S. aureus cells to quantify mechanical properties. These were explored using force spectroscopy with different trigger forces, allowing the properties to be extracted at different indentation depths. A value for the cell wall stiffness has been extracted, along with a second, higher value which is found upon indenting at higher forces. This higher value drops as the cells are exposed to high salt, sugar and detergent concentrations, implying that this measurement contains a contribution from the internal turgor pressure. We have monitored these properties as the cells progress through the cell cycle. Force maps were taken over the cells at different stages of the growth process to identify changes in the mechanics throughout the progression of growth and division. The effect of Oxacillin has also been studied, to better understand its mechanism of action. Finally mutant strains of S. aureus and a second species Bacillus subtilis have been used to link the mechanical properties of the cell walls with the chain lengths and substructures involved.

  2. Quantitative investigation of bacterial chemotaxis at the single-cell level

    Science.gov (United States)

    Min, Taejin

    Living cells sense and respond to constantly changing environmental conditions. Depending on the type of stimuli, the cell may response by altering gene expression pattern, secreting molecules, or migrating to a different environment. Directed movement of cells in response to chemical stimuli is called chemotaxis. In bacterial chemotaxis, small extracellular molecules bind receptor proteins embedded in the cell membrane, which then transmit the signal inside the cell through a cascade of protein-protein interactions. This chain of events influences the behavior of motor proteins that drive the rotation of helical filaments called flagella. Individual cells of the gut-dwelling bacteria Escherichia coli (E. coli) have many such flagella, whose collective action results in the swimming behavior of the cell. A recent study found that in absence of chemical stimuli, fluctuations in the protein cascade can cause non-Poissonian switching behavior in the flagellar motor (2). A corollary was that extension of such behavior to the whole-cell swimming level would have implications for E. coli's foraging strategy. However, existence of such behavior at the swimming cell level could not be predicted a priori, since the mapping from single flagellum behavior to the swimming behavior of a multi-flagellated cell is complex and poorly understood (3, 4). Here we characterize the chemotactic behavior of swimming E. coli cells using a novel optical trap-based measurement technique. This technique allows us to trap individual cells and monitor their swimming behavior over long time periods with high temporal resolution. We find that swimming cells exhibit non-Poissonian switching statistics between different swimming states, in a manner similar to the rotational direction-switching behavior seen in individual flagella. Furthermore, we develop a data analysis routine that allows us to characterize higher order swimming features such as reversal of swimming direction and existence of

  3. Phenotypic T cell exhaustion in a murine model of bacterial infection in the setting of pre-existing malignancy.

    Science.gov (United States)

    Mittal, Rohit; Wagener, Maylene; Breed, Elise R; Liang, Zhe; Yoseph, Benyam P; Burd, Eileen M; Farris, Alton B; Coopersmith, Craig M; Ford, Mandy L

    2014-01-01

    While much of cancer immunology research has focused on anti-tumor immunity both systemically and within the tumor microenvironment, little is known about the impact of pre-existing malignancy on pathogen-specific immune responses. Here, we sought to characterize the antigen-specific CD8+ T cell response following a bacterial infection in the setting of pre-existing pancreatic adenocarcinoma. Mice with established subcutaneous pancreatic adenocarcinomas were infected with Listeria monocytogenes, and antigen-specific CD8+ T cell responses were compared to those in control mice without cancer. While the kinetics and magnitude of antigen-specific CD8+ T cell expansion and accumulation was comparable between the cancer and non-cancer groups, bacterial antigen-specific CD8+ T cells and total CD4+ and CD8+ T cells in cancer mice exhibited increased expression of the coinhibitory receptors BTLA, PD-1, and 2B4. Furthermore, increased inhibitory receptor expression was associated with reduced IFN-γ and increased IL-2 production by bacterial antigen-specific CD8+ T cells in the cancer group. Taken together, these data suggest that cancer's immune suppressive effects are not limited to the tumor microenvironment, but that pre-existing malignancy induces phenotypic exhaustion in T cells by increasing expression of coinhibitory receptors and may impair pathogen-specific CD8+ T cell functionality and differentiation. PMID:24796533

  4. Phenotypic T cell exhaustion in a murine model of bacterial infection in the setting of pre-existing malignancy.

    Directory of Open Access Journals (Sweden)

    Rohit Mittal

    Full Text Available While much of cancer immunology research has focused on anti-tumor immunity both systemically and within the tumor microenvironment, little is known about the impact of pre-existing malignancy on pathogen-specific immune responses. Here, we sought to characterize the antigen-specific CD8+ T cell response following a bacterial infection in the setting of pre-existing pancreatic adenocarcinoma. Mice with established subcutaneous pancreatic adenocarcinomas were infected with Listeria monocytogenes, and antigen-specific CD8+ T cell responses were compared to those in control mice without cancer. While the kinetics and magnitude of antigen-specific CD8+ T cell expansion and accumulation was comparable between the cancer and non-cancer groups, bacterial antigen-specific CD8+ T cells and total CD4+ and CD8+ T cells in cancer mice exhibited increased expression of the coinhibitory receptors BTLA, PD-1, and 2B4. Furthermore, increased inhibitory receptor expression was associated with reduced IFN-γ and increased IL-2 production by bacterial antigen-specific CD8+ T cells in the cancer group. Taken together, these data suggest that cancer's immune suppressive effects are not limited to the tumor microenvironment, but that pre-existing malignancy induces phenotypic exhaustion in T cells by increasing expression of coinhibitory receptors and may impair pathogen-specific CD8+ T cell functionality and differentiation.

  5. Sensing the Structural Differences in Cellulose from Apple and Bacterial Cell Wall Materials by Raman and FT-IR Spectroscopy

    Directory of Open Access Journals (Sweden)

    Artur Zdunek

    2011-05-01

    Full Text Available Raman and Fourier Transform Infrared (FT-IR spectroscopy was used for assessment of structural differences of celluloses of various origins. Investigated celluloses were: bacterial celluloses cultured in presence of pectin and/or xyloglucan, as well as commercial celluloses and cellulose extracted from apple parenchyma. FT-IR spectra were used to estimate of the Iβ content, whereas Raman spectra were used to evaluate the degree of crystallinity of the cellulose. The crystallinity index (XCRAMAN% varied from −25% for apple cellulose to 53% for microcrystalline commercial cellulose. Considering bacterial cellulose, addition of xyloglucan has an impact on the percentage content of cellulose Iβ. However, addition of only xyloglucan or only pectins to pure bacterial cellulose both resulted in a slight decrease of crystallinity. However, culturing bacterial cellulose in the presence of mixtures of xyloglucan and pectins results in an increase of crystallinity. The results confirmed that the higher degree of crystallinity, the broader the peak around 913 cm−1. Among all bacterial celluloses the bacterial cellulose cultured in presence of xyloglucan and pectin (BCPX has the most similar structure to those observed in natural primary cell walls.

  6. [Disinfectants - bacterial cells interactions in the view of hygiene and public health].

    Science.gov (United States)

    Książczyk, Marta; Krzyżewska, Eva; Futoma-Kołoch, Bożena; Bugla-Płoskońska, Gabriela

    2015-01-01

    In recent years, the use of biocides has increased rapidly. One common example is triclosan, with wide application in households as well as medical and industrial fields, especially food industry and animal husbandry. Chemical disinfection is a major mean to control and eliminate pathogenic bacteria, particularly those with multidrug resistance (MDR) phenotype. However, exposition to biocides results in an adaptive response in microorganisms, causing them to display a wide range of resistance mechanisms. Numerous microorganisms are characterized by either natural resistance to chemical compounds or an ability to adapt to biocides using various strategies, such as: modification of cell surface structures (lipopolisaccharide), membrane fatty acids), over-expression of efflux pumps (a system for active transport of toxic compounds out of bacterial cell), enzymatic inactivation of biocides or altering biocide targets. For instance, it was shown that in vitro exposition of Salmonella Typhimurium to subinhibitory concentration of biocides (triclosan, quaternary ammonium compounds [QACs]) resulted in selection of variants resistant to tested biocides and, additionally, to acridine dyes and antibiotics. Bacillus subtilis and Micrococcus luteus strains isolated from chlorine dioxide containing disinfection devices were found to be resistant to chlorine dioxide and also to other oxidizing compounds, such as peracetic acid and hydrogen peroxide. Interaction between chemical compounds, including disinfectants and microbial cells, can create a serious threat to public health and sanitary-hygienic security. This phenomenon is connected with factor risk that intensify the probability of selection and dissemination of multidrug resistance among pathogenic bacteria. PMID:26400890

  7. Shaping T Cell – B Cell Collaboration in the Response to Human Immunodeficiency Virus Type 1 Envelope Glycoprotein gp120 by Peptide Priming

    Science.gov (United States)

    Steede, N. Kalaya; Rust, Blake J.; Hossain, Mohammad M.; Freytag, Lucy C.; Robinson, James E.; Landry, Samuel J.

    2013-01-01

    Prime-boost vaccination regimes have shown promise for obtaining protective immunity to HIV. Poorly understood mechanisms of cellular immunity could be responsible for improved humoral responses. Although CD4+ T-cell help promotes B-cell development, the relationship of CD4+ T-cell specificity to antibody specificity has not been systematically investigated. Here, protein and peptide-specific immune responses to HIV-1 gp120 were characterized in groups of ten mucosally immunized BALB/c mice. Protein and peptide reactivity of serum antibody was tested for correlation with cytokine secretion by splenocytes restimulated with individual gp120 peptides. Antibody titer for gp120 correlated poorly with the peptide-stimulated T-cell response. In contrast, titers for conformational epitopes, measured as crossreactivity or CD4-blocking, correlated with average interleukin-2 and interleukin-5 production in response to gp120 peptides. Antibodies specific for conformational epitopes and individual gp120 peptides typically correlated with T-cell responses to several peptides. In order to modify the specificity of immune responses, animals were primed with a gp120 peptide prior to immunization with protein. Priming induced distinct peptide-specific correlations of antibodies and T-cells. The majority of correlated antibodies were specific for the primed peptides or other peptides nearby in the gp120 sequence. These studies suggest that the dominant B-cell subsets recruit the dominant T-cell subsets and that T-B collaborations can be shaped by epitope-specific priming. PMID:23776539

  8. Shaping T cell - B cell collaboration in the response to human immunodeficiency virus type 1 envelope glycoprotein gp120 by peptide priming.

    Directory of Open Access Journals (Sweden)

    N Kalaya Steede

    Full Text Available Prime-boost vaccination regimes have shown promise for obtaining protective immunity to HIV. Poorly understood mechanisms of cellular immunity could be responsible for improved humoral responses. Although CD4+ T-cell help promotes B-cell development, the relationship of CD4+ T-cell specificity to antibody specificity has not been systematically investigated. Here, protein and peptide-specific immune responses to HIV-1 gp120 were characterized in groups of ten mucosally immunized BALB/c mice. Protein and peptide reactivity of serum antibody was tested for correlation with cytokine secretion by splenocytes restimulated with individual gp120 peptides. Antibody titer for gp120 correlated poorly with the peptide-stimulated T-cell response. In contrast, titers for conformational epitopes, measured as crossreactivity or CD4-blocking, correlated with average interleukin-2 and interleukin-5 production in response to gp120 peptides. Antibodies specific for conformational epitopes and individual gp120 peptides typically correlated with T-cell responses to several peptides. In order to modify the specificity of immune responses, animals were primed with a gp120 peptide prior to immunization with protein. Priming induced distinct peptide-specific correlations of antibodies and T-cells. The majority of correlated antibodies were specific for the primed peptides or other peptides nearby in the gp120 sequence. These studies suggest that the dominant B-cell subsets recruit the dominant T-cell subsets and that T-B collaborations can be shaped by epitope-specific priming.

  9. Emergence of vertebrate retroviruses and envelope capture

    International Nuclear Information System (INIS)

    Retroviruses are members of the superfamily of retroelements, mobile genetic elements that transpose via an RNA intermediate. However, retroviruses are distinct from other retroelements in that their 'transposition' is not confined to single cells but extends to neighboring cells and organisms. As such, the 'transposition' of these elements is defined as infection. It appears that a key step in the conversion of a retrotransposon into a retrovirus is the modular acquisition or capture of an envelope glycoprotein (Env) which facilitates dissemination from its initial host cell. Here we present several examples of retroviruses for which envelope capture has been identified. Indeed, capture may explain the notable conservation of env sequences among otherwise phylogenetically distant retroviruses. In a recent example, sequence homologies reported between the env of the phylogenetically distant murine leukemia viruses (MLV) and human T cell leukemia viruses (HTLV) argue in favor of an env capture by the latter. Env acquisition can provide new adaptive properties to replication-competent viruses in addition to altering their host range. Also, the captured env can alter the spectrum of physiological affects of infection in new host cells and organisms. The elucidation of such envelope exchanges and properties thereof should contribute significantly to the clarification of retroviral phylogeny, insight into retroviral pathogenesis, and to the discovery of new retroviruses

  10. Evaluation of an in vitro cell assay to select attenuated bacterial mutants of Aeromonas hydrophila and Edwardsiella tarda to channel catfish

    Science.gov (United States)

    To evaluate the feasibility of using an in vitro cell assay to select attenuated bacterial mutants. Using catfish gill cells G1B, the feasibility of using an in vitro assay instead of in vivo virulence assay using live fish to select attenuated bacterial mutants was evaluated in this study. Pearson ...

  11. Structure of the complex between teicoplanin and a bacterial cell-wall peptide: use of a carrier-protein approach

    Energy Technology Data Exchange (ETDEWEB)

    Economou, Nicoleta J.; Zentner, Isaac J. [Drexel University College of Medicine, 245 North 15th Street, Philadelphia, PA 19102 (United States); Lazo, Edwin; Jakoncic, Jean; Stojanoff, Vivian [Brookhaven National Laboratory, Upton, NY 11973 (United States); Weeks, Stephen D.; Grasty, Kimberly C.; Cocklin, Simon; Loll, Patrick J. [Drexel University College of Medicine, 245 North 15th Street, Philadelphia, PA 19102 (United States)

    2013-04-01

    Using a carrier-protein strategy, the structure of teicoplanin bound to its bacterial cell-wall target has been determined. The structure reveals the molecular determinants of target recognition, flexibility in the antibiotic backbone and intrinsic radiation sensitivity of teicoplanin. Multidrug-resistant bacterial infections are commonly treated with glycopeptide antibiotics such as teicoplanin. This drug inhibits bacterial cell-wall biosynthesis by binding and sequestering a cell-wall precursor: a d-alanine-containing peptide. A carrier-protein strategy was used to crystallize the complex of teicoplanin and its target peptide by fusing the cell-wall peptide to either MBP or ubiquitin via native chemical ligation and subsequently crystallizing the protein–peptide–antibiotic complex. The 2.05 Å resolution MBP–peptide–teicoplanin structure shows that teicoplanin recognizes its ligand through a combination of five hydrogen bonds and multiple van der Waals interactions. Comparison of this teicoplanin structure with that of unliganded teicoplanin reveals a flexibility in the antibiotic peptide backbone that has significant implications for ligand recognition. Diffraction experiments revealed an X-ray-induced dechlorination of the sixth amino acid of the antibiotic; it is shown that teicoplanin is significantly more radiation-sensitive than other similar antibiotics and that ligand binding increases radiosensitivity. Insights derived from this new teicoplanin structure may contribute to the development of next-generation antibacterials designed to overcome bacterial resistance.

  12. Structure of the complex between teicoplanin and a bacterial cell-wall peptide: use of a carrier-protein approach

    International Nuclear Information System (INIS)

    Using a carrier-protein strategy, the structure of teicoplanin bound to its bacterial cell-wall target has been determined. The structure reveals the molecular determinants of target recognition, flexibility in the antibiotic backbone and intrinsic radiation sensitivity of teicoplanin. Multidrug-resistant bacterial infections are commonly treated with glycopeptide antibiotics such as teicoplanin. This drug inhibits bacterial cell-wall biosynthesis by binding and sequestering a cell-wall precursor: a d-alanine-containing peptide. A carrier-protein strategy was used to crystallize the complex of teicoplanin and its target peptide by fusing the cell-wall peptide to either MBP or ubiquitin via native chemical ligation and subsequently crystallizing the protein–peptide–antibiotic complex. The 2.05 Å resolution MBP–peptide–teicoplanin structure shows that teicoplanin recognizes its ligand through a combination of five hydrogen bonds and multiple van der Waals interactions. Comparison of this teicoplanin structure with that of unliganded teicoplanin reveals a flexibility in the antibiotic peptide backbone that has significant implications for ligand recognition. Diffraction experiments revealed an X-ray-induced dechlorination of the sixth amino acid of the antibiotic; it is shown that teicoplanin is significantly more radiation-sensitive than other similar antibiotics and that ligand binding increases radiosensitivity. Insights derived from this new teicoplanin structure may contribute to the development of next-generation antibacterials designed to overcome bacterial resistance

  13. Simultaneous determination of gene expression and bacterial identity in single cells in defined mixtures of pure cultures

    DEFF Research Database (Denmark)

    Poulsen, Lars K.; Dalton, Helen M.; Angels, Mark; Marshall, Kevin C.; Molin, Søren; Goodman, Amanda E.

    1997-01-01

    A protocol was developed to achieve the simultaneous determination of gene expression and bacterial identity at the level of single cells: a chromogenic beta-galactosidase activity assay was combined with in situ hybridization of Fluorescently labelled oligonucleotide probes to rRNA. The method a...

  14. Adaptive Architectural Envelope

    DEFF Research Database (Denmark)

    Foged, Isak Worre; Kirkegaard, Poul Henning

    2010-01-01

    Recent years have seen an increasing variety of applications of adaptive architectural structures for improvement of structural performance by recognizing changes in their environments and loads, adapting to meet goals, and using past events to improve future performance or maintain serviceability....... The general scopes of this paper are to develop a new adaptive kinetic architectural structure, particularly a reconfigurable architectural structure which can transform body shape from planar geometries to hyper-surfaces using different control strategies, i.e. a transformation into more than one or...... two different shape alternatives. The adaptive structure is a proposal for a responsive building envelope which is an idea of a first level operational framework for present and future investigations towards performance based responsive architectures through a set of responsive typologies. A mock- up...

  15. Retinoid suppression of transglutaminase activity and envelope competence in cultured human epidermal carcinoma cells: Hydrocortisone is a potent antagonist of retinyl acetate but not retinoic acid

    OpenAIRE

    Rice, Rh; Thacher, SM; Coe, EL

    1985-01-01

    Growth of SCC-13 squamous carcinoma cultures in the presence of retinoids considerably reduced the expression of two differentiation markers, the cellular capability to form cross-linked envelopes, and the enzyme transglutaminase required for cross-linking. A limited survey of retinoids showed that all-trans retinoic acid, 13-cis retinoic acid, and arotinoid Ro 13-6298 were highly effective in the absence of hydrocortisone and were only slightly antagonized by its presence in the medium. In c...

  16. Phospholipase D promotes Arcanobacterium haemolyticum adhesion via lipid raft remodeling and host cell death following bacterial invasion

    Directory of Open Access Journals (Sweden)

    Carlson Petteri

    2010-10-01

    Full Text Available Abstract Background Arcanobacterium haemolyticum is an emerging bacterial pathogen, causing pharyngitis and more invasive infections. This organism expresses an unusual phospholipase D (PLD, which we propose promotes bacterial pathogenesis through its action on host cell membranes. The pld gene is found on a genomic region of reduced %G + C, suggesting recent horizontal acquisition. Results Recombinant PLD rearranged HeLa cell lipid rafts in a dose-dependent manner and this was inhibited by cholesterol sequestration. PLD also promoted host cell adhesion, as a pld mutant had a 60.3% reduction in its ability to adhere to HeLa cells as compared to the wild type. Conversely, the pld mutant appeared to invade HeLa cells approximately two-fold more efficiently as the wild type. This finding was attributable to a significant loss of host cell viability following secretion of PLD from intracellular bacteria. As determined by viability assay, only 15.6% and 82.3% of HeLa cells remained viable following invasion by the wild type or pld mutant, respectively, as compared to untreated HeLa cells. Transmission electron microscopy of HeLa cells inoculated with A. haemolyticum strains revealed that the pld mutant was contained within intracellular vacuoles, as compared to the wild type, which escaped the vacuole. Wild type-infected HeLa cells also displayed the hallmarks of necrosis. Similarly inoculated HeLa cells displayed no signs of apoptosis, as measured by induction of caspase 3/7, 8 or 9 activities. Conclusions These data indicate that PLD enhances bacterial adhesion and promotes host cell necrosis following invasion, and therefore, may be important in the disease pathogenesis of A. haemolyticum infections.

  17. A 16 × 16 CMOS Capacitive Biosensor Array Towards Detection of Single Bacterial Cell.

    Science.gov (United States)

    Couniot, Numa; Francis, Laurent A; Flandre, Denis

    2016-04-01

    We present a 16 × 16 CMOS biosensor array aiming at impedance detection of whole-cell bacteria. Each 14 μm × 16 μm pixel comprises high-sensitive passivated microelectrodes connected to an innovative readout interface based on charge sharing principle for capacitance-to-voltage conversion and subthreshold gain stage to boost the sensitivity. Fabricated in a 0.25 μm CMOS process, the capacitive array was experimentally shown to perform accurate dielectric measurements of the electrolyte up to electrical conductivities of 0.05 S/m, with maximal sensitivity of 55 mV/fF and signal-to-noise ratio (SNR) of 37 dB. As biosensing proof of concept, real-time detection of Staphylococcus epidermidis binding events was experimentally demonstrated and provides detection limit of ca. 7 bacteria per pixel and sensitivity of 2.18 mV per bacterial cell. Models and simulations show good matching with experimental results and provide a comprehensive analysis of the sensor and circuit system. Advantages, challenges and limits of the proposed capacitive biosensor array are finally described with regards to literature. With its small area and low power consumption, the present capacitive array is particularly suitable for portable point-of-care (PoC) diagnosis tools and lab-on-chip (LoC) systems. PMID:25974947

  18. The impact of hypoxia on intestinal epithelial cell functions: consequences for invasion by bacterial pathogens.

    Science.gov (United States)

    Zeitouni, Nathalie E; Chotikatum, Sucheera; von Köckritz-Blickwede, Maren; Naim, Hassan Y

    2016-12-01

    The maintenance of oxygen homeostasis in human tissues is mediated by several cellular adaptations in response to low-oxygen stress, called hypoxia. A decrease in tissue oxygen levels is initially counteracted by increasing local blood flow to overcome diminished oxygenation and avoid hypoxic stress. However, studies have shown that the physiological oxygen concentrations in several tissues are much lower than atmospheric (normoxic) conditions, and the oxygen supply is finely regulated in individual cell types. The gastrointestinal tract has been described to subsist in a state of physiologically low oxygen level and is thus depicted as a tissue in the state of constant low-grade inflammation. The intestinal epithelial cell layer plays a vital role in the immune response to inflammation and infections that occur within the intestinal tissue and is involved in many of the adaptation responses to hypoxic stress. This is especially relevant in the context of inflammatory disorders, such as inflammatory bowel disease (IBD). Therefore, this review aims to describe the intestinal epithelial cellular response to hypoxia and the consequences for host interactions with invading gastrointestinal bacterial pathogens. PMID:27002817

  19. Mitomycin resistance in mammalian cells expressing the bacterial mitomycin C resistance protein MCRA.

    Science.gov (United States)

    Belcourt, M F; Penketh, P G; Hodnick, W F; Johnson, D A; Sherman, D H; Rockwell, S; Sartorelli, A C

    1999-08-31

    The mitomycin C-resistance gene, mcrA, of Streptomyces lavendulae produces MCRA, a protein that protects this microorganism from its own antibiotic, the antitumor drug mitomycin C. Expression of the bacterial mcrA gene in mammalian Chinese hamster ovary cells causes profound resistance to mitomycin C and to its structurally related analog porfiromycin under aerobic conditions but produces little change in drug sensitivity under hypoxia. The mitomycins are prodrugs that are enzymatically reduced and activated intracellularly, producing cytotoxic semiquinone anion radical and hydroquinone reduction intermediates. In vitro, MCRA protects DNA from cross-linking by the hydroquinone reduction intermediate of these mitomycins by oxidizing the hydroquinone back to the parent molecule; thus, MCRA acts as a hydroquinone oxidase. These findings suggest potential therapeutic applications for MCRA in the treatment of cancer with the mitomycins and imply that intrinsic or selected mitomycin C resistance in mammalian cells may not be due solely to decreased bioactivation, as has been hypothesized previously, but instead could involve an MCRA-like mechanism. PMID:10468636

  20. Modified bacterial cellulose scaffolds for localized doxorubicin release in human colorectal HT-29 cells.

    Science.gov (United States)

    Cacicedo, Maximiliano L; León, Ignacio E; Gonzalez, Jimena S; Porto, Luismar M; Alvarez, Vera A; Castro, Guillermo R

    2016-04-01

    Bacterial cellulose (BC) films modified by the in situ method with the addition of alginate (Alg) during the microbial cultivation of Gluconacetobacter hansenii under static conditions increased the loading of doxorubicin by at least three times. Biophysical analysis of BC-Alg films by scanning electron microscopy, thermogravimetry, X-ray diffraction and FTIR showed a highly homogeneous interpenetrated network scaffold without changes in the BC crystalline structure but with an increased amorphous phase. The main molecular interactions determined by FTIR between both biopolymers clearly suggest high compatibility. These results indicate that alginate plays a key role in the biophysical properties of the hybrid BC matrix. BC-Alg scaffold analysis by nitrogen adsorption isotherms revealed by the Brunauer-Emmett-Teller (BET) method an increase in surface area of about 84% and in pore volume of more than 200%. The Barrett-Joyner-Halenda (BJH) model also showed an increase of about 25% in the pore size compared to the BC film. Loading BC-Alg scaffolds with different amounts of doxorubicin decreased the cell viability of HT-29 human colorectal adenocarcinoma cell line compared to the free Dox from around 95-53% after 24h and from 63% to 37% after 48 h. Dox kinetic release from the BC-Alg nanocomposite displayed hyperbolic curves related to the different amounts of drug payload and was stable for at least 14 days. The results of the BC-Alg nanocomposites show a promissory potential for anticancer therapies of solid tumors. PMID:26784658

  1. Biosynthesis of Bacterial Cellulose/Carboxylic Multi-Walled Carbon Nanotubes for Enzymatic Biofuel Cell Application

    Directory of Open Access Journals (Sweden)

    Pengfei Lv

    2016-03-01

    Full Text Available Novel nanocomposites comprised of bacterial cellulose (BC with carboxylic multi-walled carbon nanotubes (c-MWCNTs incorporated into the BC matrix were prepared through a simple method of biosynthesis. The biocathode and bioanode for the enzyme biological fuel cell (EBFC were prepared using BC/c-MWCNTs composite injected by laccase (Lac and glucose oxidase (GOD with the aid of glutaraldehyde (GA crosslinking. Biosynthesis of BC/c-MWCNTs composite was characterized by digital photos, scanning electron microscope (SEM, and Fourier Transform Infrared (FTIR. The experimental results indicated the successful incorporation of c-MWCNTs into the BC. The electrochemical and biofuel performance were evaluated by cyclic voltammetry (CV and linear sweep voltammetry (LSV. The power density and current density of EBFCs were recorded at 32.98 µW/cm3 and 0.29 mA/cm3, respectively. Additionally, the EBFCs also showed acceptable stability. Preliminary tests on double cells indicated that renewable BC have great potential in the application field of EBFCs.

  2. Studies on interfacial interactions of TiO2 nanoparticles with bacterial cells under light and dark conditions

    Indian Academy of Sciences (India)

    Swayamprava Dalai; Sunandan Pakrashi; Sujay Chakravarty; Shamima Hussain; N Chandrasekaran; Amitava Mukherjee

    2014-05-01

    The probable underlying mechanism(s) of bacterial cell–TiO2 nanoparticles (TiO2 NPs) interaction in the absence of photo-irradiation has been less studied since most of the prior cytotoxicity studies focused on irradiated TiO2. The present study draws attention to the possible role of cell surface–TiO2 NP interactions under dark conditions, through an array of spectroscopic and microscopic investigations. A dominant freshwater bacterial isolate, Bacillus licheniformis, which interacted with environmentally relevant concentrations of TiO2 NPs (1 g/mL), was analysed and compared under both light and dark conditions. Aggregation of cells upon NP interaction and adsorption of NPs onto the cell membrane was evident from the scanning electron micrographs under both light and dark conditions. The FT–IR and FT–Raman spectra suggested stress response of bacterial cells by elevated protein and polysaccharide content in the cell–NP interaction. The X-ray photoelectron spectroscopic data substantiated the reduction of titanium from Ti(IV) to Ti(III) species which might have contributed to the redox interactions on the cell surface under light as well as dark conditions. The internalization of NPs in the cytoplasm were obvious from the transmission electron micrographs. The consequent cell death/damage was confirmed through fluorescence spectroscopy and microscopy. To conclude, the current study established the substantial role of interfacial interactions in cytotoxicity of the TiO2 NPs irrespective of the irradiation conditions.

  3. Interference of bacterial cell-to-cell communication: A new concept of antimicrobial chemotherapy breaks antibiotic

    Directory of Open Access Journals (Sweden)

    HidetadaHirakawa

    2013-05-01

    Full Text Available Bacteria use a cell-to-cell communication activity termed “Quorum sensing” to coordinate group behaviors in a cell-density dependent manner. Quorum sensing influences the expression profile of diverse genes, including antibiotic tolerance and virulence determinants, via specific chemical compounds called “Auto-inducers”. During quorum sensing, Gram-negative bacteria typically use an acylated homoserine lactone (AHL called auto-inducer 1 (AI-1. Since the first discovery of quorum sensing in a marine bacterium, it has been recognized that more than 100 species possess this mechanism of cell-to-cell communication. In addition to being of interest from a biological standpoint, quorum sensing is a potential target for antimicrobial chemotherapy. This unique concept of antimicrobial control relies on reducing the burden of virulence rather than killing the bacteria. It is believed that this approach will not only suppress the development of antibiotic resistance, but will also improve the treatment of refractory infections triggered by multi-drug resistant (MDR pathogens. In this paper, we review and track recent progress in studies on AHL inhibitors/modulators from a biological standpoint. It has been discovered that both natural and synthetic compounds can disrupt quorum sensing by a variety of means, such as jamming signal transduction, inhibition of signal production and break-down and trapping of signal compounds. We also focus on the regulatory elements that attenuate quorum sensing activities and discuss their unique properties. Understanding the biological roles of regulatory elements might be useful in developing inhibitor applications and understanding how quorum sensing is controlled.

  4. Activation of p38α in T cells regulates the intestinal host defense against attaching and effacing bacterial infections

    OpenAIRE

    Shim, Eun-Jin; Bang, Bo Ram; Kang, Seung-Goo; Ma, Jianhui; Otsuka, Motoyuki; Kang, Jiman; Stahl, Martin; Han, Jiahuai; Xiao, Changchun; Vallance, Bruce A.; Kang, Young Jun

    2013-01-01

    Intestinal infections by attaching and effacing (A/E) bacterial pathogens cause severe colitis and bloody diarrhea. Although p38α in intestine epithelial cells (IEC) plays an important role in promoting protection against A/E bacteria by regulating T cell recruitment, its impact on immune responses remains unclear. In this study, we show that activation of p38α in T cells is critical for the clearance of the A/E pathogen Citrobacter rodentium. Mice deficient of p38α in T cells, but not in mac...

  5. Categories with envelopes and imprints

    CERN Document Server

    Akbarov, Sergei

    2011-01-01

    An envelope in a category is a construction generalizing operations of "exterior completion", like completion of a locally convex space. Dually, an imprint generalizes operations of "interior enrichment", like saturation of a locally convex space. We give abstract definition for envelopes and imprints, prove existence of these objects in the categories of stereotype spaces and of stereotype algebras, and give some examples.

  6. Vigorous, but differential mononuclear cell response of cirrhotic patients to bacterial ligands

    Institute of Scientific and Technical Information of China (English)

    Varenka J Barbero-Becerra; María Concepción Gutiérrez-Ruiz; Carmen Maldonado-Bernal; Félix I Téllez-Avila; Roberto Alfaro-Lara; Florencia Vargas-Vorácková

    2011-01-01

    AIM: To study the role of gram-positive and gram-negative bacteria in the pathogenesis of liver injury, specifically the activation of inflammatory mediators. METHODS: Peripheral blood mononuclear cells of 20 out-patients were studied, 10 of them with cirrhosis. Peripheral blood mononuclear cells were isolated and exposed to lipopolysaccharide or lipoteichoic acid. CD14, Toll-like receptor 2 and 4 expression was determined by flow cytometry, and tumor necrosis factor (TNF) α, interleukin (IL)-1β, IL-6, IL-12 and IL-10 secretion in supernatants was determined by ELISA. RESULTS: Higher CD14, Toll-like receptor 2 and 4 expression was observed in peripheral blood mononuclear cells from cirrhotic patients, (P < 0.01, P < 0.006, P < 0.111) respectively. Lipopolysaccharide and lipoteichoic acid induced a further increase in CD14 expression (P < 0.111 lipopolysaccharide, P < 0.013 lipoteichoic acid), and a decrease in Toll-like receptor 2 (P < 0.008 lipopolysaccharide, P < 0.008 lipoteichoic acid) and Toll-like receptor 4 (P < 0.008 lipopolysaccharide, P < 0.028 lipoteichoic acid) expression. With the exception of TNFα, absolute cytokine secretion of peripheral blood mononuclear cells was lower in cirrhotic patients under nonexposure conditions (P < 0.070 IL-6, P < 0.009 IL-1β, P < 0.022 IL-12). Once exposed to lipopolysaccharide or lipoteichoic acid, absolute cytokine secretion of peripheral blood mononuclear cells was similar in cirrhotic and non-cirrhotic patients, determining a more vigorous response in the former (P < 0.005 TNFα, IL-1β, IL-6, IL-2 and IL-10 lipopolysaccharide; P < 0.037 TNFα; P < 0.006 IL-1β; P < 0.005 IL-6; P < 0.007 IL-12; P < 0.014 IL-10 lipoteichoic acid). Response of peripheral blood mononuclear cells was more intense after lipopolysaccharide than after lipoteichoic acid exposure. CONCLUSION: Peripheral blood mononuclear cells of cirrhotic patients are able to respond to a sudden bacterial ligand exposure, particularly lipopolysaccharide

  7. The nucleotide sequence of the high-leukemogenic murine retrovirus SL3-3 reveals a patch of mink cell focus forming-like sequences upstream of the ecotropic envelope gene. Brief report

    DEFF Research Database (Denmark)

    Lund, Anders Henrik; Pedersen, F S

    1999-01-01

    We report the complete nucleotide sequence of the potent T-lymphomagenic murine retrovirus SL3-3. The non-LTR regions of the virus show 98% sequence identity to the endogenous ecotropic Akv murine leukemia virus. While the region encoding the surface envelope protein is completely identical to that...... of Akv, a approximately 200 nucleotide stretch in the integrase encoding region upstream of env is similar to the sequence of mink cell focus forming (MCF) viruses and shows a complete match with the mouse retrovirus 10A1. The history of SL3-3 may therefore include recombination involving an Akv...

  8. Nonmalignant T cells stimulate growth of T-cell lymphoma cells in the presence of bacterial toxins

    DEFF Research Database (Denmark)

    Woetmann, Anders; Lovato, Paola; Eriksen, Karsten W; Krejsgaard, Thorbjørn; Labuda, Tord; Zhang, Qian; Mathiesen, Anne-Merethe; Geisler, Carsten; Svejgaard, Arne; Wasik, Mariusz A; Odum, Niels

    2007-01-01

    malignant CTCL cells express MHC class II molecules that are high-affinity receptors for SE. Although treatment with SE has no direct effect on the growth of the malignant CTCL cells, the SE-treated CTCL cells induce vigorous proliferation of the SE-responsive nonmalignant T cells. In turn, the nonmalignant......-promoting effect depends on direct cell-cell contact and soluble factors such as interleukin-2. In conclusion, we demonstrate that SE triggers a bidirectional cross talk between nonmalignant T cells and malignant CTCL cells that promotes growth of the malignant cells. This represents a novel mechanism by which...

  9. Bacterial β-(1,3)-glucan prevents DSS-induced IBD by restoring the reduced population of regulatory T cells.

    Science.gov (United States)

    Lee, Kwang-Ho; Park, Min; Ji, Kon-Young; Lee, Hwa-Youn; Jang, Ji-Hun; Yoon, Il-Joo; Oh, Seung-Su; Kim, Su-Man; Jeong, Yun-Hwa; Yun, Chul-Ho; Kim, Mi-Kyoung; Lee, In-Young; Choi, Ha-Rim; Ko, Ki-sung; Kang, Hyung-Sik

    2014-10-01

    Bacterial β-(1,3)-glucan has more advantages in terms of cost, yield and efficiency than that derived from mushrooms, plants, yeasts and fungi. We have previously developed a novel and high-yield β-(1,3)-glucan produced by Agrobacterium sp. R259. This study aimed to elucidate the functional mechanism and therapeutic efficacy of bacterial β-(1,3)-glucan in dextran sulfate sodium (DSS)-induced inflammatory bowel disease (IBD).Mice were orally pretreated with bacterial β-(1,3)-glucan at daily doses of 2.5 or 5mg/kg for 2 weeks. After 6 days of DSS treatment, clinical assessment of IBD severity and expression of pro-inflammatory cytokines were evaluated. In vivo cell proliferation was examined by immunohistochemistry using Ki-67 and ER-TR7 antibodies. The frequency of regulatory T cells (Tregs) was analyzed by flow cytometry. Natural killer (NK) activity and IgA level were evaluated using NK cytotoxicity assay and ELISA.The deterioration of body weight gain, colonic architecture, disease score and histological score was recovered in DSS-induced IBD mice when pretreated with bacterial β-(1,3)-glucan. The recruitment of macrophages and the gene expression of proinflammatory cytokines, such as IL-1β, IL-6 and IL-17A/F, were markedly decreased in the colon of β-(1,3)-glucan-pretreated mice. β-(1,3)-Glucan induced the recovery of Tregs in terms of their frequency in DSS-induced IBD mice. Intriguingly, β-(1,3)-glucan reversed the functional defects of NK cells and excessive IgA production in DSS-induced IBD mice.We conclude that bacterial β-(1,3)-glucan prevented the progression of DSS-induced IBD by recovering the reduction of Tregs, functional defect of NK cells and excessive IgA production. PMID:25092569

  10. Role of the T cell receptor ligand affinity in T cell activation by bacterial superantigens

    DEFF Research Database (Denmark)

    Andersen, P S; Geisler, C; Buus, S; Mariuzza, R A; Karjalainen, K

    2001-01-01

    the SEC3 variants correlated with enhanced binding without any optimum in the binding range covered by native TCR ligands. Comparable studies using anti-TCR antibodies of known affinity confirmed these observations. By comparing the biological potency of the two sets of ligands, we found a significant...... correlation between ligand affinity and ligand potency indicating that it is the density of receptor-ligand complexes in the T cell contact area that determines TCR signaling strength....

  11. In vitro behaviors of rat mesenchymal stem cells on bacterial celluloses with different moduli

    International Nuclear Information System (INIS)

    Compressive moduli of bacteria-synthesized cellulose (BC) were altered by two drying techniques: ambient-air drying and freeze drying. While no significant differences in dry weight were found, their cross-sectional structures and thickness varied greatly. Freeze dried BCs had loose cross-sectional structures and a thickness of ∼ 4.7 mm, whereas air dried BCs had more compacted cross-sectional structures and a thickness of ∼ 0.1 mm. The compressive moduli of the rehydrated freeze dried and rehydrated air dried BCs were measured to be 21.06 ± 0.22 kPa and 90.09 ± 21.07 kPa, respectively. When rat mesenchymal stem cells (rMSCs) were seeded on these BCs, they maintained a round morphology in the first 3 days of cultivation. More spread-out morphology and considerable proliferation on freeze dried BCs were observed in 7 days, but not on air-dried BCs. The cells were further grown for 3 weeks in the absence and presence of differentiation agents. Without using any differentiation agents, no detectable differentiation was noticed for rMSCs further cultivated on both types of BC. With differentiation inducing agents, chondrogenic differentiation, visualized by histological staining, was observed in some area of the rehydrated freeze dried BCs; while osteogenic differentiation was noticed on the stiffer rehydrated air dried BCs. - Graphical abstract: In the presence of induction agents, rat mesenchymal stem cells (rMSCs) preferentially differentiated into osteocytes on stiffer air dried BC films. - Highlights: • Bacterial cellulose (BC) sheets with different moduli generated by drying differently • Air-dried BC exhibited a modulus similar to that of bone. • Freeze-dried BC showed a modulus in the range of that of muscle. • Air-dried BC promoted the differentiation of rMSCs into osteocytes. • Freeze-dried BC promoted the differentiation of rMSCs into chondrocytes

  12. In vitro behaviors of rat mesenchymal stem cells on bacterial celluloses with different moduli

    Energy Technology Data Exchange (ETDEWEB)

    Taokaew, Siriporn [Department of Chemical Engineering, Faculty of Engineering, Chulalongkorn University, Bangkok 10330 (Thailand); Department of Chemical and Biomolecular Engineering, The University of Akron, Akron, OH 44325-3906 (United States); Phisalaphong, Muenduen [Department of Chemical Engineering, Faculty of Engineering, Chulalongkorn University, Bangkok 10330 (Thailand); Zhang Newby, Bi-min, E-mail: bimin@uakron.edu [Department of Chemical and Biomolecular Engineering, The University of Akron, Akron, OH 44325-3906 (United States)

    2014-05-01

    Compressive moduli of bacteria-synthesized cellulose (BC) were altered by two drying techniques: ambient-air drying and freeze drying. While no significant differences in dry weight were found, their cross-sectional structures and thickness varied greatly. Freeze dried BCs had loose cross-sectional structures and a thickness of ∼ 4.7 mm, whereas air dried BCs had more compacted cross-sectional structures and a thickness of ∼ 0.1 mm. The compressive moduli of the rehydrated freeze dried and rehydrated air dried BCs were measured to be 21.06 ± 0.22 kPa and 90.09 ± 21.07 kPa, respectively. When rat mesenchymal stem cells (rMSCs) were seeded on these BCs, they maintained a round morphology in the first 3 days of cultivation. More spread-out morphology and considerable proliferation on freeze dried BCs were observed in 7 days, but not on air-dried BCs. The cells were further grown for 3 weeks in the absence and presence of differentiation agents. Without using any differentiation agents, no detectable differentiation was noticed for rMSCs further cultivated on both types of BC. With differentiation inducing agents, chondrogenic differentiation, visualized by histological staining, was observed in some area of the rehydrated freeze dried BCs; while osteogenic differentiation was noticed on the stiffer rehydrated air dried BCs. - Graphical abstract: In the presence of induction agents, rat mesenchymal stem cells (rMSCs) preferentially differentiated into osteocytes on stiffer air dried BC films. - Highlights: • Bacterial cellulose (BC) sheets with different moduli generated by drying differently • Air-dried BC exhibited a modulus similar to that of bone. • Freeze-dried BC showed a modulus in the range of that of muscle. • Air-dried BC promoted the differentiation of rMSCs into osteocytes. • Freeze-dried BC promoted the differentiation of rMSCs into chondrocytes.

  13. Bacterial biofilm mechanical properties persist upon antibiotic treatment and survive cell death

    International Nuclear Information System (INIS)

    Bacteria living on surfaces form heterogeneous three-dimensional consortia known as biofilms, where they exhibit many specific properties one of which is an increased tolerance to antibiotics. Biofilms are maintained by a polymeric network and display physical properties similar to that of complex fluids. In this work, we address the question of the impact of antibiotic treatment on the physical properties of biofilms based on recently developed tools enabling the in situ mapping of biofilm local mechanical properties at the micron scale. This approach takes into account the material heterogeneity and reveals the spatial distribution of all the small changes that may occur in the structure. With an Escherichia coli biofilm, we demonstrate using in situ fluorescent labeling that the two antibiotics ofloxacin and ticarcillin—targeting DNA replication and membrane assembly, respectively—induced no detectable alteration of the biofilm mechanical properties while they killed the vast majority of the cells. In parallel, we show that a proteolytic enzyme that cleaves extracellular proteins into short peptides, but does not alter bacterial viability in the biofilm, clearly affects the mechanical properties of the biofilm structure, inducing a significant increase of the material compliance. We conclude that conventional biofilm control strategy relying on the use of biocides targeting cells is missing a key target since biofilm structural integrity is preserved. This is expected to efficiently promote biofilm resilience, especially in the presence of persister cells. In contrast, the targeting of polymer network cross-links—among which extracellular proteins emerge as major players—offers a promising route for the development of rational multi-target strategies to fight against biofilms. (paper)

  14. Efficiency of fluorescence in situ hybridization for bacterial cell identification in temporary river sediments with contrasting water content.

    Science.gov (United States)

    Fazi, Stefano; Amalfitano, Stefano; Pizzetti, Ilaria; Pernthaler, Jakob

    2007-09-01

    We studied the efficiency of two hybridization techniques for the analysis of benthic bacterial community composition under varying sediment water content. Microcosms were set up with sediments from four European temporary rivers. Wet sediments were dried, and dry sediments were artificially rewetted. The percentage of bacterial cells detected by fluorescence in situ hybridization with fluorescently monolabeled probes (FISH) significantly increased from dry to wet sediments, showing a positive correlation with the community activity measured via incorporation of (3)H leucine. FISH and signal amplification by catalyzed reporter deposition (CARD-FISH) could significantly better detect cells with low activity in dried sediments. Through the application of an optimized cell permeabilization protocol, the percentage of hybridized cells by CARD-FISH showed comparable values in dry and wet conditions. This approach was unrelated to (3)H leucine incorporation rates. Moreover, the optimized protocol allowed a significantly better visualization of Gram-positive Actinobacteria in the studied samples. CARD-FISH is, therefore, proposed as an effective technique to compare bacterial communities residing in sediments with contrasting water content, irrespective of differences in the activity state of target cells. Considering the increasing frequencies of flood and drought cycles in European temporary rivers, our approach may help to better understand the dynamics of microbial communities in such systems. PMID:17452089

  15. Bacterial carbonatogenesis

    International Nuclear Information System (INIS)

    Several series of experiments in the laboratory as well as in natural conditions teach that the production of carbonate particles by heterotrophic bacteria follows different ways. The 'passive' carbonatogenesis is generated by modifications of the medium that lead to the accumulation of carbonate and bicarbonate ions and to the precipitation of solid particles. The 'active' carbonatogenesis is independent of the metabolic pathways. The carbonate particles are produced by ionic exchanges through the cell membrane following still poorly known mechanisms. Carbonatogenesis appears to be the response of heterotrophic bacterial communities to an enrichment of the milieu in organic matter. The active carbonatogenesis seems to start first. It is followed by the passive one which induces the growth of initially produced particles. The yield of heterotrophic bacterial carbonatogenesis and the amounts of solid carbonates production by bacteria are potentially very high as compared to autotrophic or chemical sedimentation from marine, paralic or continental waters. Furthermore, the bacterial processes are environmentally very ubiquitous; they just require organic matter enrichment. Thus, apart from purely evaporite and autotrophic ones, all Ca and/or Mg carbonates must be considered as from heterotrophic bacterial origin. By the way, the carbon of carbonates comes from primary organic matter. Such considerations ask questions about some interpretations from isotopic data on carbonates. Finally, bacterial heterotrophic carbonatogenesis appears as a fundamental phase in the relationships between atmosphere and lithosphere and in the geo-biological evolution of Earth. (author)

  16. Bacterial gastroenteritis

    Science.gov (United States)

    Infectious diarrhea - bacterial gastroenteritis; Acute gastroenteritis; Gastroenteritis - bacterial ... Bacterial gastroenteritis can affect 1 person or a group of people who all ate the same food. It is ...

  17. A novel functional T cell hybridoma recognizes macrophage cell death induced by bacteria: a possible role for innate lymphocytes in bacterial infection.

    Science.gov (United States)

    Kubota, Koichi

    2006-06-15

    We have established a novel TCRalphabeta (TCRVbeta6)(+)CD4(-)CD8(-) T cell hybridoma designated B6HO3. When the B6HO3 cells were cocultured with bacterial-infected J774 macrophage-like cells, IFN-gamma production by B6HO3 cells was triggered through direct cell-cell contact with dying J774 cells infected with Listeria monocytogenes (LM), Shigella flexneri, or Salmonella typhimurium that expressed the type III secretion system, but not with intact J774 cells infected with heat-killed LM, nonhemolytic lysteriolysin O-deficient (Hly(-)) LM, plasmid-cured Shigella, or stationary-phase Salmonella. However, the triggering of B6HO3 cells for IFN-gamma production involved neither dying hepatoma cells infected with LM nor dying J774 cells caused by gliotoxin treatment or freeze thawing. Cycloheximide and Abs to H-2K(d), H-2D(d), Ia(d), CD1d, TCRVbeta6, and IL-12 did not inhibit the contact-dependent IFN-gamma response, indicating that this IFN-gamma response did not require de novo protein synthesis in bacterial-infected J774 cells and was TCR and IL-12 independent. Thus, in an as yet undefined way, B6HO3 hybridoma recognizes a specialized form of macrophage cell death resulting from bacterial infection and consequently produces IFN-gamma. Moreover, contact-dependent interaction of minor subsets of splenic alphabeta T cells, including NKT cells with dying LM-infected J774 and bone marrow-derived macrophage (BMM) cells, proved to provide an IFN-gamma-productive stimulus for these minor T cell populations, to which the parental T cell of the B6HO3 hybridoma appeared to belong. Unexpectedly, subsets of gammadelta T and NK cells similarly responded to dying LM-infected macrophage cells. These results propose that innate lymphocytes may possess a recognition system sensing macrophage cell "danger" resulting from bacterial infection. PMID:16751404

  18. An ethanol biosensor based on a bacterial cell-immobilized eggshell membrane

    Institute of Scientific and Technical Information of China (English)

    Guang Ming Wen; Shao Min Shuang; Chuan Dong; Martin M.F. Choi

    2012-01-01

    An ethanol biosensor was fabricated based on a Methylobacterium organophilium-immobilized eggshell membrane and an oxygen (O2) electrode.A linear response for ethanol was obtained in the range of 0.050-7.5 mmol/L with a detection limit of 0.025 mmol/L (S/N =3) and a R.S.D.of 2.1%.The response time was less than 100 s at room temperature and ambient pressure.The optimal loading of bacterial cells on the biosensor membrane is 40 mg (wet weight).The optimal working conditions for the microbial biosensor are pH 7.0 phosphate buffer (50 mmol/L) at 20-25 ℃.The interference test,operational and storage stability of the biosensor are studied in detail.Finally,the biosensor is applied to determine the ethanol contents in various alcohol samples and the results are comparable to that obtained by gas chromatographic method and the results are satisfactory.Our proposed biosensor provides a convenient,simple and reliable method to determine ethanol content in alcoholic drinks.

  19. Volumetric measurements of bacterial cells and extracellular polymeric substance glycoconjugates in biofilms.

    Science.gov (United States)

    Staudt, C; Horn, H; Hempel, D C; Neu, T R

    2004-12-01

    In this study an enrichment culture developed from activated sludge was used to investigate the architecture of fully hydrated multispecies biofilms. The assessment of biofilm structure and volume was carried out using confocal laser scanning microscopy (CLSM). Bacterial cell distribution was determined with the nucleic acid-specific stain SYTO 60, whereas glycoconjugates of extracellular polymeric substances (EPS) were stained with the Alexa-488-labeled lectin of Aleuria aurantia. Digital image analysis was employed for visualization and quantification of three-dimensional CLSM data sets. The specific volumes of the polymeric and cellular biofilm constituents were quantified. In addition, gravimetric measurements were done to determine dry mass and thickness of the biofilms. The data recorded by the CLSM technique and the gravimetric data were then compared. It was shown that the biofilm thicknesses determined with both methods agree well for slow-growing heterotrophic and chemoautotrophic biofilms. In addition, for slow-growing biofilms, the volumes and masses calculated from CLSM and the biomass calculated from gravimetric measurements were also comparable. For fast-growing heterotrophic biofilms cultivated with high glucose concentrations the data sets fit to a lesser degree, but still showed the same common trend. Compared with traditional gravimetric measurements, CLSM allowed differential recording of multiple biofilm parameters with subsequent three-dimensional visualization and quantification. The quantitative three-dimensional results recorded by CLSM are an important basis for understanding, controlling, exploiting, and modeling of biofilms. PMID:15470707

  20. Vimentin in Bacterial Infections

    DEFF Research Database (Denmark)

    Mak, Tim N; Brüggemann, Holger

    2016-01-01

    -vimentin interactions are presented in this review: the role of vimentin in pathogen-binding on the cell surface and subsequent bacterial invasion and the interaction of cytosolic vimentin and intracellular pathogens with regards to innate immune signaling. Mechanistic insight is presented involving distinct bacterial......Despite well-studied bacterial strategies to target actin to subvert the host cell cytoskeleton, thus promoting bacterial survival, replication, and dissemination, relatively little is known about the bacterial interaction with other components of the host cell cytoskeleton, including intermediate...... filaments (IFs). IFs have not only roles in maintaining the structural integrity of the cell, but they are also involved in many cellular processes including cell adhesion, immune signaling, and autophagy, processes that are important in the context of bacterial infections. Here, we summarize the knowledge...

  1. Multifamily Envelope Leakage Model

    Energy Technology Data Exchange (ETDEWEB)

    Faakye, Omari [Consortium for Advanced Residential Buildings, Norwalk, CT (United States); Griffiths, Dianne [Consortium for Advanced Residential Buildings, Norwalk, CT (United States)

    2015-05-08

    “The cost for blower testing is high, because it is labor intensive, and it may disrupt occupants in multiple units. This high cost and disruption deter program participants, and dissuade them from pursuing energy improvements that would trigger air leakage testing, such as improvements to the building envelope.” This statement found in a 2012 report by Heschong Mahone Group for several California interests emphasizes the importance of reducing the cost and complexity of blower testing in multifamily buildings. Energy efficiency opportunities are being bypassed. The cost of single blower testing is on the order of $300. The cost for guarded blower door testing—the more appropriate test for assessing energy savings opportunities—could easily be six times that, and that’s only if you have the equipment and simultaneous access to multiple apartments. Thus, the proper test is simply not performed. This research seeks to provide an algorithm for predicting the guarded blower door test result based upon a single, total blower door test.

  2. Impact of bacteria and bacterial components on osteogenic and adipogenic differentiation of adipose-derived mesenchymal stem cells

    International Nuclear Information System (INIS)

    Adult mesenchymal stem cells (MSC) are present in several tissues, e.g. bone marrow, heart muscle, brain and subcutaneous adipose tissue. In invasive infections MSC get in contact with bacteria and bacterial components. Not much is known about how bacterial pathogens interact with MSC and how contact to bacteria influences MSC viability and differentiation potential. In this study we investigated the impact of three different wound infection relevant bacteria, Escherichia coli, Staphylococcus aureus, and Streptococcus pyogenes, and the cell wall components lipopolysaccharide (LPS; Gram-negative bacteria) and lipoteichoic acid (LTA; Gram-positive bacteria) on viability, proliferation, and osteogenic as well as adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells (adMSC). We show that all three tested species were able to attach to and internalize into adMSC. The heat-inactivated Gram-negative E. coli as well as LPS were able to induce proliferation and osteogenic differentiation but reduce adipogenic differentiation of adMSC. Conspicuously, the heat-inactivated Gram-positive species showed the same effects on proliferation and adipogenic differentiation, while its cell wall component LTA exhibited no significant impact on adMSC. Therefore, our data demonstrate that osteogenic and adipogenic differentiation of adMSC is influenced in an oppositional fashion by bacterial antigens and that MSC-governed regeneration is not necessarily reduced under infectious conditions. - Highlights: • Staphylococcus aureus, Streptococcus pyogenes and Escherichia coli bind to and internalize into adMSC. • Heat-inactivated cells of these bacterial species trigger proliferation of adMSC. • Heat-inactivated E. coli and LPS induce osteogenic differentiation of adMSC. • Heat-inactivated E. coli and LPS reduce adipogenic differentiation of adMSC. • LTA does not influence adipogenic or osteogenic differentiation of adMSC

  3. Impact of bacteria and bacterial components on osteogenic and adipogenic differentiation of adipose-derived mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Fiedler, Tomas, E-mail: tomas.fiedler@med.uni-rostock.de [Institute for Medical Microbiology, Virology, and Hygiene, Rostock University Medical Center, Schillingallee 70, D-18057 Rostock (Germany); Salamon, Achim; Adam, Stefanie; Herzmann, Nicole [Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany); Taubenheim, Jan [Institute for Medical Microbiology, Virology, and Hygiene, Rostock University Medical Center, Schillingallee 70, D-18057 Rostock (Germany); Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany); Peters, Kirsten [Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany)

    2013-11-01

    Adult mesenchymal stem cells (MSC) are present in several tissues, e.g. bone marrow, heart muscle, brain and subcutaneous adipose tissue. In invasive infections MSC get in contact with bacteria and bacterial components. Not much is known about how bacterial pathogens interact with MSC and how contact to bacteria influences MSC viability and differentiation potential. In this study we investigated the impact of three different wound infection relevant bacteria, Escherichia coli, Staphylococcus aureus, and Streptococcus pyogenes, and the cell wall components lipopolysaccharide (LPS; Gram-negative bacteria) and lipoteichoic acid (LTA; Gram-positive bacteria) on viability, proliferation, and osteogenic as well as adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells (adMSC). We show that all three tested species were able to attach to and internalize into adMSC. The heat-inactivated Gram-negative E. coli as well as LPS were able to induce proliferation and osteogenic differentiation but reduce adipogenic differentiation of adMSC. Conspicuously, the heat-inactivated Gram-positive species showed the same effects on proliferation and adipogenic differentiation, while its cell wall component LTA exhibited no significant impact on adMSC. Therefore, our data demonstrate that osteogenic and adipogenic differentiation of adMSC is influenced in an oppositional fashion by bacterial antigens and that MSC-governed regeneration is not necessarily reduced under infectious conditions. - Highlights: • Staphylococcus aureus, Streptococcus pyogenes and Escherichia coli bind to and internalize into adMSC. • Heat-inactivated cells of these bacterial species trigger proliferation of adMSC. • Heat-inactivated E. coli and LPS induce osteogenic differentiation of adMSC. • Heat-inactivated E. coli and LPS reduce adipogenic differentiation of adMSC. • LTA does not influence adipogenic or osteogenic differentiation of adMSC.

  4. N-acetylcysteine and the human serum components that inhibit bacterial invasion of gingival epithelial cells prevent experimental periodontitis in mice

    OpenAIRE

    Alam, Jehan; Baek, Keum Jin; Choi, Yun Sik; Kim, Yong Cheol; Choi, Youngnim

    2014-01-01

    Purpose We previously reported that human serum significantly reduces the invasion of various oral bacterial species into gingival epithelial cells in vitro. The aims of the present study were to characterize the serum component(s) responsible for the inhibition of bacterial invasion of epithelial cells and to examine their effect on periodontitis induced in mice. Methods Immortalized human gingival epithelial (HOK-16B) cells were infected with various 5- (and 6-) carboxy-fluorescein diacetat...

  5. Origin of envelope proteins of a leukemia virus

    International Nuclear Information System (INIS)

    The roles of avian myeloblastosis virus (AMV) and host myeloblast cells in controlling the protein composition of virus envelope and host cell membrane are being studied by examining an ATPase enzyme in the virus and cells. New culture techniques for virus producing myeloblasts have been developed. (U.S.)

  6. Bacterial membrane activity of a-peptide/b-peptoid chimeras: Influence of amino acid composition and chain length on the activity against different bacterial strains

    DEFF Research Database (Denmark)

    Hein-Kristensen, Line; Knapp, Kolja M; Franzyk, Henrik;

    2011-01-01

    BACKGROUND: Characterization and use of antimicrobial peptides (AMPs) requires that their mode of action is determined. The interaction of membrane-active peptides with their target is often established using model membranes, however, the actual permeabilization of live bacterial cells and...... permeabilization of the bacterial cell envelope, and the outer membrane may act as a barrier in Gram-negative bacteria. The tolerance of S. marcescens to chimeras may be due to differences in the composition of the lipopolysaccharide layer also responsible for its resistance to polymyxin B....... subsequent killing is usually not tested. In this report, six α-peptide/β-peptoid chimeras were examined for the effect of amino acid/peptoid substitutions and chain length on the membrane perturbation and subsequent killing of food-borne and clinical bacterial isolates. RESULTS: All six AMP analogues...

  7. Safe operating envelope

    International Nuclear Information System (INIS)

    This is a slide-based oral presentation given to the COG/IAEA: Fifth technical committee meeting on 'Exchange of operating experience of pressurized heavy water reactors' held in Mangalia, Romania on 7-10 September 1998. The plant states and operating conditions are defined as resulting from the consequences of a Licensing Basis Event occurrence. Three categories of important plant parameters are considered: A - directly associated with either the mechanical or instrumentation and control aspects of the Special Safety Systems; B - process conditions related to trip parameters; C - other parameters that effect outcome of an accident. In A category the following aspects were addressed: Instrument loop uncertainties and time lags and delays; Tank levels, pressures and chemistry; Valve operating times and characteristics; Pump performance; The number of redundant pieces of equipment. Relating to category B the process conditions implied in trip initiation parameters as they effect margin to trip are discussed and illustrated by the cases of pressurizer level, RB pressure, etc. Finally, in the last category, the parameters effecting accident outcome are considered, i.e. either process variables, or equipment associated with safety related or safety support systems. The following cases are analysed PHTS Isotopic/temperatures/flows, moderator outlet temperature, RCW/RSW temperature, number of available aux. boiler feedpumps. In connection with the plant states the following matters are analyzed: fueling; boiler tube leak; defect fuel; non-equilibrium core; shutdown and reduced power operation; two pump operation. Concerning the Safety Operating Envelope (SOE) the following issues are presented: OP and P (high level overview), supported by OM tests and surveillance, by taking into account the use Tech Specs for the future, the role of safety analysis, historical perspective at PLGS. Finally, the DOA (Design/Operation/Analysis) Program at PLGS is described and the following

  8. Phenotypic T Cell Exhaustion in a Murine Model of Bacterial Infection in the Setting of Pre-Existing Malignancy

    OpenAIRE

    Mittal, Rohit; Wagener, Maylene; Breed, Elise R.; Liang, Zhe; Yoseph, Benyam P.; Burd, Eileen M.; Farris, Alton B.; Coopersmith, Craig M; Ford, Mandy L.

    2014-01-01

    While much of cancer immunology research has focused on anti-tumor immunity both systemically and within the tumor microenvironment, little is known about the impact of pre-existing malignancy on pathogen-specific immune responses. Here, we sought to characterize the antigen-specific CD8+ T cell response following a bacterial infection in the setting of pre-existing pancreatic adenocarcinoma. Mice with established subcutaneous pancreatic adenocarcinomas were infected with Listeria monocytogen...

  9. Effect of bacterial lectin on acceleration of fat cell lipolysis at in vitro diode laser treatment using encapsulated ICG

    Science.gov (United States)

    Yanina, Irina Yu.; Kochubey, Vyacheslav I.; Tuchin, Valery V.; Portnov, Sergey A.; Svenskaya, Yuliya I.; Gorin, Dmitry A.; Ponomareva, Elena G.; Nikitina, Valentina E.

    2012-03-01

    The influence of bacterial lectin on photochemically induced fat cell lipolysis was studied. Resulting capsules were tested for ICG absorption by optical spectra measurements. To separate released and encapsulated ICG supernatant was removed and capsules were redispered in pure deionized water. Supernatant and capsule suspension spectra were measured separately. It was also found that pretreatment of tissue by lectin leads to acceleration of lipolysis at photochemical treatment. The data obtained can be used to enhance efficiency of photochemical therapy.

  10. Radioprotection of mice by the bacterial extract Broncho-VaxonR: haemopoietic stem cells and survival enhancement

    International Nuclear Information System (INIS)

    Pretreatment of mice with 50-1000 μg of the bacterial extract Broncho-VaxomR (BV, free of endotoxin) before sublethal irradiation induced an increase in the number of endogenous haemopoietic stem cells (E-CFU). The degree of radioprotection was dependent on both the time of administration and the dose of BV. An optimal E-CFU survival was observed when 500 μg of BV was administered i.p. 24h before irradiation. (author)

  11. CD4+ T Cells and Toll-Like Receptors Recognize Salmonella Antigens Expressed in Bacterial Surface Organelles

    OpenAIRE

    Bergman, Molly A.; Cummings, Lisa A.; Barrett, Sara L. Rassoulian; Smith, Kelly D.; Lara, J. Cano; Aderem, Alan; Cookson, Brad T.

    2005-01-01

    A better understanding of immunity to infection is revealed from the characteristics of microbial ligands recognized by host immune responses. Murine infection with the intracellular bacterium Salmonella generates CD4+ T cells that specifically recognize Salmonella proteins expressed in bacterial surface organelles such as flagella and membrane vesicles. These natural Salmonella antigens are also ligands for Toll-like receptors (TLRs) or avidly associated with TLR ligands such as lipopolysacc...

  12. Identification of multiple physicochemical and structural properties associated with soluble expression of eukaryotic proteins in cell-free bacterial extracts

    OpenAIRE

    AlexanderA.Tokmakov

    2014-01-01

    Bacterial extracts are widely used to synthesize recombinant proteins. Vast data volumes have been accumulated in cell-free expression databases, covering a whole range of existing proteins. It makes possible comprehensive bioinformatics analysis and identification of multiple features associated with protein solubility and aggregation. In the present paper, an approach to identify the multiple physicochemical and structural properties of amino acid sequences associated with soluble expressio...

  13. The LHC in an envelope

    CERN Multimedia

    2007-01-01

    The series of envelopes featuring CERN issued this summer was a huge success. The French postal services of the Pays de Gex will shortly be launching the second set of pre-paid envelopes issued in collaboration with the Laboratory this year, this time highlighting the LHC. Five thousand envelopes describing the accelerator’s capabilities will go on sale on 12 November, and some of the packs will even contain a small sample of the cables from the heart of the LHC magnets. The sets of ten pre-paid envelopes will tell you everything about CERN’s flagship accelerator, from its astounding technical capabilities to its spin-offs in the fields of technology and human resources. Each envelope will feature a different attribute or spin-off of the LHC. People will be invited to consult CERN’s public website for more detailed explanations if they want to know more. The new envelopes will be available from five post offices in the Pays de Gex (Ferney-Voltaire, Prévessin...

  14. The LHC on an envelope

    CERN Multimedia

    2007-01-01

    The series of envelopes featuring CERN issued this summer was a huge success. The French postal services of the Pays de Gex will shortly be launching the second set of pre-paid envelopes issued in collaboration with the Laboratory this year, this time highlighting the LHC. Five thousand envelopes describing the accelerator’s capabilities will go on sale on 12 November, and some of the packs will even contain a small sample of the cables from the heart of the LHC magnets. The sets of ten pre-paid envelopes will tell you everything about CERN’s flagship accelerator, from its astounding technical capabilities to its spin-offs in the fields of technology and human resources. Each envelope will feature a different attribute or spin-off of the LHC. People will be invited to consult CERN’s public website for more detailed explanations if they want to know more. The new envelopes will be available from five post offices in the Pays ...

  15. Yeast cell wall extract induces disease resistance against bacterial and fungal pathogens in Arabidopsis thaliana and Brassica crop.

    Science.gov (United States)

    Narusaka, Mari; Minami, Taichi; Iwabuchi, Chikako; Hamasaki, Takashi; Takasaki, Satoko; Kawamura, Kimito; Narusaka, Yoshihiro

    2015-01-01

    Housaku Monogatari (HM) is a plant activator prepared from a yeast cell wall extract. We examined the efficacy of HM application and observed that HM treatment increased the resistance of Arabidopsis thaliana and Brassica rapa leaves to bacterial and fungal infections. HM reduced the severity of bacterial leaf spot and anthracnose on A. thaliana and Brassica crop leaves with protective effects. In addition, gene expression analysis of A. thaliana plants after treatment with HM indicated increased expression of several plant defense-related genes. HM treatment appears to induce early activation of jasmonate/ethylene and late activation of salicylic acid (SA) pathways. Analysis using signaling mutants revealed that HM required SA accumulation and SA signaling to facilitate resistance to the bacterial pathogen Pseudomonas syringae pv. maculicola and the fungal pathogen Colletotrichum higginsianum. In addition, HM-induced resistance conferred chitin-independent disease resistance to bacterial pathogens in A. thaliana. These results suggest that HM contains multiple microbe-associated molecular patterns that activate defense responses in plants. These findings suggest that the application of HM is a useful tool that may facilitate new disease control methods. PMID:25565273

  16. Yeast cell wall extract induces disease resistance against bacterial and fungal pathogens in Arabidopsis thaliana and Brassica crop.

    Directory of Open Access Journals (Sweden)

    Mari Narusaka

    Full Text Available Housaku Monogatari (HM is a plant activator prepared from a yeast cell wall extract. We examined the efficacy of HM application and observed that HM treatment increased the resistance of Arabidopsis thaliana and Brassica rapa leaves to bacterial and fungal infections. HM reduced the severity of bacterial leaf spot and anthracnose on A. thaliana and Brassica crop leaves with protective effects. In addition, gene expression analysis of A. thaliana plants after treatment with HM indicated increased expression of several plant defense-related genes. HM treatment appears to induce early activation of jasmonate/ethylene and late activation of salicylic acid (SA pathways. Analysis using signaling mutants revealed that HM required SA accumulation and SA signaling to facilitate resistance to the bacterial pathogen Pseudomonas syringae pv. maculicola and the fungal pathogen Colletotrichum higginsianum. In addition, HM-induced resistance conferred chitin-independent disease resistance to bacterial pathogens in A. thaliana. These results suggest that HM contains multiple microbe-associated molecular patterns that activate defense responses in plants. These findings suggest that the application of HM is a useful tool that may facilitate new disease control methods.

  17. Cell-to-Cell Propagation of the Bacterial Toxin CNF1 via Extracellular Vesicles: Potential Impact on the Therapeutic Use of the Toxin

    Directory of Open Access Journals (Sweden)

    Alessia Fabbri

    2015-11-01

    Full Text Available Eukaryotic cells secrete extracellular vesicles (EVs, either constitutively or in a regulated manner, which represent an important mode of intercellular communication. EVs serve as vehicles for transfer between cells of membrane and cytosolic proteins, lipids and RNA. Furthermore, certain bacterial protein toxins, or possibly their derived messages, can be transferred cell to cell via EVs. We have herein demonstrated that eukaryotic EVs represent an additional route of cell-to-cell propagation for the Escherichia coli protein toxin cytotoxic necrotizing factor 1 (CNF1. Our results prove that EVs from CNF1 pre-infected epithelial cells can induce cytoskeleton changes, Rac1 and NF-κB activation comparable to that triggered by CNF1. The observation that the toxin is detectable inside EVs derived from CNF1-intoxicated cells strongly supports the hypothesis that extracellular vesicles can offer to the toxin a novel route to travel from cell to cell. Since anthrax and tetanus toxins have also been reported to engage in the same process, we can hypothesize that EVs represent a common mechanism exploited by bacterial toxins to enhance their pathogenicity.

  18. Prophylaxis with levofloxacin: impact on bacterial susceptibility and epidemiology in a hematopoietic stem cell transplant unit

    Directory of Open Access Journals (Sweden)

    Livia Amaral Alonso Lopes

    2014-01-01

    Full Text Available Background: The emergence of resistance has been demonstrated in cancer treatment centers where prophylaxis with fluoroquinolone is used. Objective: Considering the importance of epidemiological monitoring as a strategy in choosing protocols involving antibiotics, this study aimed to evaluate the emergence of quinolone resistance and changes in the local epidemiology in a hematopoietic stem cell transplant service. Methods: For this study, 60 positive cultures before the prophylactic use of levofloxacin (period A: 2007-2008 and 118 cultures after starting the use of prophylactic levofloxacin (period B: 2010-2011 were evaluated. Results: Resistance increased for all the different types of bacteria isolated (from 46.0% to 76.5%; p-value = 0.0002. Among Gram-negative bacteria, resistance increased from 21.4% to 60.7% (p-value = 0.0163 and among Gram-positive bacteria, it increased from 55.6% to 82.9% (p-value = 0.0025. The use of levofloxacin increased from 19.44 defined daily doses per 1,000 patient-days in period A to 166.64 in period B. The use of broad spectrum antibiotics remained unchanged. Considering bacteria associated with infection, 72 and 76 were isolated in periods A and B, respectively. There was a reduction in the rate of Gramnegative bacteria in cultures associated with infection (3.81 vs. 2.00 cultures/1,000 patientdays; p-value = 0.008. Conclusion: The study of prophylaxis with levofloxacin demonstrated that there was a decrease in infections by Gram-negative bacteria; however, bacterial resistance increased, even though the use of broad-spectrum antibiotics remained unchanged. Constant monitoring of local epidemiology combined with research on clinical outcomes is needed to evaluate the effectiveness of prophylaxis.

  19. Bacterioplankton Production in Humic Lake Örträsket in Relation to Input of Bacterial Cells and Input of Allochthonous Organic Carbon.

    Science.gov (United States)

    Bergström; Jansson

    2000-02-01

    In order to compare riverine bacteria input with lake water bacterial production and grazing loss with output loss, a bacterial cell budget was constructed for humic Lake Örträsket in northern Sweden. The riverine input of bacterial cells in 1997 represented 29% of the number of bacterial cells produced within the layer of the lake affected by inlet water. A large share of the in situ lake bacterial production was consumed by grazers, mainly flagellates, which stresses the importance of bacteria as energy mobilizers for the pelagic food web in the lake. The bacterial production in Lake Örträsket, which is almost entirely dependent on humic material as an energy source, was clearly stimulated by high flow episodes which brought high amounts of little degraded material into the lake. During base flow condition the bacterial production in the inlet rivers was high, which led to an input of more degraded material to the lake. This material did not stimulate the lake bacterial production. Internal factors that determined the utilization of the allochthonous DOC in the lake were the retention time and the exposure to light and high temperatures. Thus, the potential for in situ production of bacteria in Lake Örträsket was to a large extent a function of how precipitation and runoff conditions affected terrestrial losses and river transport of humic material. PMID:10833223

  20. Short-term variability in bacterial abundance, cell properties, and incorporation of leucine and thymidine in subarctic sea ice

    DEFF Research Database (Denmark)

    Kaartokallio, H.; Søgaard, D.H.; Norman, L.;

    2013-01-01

    cell population properties (by flow cytometry) in subarctic sea ice in SW Greenland. Short-term temporal variability was moderate, and steep environmental gradients, typical for sea ice, were the main drivers of the variability in bacterial cell properties and activity. Low nucleic acid (LNA) bacteria...... brightly fluorescing intracellular inclusions after Nile Blue A staining. High Leu saturating concentrations coupled with the occurrence of PHA-producing organisms further highlight the similarity of sea ice internal habitats to biofilm-like systems rather than to open-water systems....

  1. Pili Binding to Asialo-GM1 on Epithelial Cells Can Mediate Cytotoxicity or Bacterial Internalization by Pseudomonas aeruginosa

    OpenAIRE

    Comolli, James C; Waite, Leslie L.; Keith E Mostov; Joanne N. Engel

    1999-01-01

    The interaction of Pseudomonas aeruginosa type IV pili and the glycosphingolipid asialo-GM1 (aGM1) can mediate bacterial adherence to epithelial cells, but the steps subsequent to this adherence have not been elucidated. To investigate the result of the interaction of pili and aGM1, we used polarized epithelial monolayers of Madin-Darby canine kidney (MDCK) cells in culture, which contained little detectable aGM1 on their apical surface but were able to incorporate exogenous aGM1. Compared to...

  2. A LytM Domain Dictates the Localization of Proteins to the Mother Cell-Forespore Interface during Bacterial Endospore Formation▿ †

    OpenAIRE

    Meisner, Jeffrey; Moran, Charles P.

    2010-01-01

    A large number of proteins are known to reside at specific subcellular locations in bacterial cells. However, the molecular mechanisms by which many of these proteins are anchored at these locations remains unclear. During endospore formation in Bacillus subtilis, several integral membrane proteins are located specifically at the interface of the two adjacent cells of the developing sporangium, the mother cell and forespore. The mother cell membrane protein SpoIIIAH recognizes the cell-cell i...

  3. The envelope of mycobacteria.

    Science.gov (United States)

    Brennan, P J; Nikaido, H

    1995-01-01

    Mycobacteria, members of which cause tuberculosis and leprosy, produce cell walls of unusually low permeability, which contribute to their resistance to therapeutic agents. Their cell walls contain large amounts of C60-C90 fatty acids, mycolic acids, that are covalently linked to arabinogalactan. Recent studies clarified the unusual structures of arabinogalactan as well as of extractable cell wall lipids, such as trehalose-based lipooligosaccharides, phenolic glycolipids, and glycopeptidolipids. Most of the hydrocarbon chains of these lipids assemble to produce an asymmetric bilayer of exceptional thickness. Structural considerations suggest that the fluidity is exceptionally low in the innermost part of bilayer, gradually increasing toward the outer surface. Differences in mycolic acid structure may affect the fluidity and permeability of the bilayer, and may explain the different sensitivity levels of various mycobacterial species to lipophilic inhibitors. Hydrophilic nutrients and inhibitors, in contrast, traverse the cell wall presumably through channels of recently discovered porins. PMID:7574484

  4. Effect of Bacterial Lipopolysaccharide Contamination on Gutta Percha- versus Resilon-Induced Human Monocyte Cell Line Toxicity.

    Directory of Open Access Journals (Sweden)

    Jamshid Hadjati

    2015-04-01

    Full Text Available Cytotoxic effects of obturation materials were tested in presence and absence of endotoxin on human monocytes in vitro.Human monocytes from THP-1 cell line were cultured. Three millimeters from the tip of each Resilon and gutta percha points were cut and directly placed at the bottom of the culture wells. Cultured cells were exposed to gutta percha (groups G1 and G2 and Resilon (R1 and R2. Ten μg/ml bacterial lipopolysaccharide (LPS was added to the culture wells in groups G1 and R1. Positive control included the bacterial LPS without the root canal filling material and the negative control contained the cells in culture medium only. Viability of cells was tested in all groups after 24, 48, and 72 hours using the methylthiazolyldiphenyl-tetrazolium bromide (MTT assay for at least 3 times to obtain reproducible results. Optical density values were read and the data were analyzed using three-way ANOVA and post hoc statistical test.The results showed that cells in G2 had the lowest rate of viability at 24 hours, but the lowest rate of viable cells was recorded in G1 at 48 and 72 hours. The effect of LPS treatment was not statistically significant. Resilon groups showed cell viability values higher than those of gutta percha groups, although statistically non-significant (P=0.105. Cell viability values were lower in gutta percha than Resilon groups when LPS-treated and LPS-untreated groups were compared independently at each time point.It could be concluded that none of the tested root canal filling materials had toxic effects on cultured human monocyte cells whether in presence or absence of LPS contamination.

  5. Vimentin in Bacterial Infections.

    Science.gov (United States)

    Mak, Tim N; Brüggemann, Holger

    2016-01-01

    Despite well-studied bacterial strategies to target actin to subvert the host cell cytoskeleton, thus promoting bacterial survival, replication, and dissemination, relatively little is known about the bacterial interaction with other components of the host cell cytoskeleton, including intermediate filaments (IFs). IFs have not only roles in maintaining the structural integrity of the cell, but they are also involved in many cellular processes including cell adhesion, immune signaling, and autophagy, processes that are important in the context of bacterial infections. Here, we summarize the knowledge about the role of IFs in bacterial infections, focusing on the type III IF protein vimentin. Recent studies have revealed the involvement of vimentin in host cell defenses, acting as ligand for several pattern recognition receptors of the innate immune system. Two main aspects of bacteria-vimentin interactions are presented in this review: the role of vimentin in pathogen-binding on the cell surface and subsequent bacterial invasion and the interaction of cytosolic vimentin and intracellular pathogens with regards to innate immune signaling. Mechanistic insight is presented involving distinct bacterial virulence factors that target vimentin to subvert its function in order to change the host cell fate in the course of a bacterial infection. PMID:27096872

  6. Vimentin in Bacterial Infections

    Directory of Open Access Journals (Sweden)

    Tim N. Mak

    2016-04-01

    Full Text Available Despite well-studied bacterial strategies to target actin to subvert the host cell cytoskeleton, thus promoting bacterial survival, replication, and dissemination, relatively little is known about the bacterial interaction with other components of the host cell cytoskeleton, including intermediate filaments (IFs. IFs have not only roles in maintaining the structural integrity of the cell, but they are also involved in many cellular processes including cell adhesion, immune signaling, and autophagy, processes that are important in the context of bacterial infections. Here, we summarize the knowledge about the role of IFs in bacterial infections, focusing on the type III IF protein vimentin. Recent studies have revealed the involvement of vimentin in host cell defenses, acting as ligand for several pattern recognition receptors of the innate immune system. Two main aspects of bacteria-vimentin interactions are presented in this review: the role of vimentin in pathogen-binding on the cell surface and subsequent bacterial invasion and the interaction of cytosolic vimentin and intracellular pathogens with regards to innate immune signaling. Mechanistic insight is presented involving distinct bacterial virulence factors that target vimentin to subvert its function in order to change the host cell fate in the course of a bacterial infection.

  7. The ABC's of Cell Division: Regulation of Peptidoglycan Amidase Activity during Cytokinesis in Escherichia coli

    OpenAIRE

    Yang, Desiree Choy

    2012-01-01

    The bacterial cell wall, composed of peptidoglycan (PG), is an essential component of the cell envelope. This macromolecular structure fortifies the cell membrane, determines cell shape, and helps prevent osmotic lysis. The synthesis and remodeling/recycling of this polymer is mediated by PG synthases and hydrolases, respectively. Proper control of the PG hydrolases is particularly important since misregulation of these enzymes can lead to lethal breaches in the cell wall. Surprisingly, howev...

  8. Enteric bacterial invasion of intestinal epithelial cells in vitro is dramatically enhanced using a vertical diffusion chamber model.

    Science.gov (United States)

    Naz, Neveda; Mills, Dominic C; Wren, Brendan W; Dorrell, Nick

    2013-01-01

    The interactions of bacterial pathogens with host cells have been investigated extensively using in vitro cell culture methods. However as such cell culture assays are performed under aerobic conditions, these in vitro models may not accurately represent the in vivo environment in which the host-pathogen interactions take place. We have developed an in vitro model of infection that permits the coculture of bacteria and host cells under different medium and gas conditions. The Vertical Diffusion Chamber (VDC) model mimics the conditions in the human intestine where bacteria will be under conditions of very low oxygen whilst tissue will be supplied with oxygen from the blood stream. Placing polarized intestinal epithelial cell (IEC) monolayers grown in Snapwell inserts into a VDC creates separate apical and basolateral compartments. The basolateral compartment is filled with cell culture medium, sealed and perfused with oxygen whilst the apical compartment is filled with broth, kept open and incubated under microaerobic conditions. Both Caco-2 and T84 IECs can be maintained in the VDC under these conditions without any apparent detrimental effects on cell survival or monolayer integrity. Coculturing experiments performed with different C. jejuni wild-type strains and different IEC lines in the VDC model with microaerobic conditions in the apical compartment reproducibly result in an increase in the number of interacting (almost 10-fold) and intracellular (almost 100-fold) bacteria compared to aerobic culture conditions. The environment created in the VDC model more closely mimics the environment encountered by C. jejuni in the human intestine and highlights the importance of performing in vitro infection assays under conditions that more closely mimic the in vivo reality. We propose that use of the VDC model will allow new interpretations of the interactions between bacterial pathogens and host cells. PMID:24192850

  9. In situ deposition of platinum nanoparticles on bacterial cellulose membranes and evaluation of PEM fuel cell performance

    International Nuclear Information System (INIS)

    In situ deposition of platinum (Pt) nanoparticles on bacterial cellulose membranes (BC) for a fuel cell application was studied. The platinum/bacterial cellulose (Pt/BC) membranes under different experimental conditions were characterized by using SEM (scanning electron microscopy), TEM (transmission electron microscopy), EDS (energy dispersive spectroscopy), XRD (X-ray diffractometry) and TG (thermo-gravimetric analysis) techniques. TEM images and XRD patterns both lead to the observation of spherical metallic platinum nanoparticles with mean diameter of 3-4 nm well impregnated into the BC fibrils. TG curves revealed these Pt/BC composite materials had the high thermal stability. The electrosorption of hydrogen was investigated by CV (cyclic voltammetry). It was found that Pt/BC catalysts have high electrocatalytic activity in the hydrogen oxidation reaction. The single cell performance of Pt/BC was tested at 20 deg. C, 30 deg. C, and 40 deg. C under non-humidified conditions. Preliminary tests on a single cell indicate that renewable BC is a good prospect to be explored as membrane in fuel cell field [B.R. Evans, H.M. O'Neill, V.P. Malyvanh, I. Lee, J. Woodward, Biosens. Bioelectron. 18 (2003) 917].

  10. In situ deposition of platinum nanoparticles on bacterial cellulose membranes and evaluation of PEM fuel cell performance

    Energy Technology Data Exchange (ETDEWEB)

    Yang Jiazhi [School of Chemistry and Chemical Engineering, Nan Jing University of Science and Technology, Nanjing 210094 (China); Sun Dongping [School of Chemistry and Chemical Engineering, Nan Jing University of Science and Technology, Nanjing 210094 (China)], E-mail: dongpingsun@163.com; Li Jun; Yang Xujie; Yu Junwei; Hao Qingli [School of Chemistry and Chemical Engineering, Nan Jing University of Science and Technology, Nanjing 210094 (China); Liu Wenming [Eco-materials and Renewable Energy Research Center, Department of Materials Science and Engineering and National Laboratory of Solid State Microstructures, Nanjing University, Nanjing 210093 (China); Liu Jianguo [Eco-materials and Renewable Energy Research Center, Department of Materials Science and Engineering and National Laboratory of Solid State Microstructures, Nanjing University, Nanjing 210093 (China)], E-mail: jianguoliu@nju.edu.cn; Zou Zhigang; Gu Jun [Eco-materials and Renewable Energy Research Center, Department of Materials Science and Engineering and National Laboratory of Solid State Microstructures, Nanjing University, Nanjing 210093 (China)

    2009-11-01

    In situ deposition of platinum (Pt) nanoparticles on bacterial cellulose membranes (BC) for a fuel cell application was studied. The platinum/bacterial cellulose (Pt/BC) membranes under different experimental conditions were characterized by using SEM (scanning electron microscopy), TEM (transmission electron microscopy), EDS (energy dispersive spectroscopy), XRD (X-ray diffractometry) and TG (thermo-gravimetric analysis) techniques. TEM images and XRD patterns both lead to the observation of spherical metallic platinum nanoparticles with mean diameter of 3-4 nm well impregnated into the BC fibrils. TG curves revealed these Pt/BC composite materials had the high thermal stability. The electrosorption of hydrogen was investigated by CV (cyclic voltammetry). It was found that Pt/BC catalysts have high electrocatalytic activity in the hydrogen oxidation reaction. The single cell performance of Pt/BC was tested at 20 deg. C, 30 deg. C, and 40 deg. C under non-humidified conditions. Preliminary tests on a single cell indicate that renewable BC is a good prospect to be explored as membrane in fuel cell field [B.R. Evans, H.M. O'Neill, V.P. Malyvanh, I. Lee, J. Woodward, Biosens. Bioelectron. 18 (2003) 917].

  11. Possible implication of bacterial infection in acute graft-versus-host disease after allogeneic hematopoietic stem cell transplantation

    Directory of Open Access Journals (Sweden)

    Shigeo eFuji

    2014-04-01

    Full Text Available Graft-versus-host disease (GVHD is still one of the major causes of morbidity and mortality in allogeneic hematopoietic stem cell transplantation (HSCT. In the pathogenesis of acute GVHD, it has been established that donor-derived T cells activated in the recipient play a major role in GVHD in initiation and maintenance within an inflammatory cascade. To reduce the risk of GVHD, intensification of GVHD prophylaxis like T cell depletion is effective, but it inevitably increases the risk of infectious diseases and abrogates beneficial graft-versus-leukemia effects. Although various cytokines are considered to play an important role in the pathogenesis of GVHD, GVHD initiation is such a complex process that cannot be prevented by means of single inflammatory cytokine inhibition. Thus, efficient methods to control the whole inflammatory milieu both on cellular and humoral view are needed. In this context, infectious diseases can theoretically contribute to an elevation of inflammatory cytokines after allogeneic HSCT and activation of various subtypes of immune effector cells, which might in summary lead to an aggravation of acute GVHD. The appropriate treatments or prophylaxis of bacterial infection during the early phase after allogeneic HSCT might be beneficial to reduce not only infectious-related but also GVHD-related mortality. Here, we aim to review the literature addressing the interactions of bacterial infections and GVHD after allogeneic HSCT.

  12. An HIV-1 envelope immunogen with W427S mutation in CD4 binding site induced more T follicular helper memory cells and reduced non-specific antibody responses.

    Directory of Open Access Journals (Sweden)

    Hao-Tong Yu

    Full Text Available The CD4 binding site (CD4BS of the HIV-1 envelope glycoprotein (Env contains epitopes for broadly neutralizing antibody (nAb and is the target for the vaccine development. However, the CD4BS core including residues 425-430 overlaps the B cell superantigen site and may be related to B cell exhaustion in HIV-1 infection. Furthermore, production of nAb and high-affinity plasma cells needs germinal center reaction and the help of T follicular helper (Tfh cells. We believe that strengthening the ability of Env CD4BS in inducing Tfh response and decreasing the effects of the superantigen are the strategies for eliciting nAb and development of HIV-1 vaccine. We constructed a gp120 mutant W427S of an HIV-1 primary R5 strain and examined its ability in the elicitation of Ab and the production of Tfh by immunization of BALB/c mice. We found that the trimeric wild-type gp120 can induce more non-specific antibody-secreting plasma cells, higher serum IgG secretion, and more Tfh cells by splenocyte. The modified W427S gp120 elicits higher levels of specific binding antibodies as well as nAbs though it produces less Tfh cells. Furthermore, higher Tfh cell frequency does not correlate to the specific binding Abs or nAbs indicating that the wild-type gp120 induced some non-specific Tfh that did not contribute to the production of specific Abs. This gp120 mutant led to more memory Tfh production, especially, the effector memory Tfh cells. Taken together, W427S gp120 could induce higher level of specific binding and neutralizing Ab production that may be associated with the reduction of non-specific Tfh but strengthening of the memory Tfh.

  13. Proteinaceous determinants of surface colonization in bacteria: Bacterial adhesion and biofilm formation from a protein secretion perspective

    Directory of Open Access Journals (Sweden)

    MickaelDesvaux

    2013-10-01

    Full Text Available Bacterial colonization of biotic or abiotic surfaces results from two quite distinct physiological processes, namely bacterial adhesion and biofilm formation. Broadly speaking, a biofilm is defined as the sessile development of microbial cells. Biofilm formation arises following bacterial adhesion but not all single bacterial cells adhering reversibly or irreversibly engage inexorably into a sessile mode of growth. Among molecular determinants promoting bacterial colonization, surface proteins are the most functionally diverse active components. To be present on the bacterial cell surface, though, a protein must be secreted in the first place. Considering the close association of secreted proteins with their cognate secretion systems, the secretome (which refers both to the secretion systems and their protein substrates is a key concept to apprehend the protein secretion and related physiological functions. The protein secretion systems are here considered in light of the differences in the cell-envelope architecture between diderm-LPS (archetypal Gram-negative, monoderm (archetypal Gram-positive and diderm-mycolate (archetypal acid-fast bacteria. Besides, their cognate secreted proteins engaged in the bacterial colonization process are regarded from single protein to supramolecular protein structure as well as the non-classical protein secretion. This state-of-the-art on the complement of the secretome (the secretion systems and their cognate effectors involved in the surface colonization process in diderm-LPS and monoderm bacteria paves the way for future research directions in the field.

  14. Natural Killer Cells and Helicobacter pylori Infection: Bacterial Antigens and Interleukin-12 Act Synergistically To Induce Gamma Interferon Production

    Science.gov (United States)

    Yun, Cheol H.; Lundgren, Anna; Azem, Josef; Sjöling, Åsa; Holmgren, Jan; Svennerholm, Ann-Mari; Lundin, B. Samuel

    2005-01-01

    Helicobacter pylori is known to induce a local immune response, which is characterized by activation of lymphocytes and the production of IFN-γ in the stomach mucosa. Since not only T cells, but also natural killer (NK) cells, are potent producers of gamma interferon (IFN-γ), we investigated whether NK cells play a role in the immune response to H. pylori infection. Our results showed that NK cells were present in both the gastric and duodenal mucosae but that H. pylori infection did not affect the infiltration of NK cells into the gastrointestinal area. Furthermore, we could show that NK cells could be activated directly by H. pylori antigens, as H. pylori bacteria, as well as lysate from H. pylori, induced the secretion of IFN-γ by NK cells. NK cells were also activated without direct contact when separated from the bacteria by an epithelial cell layer, indicating that the activation of NK cells by H. pylori can also occur in vivo, in the infected stomach mucosa. Moreover, the production of IFN-γ by NK cells was greatly enhanced when a small amount of interleukin-12 (IL-12) was added, and this synergistic effect was associated with increased expression of the IL-12 receptor β2. It was further evident that bacterial lysate alone was sufficient to induce the activation of cytotoxicity-related molecules. In conclusion, we demonstrated that NK cells are present in the gastroduodenal mucosa of humans and that NK cells produce high levels of IFN-γ when stimulated with a combination of H. pylori antigen and IL-12. We propose that NK cells play an active role in the local immune response to H. pylori infection. PMID:15731046

  15. Moisture Dynamics in Building Envelopes

    DEFF Research Database (Denmark)

    Peuhkuri, Ruut Hannele

    2003-01-01

    moisture conditions in the exterior weather and indoor climate the materials dynamically absorb and release moisture. The complexity of the impact of these conditions on the resulting moisture transport and content of the materials has been studied in this Thesis with controlled laboratory tests. The first......The overall scope of this Thesis "Moisture dynamics in building envelopes" has been to characterise how the various porous insulation materials investigated performed hygrothermally under conditions similar to those in a typical building envelope. As a result of the changing temperature and...

  16. Moisture dynamics in building envelopes

    Energy Technology Data Exchange (ETDEWEB)

    Peuhkuri, R.

    2003-07-01

    The overall scope of this Thesis 'Moisture dynamics in building envelopes' has been to characterise how the various porous insulation materials investigated performed hygro thermally under conditions similar to those in a typical building envelope. As a result of the changing temperature and moisture conditions in the exterior weather and indoor climate the materials dynamically absorb and release moisture. The complexity of the impact of these conditions on the resulting moisture transport and content of the materials has been studied in this Thesis with controlled laboratory tests. (au)

  17. Envelope Inflation or Stellar Wind?

    Science.gov (United States)

    Ro, S.; Matzner, C. D.

    We an optically-thick, transonic, steady wind model for a H-free Wolf-Rayet star. A bifurcation is found across a critical mass loss rate Mb. Slower winds M < Mb extend by several hydrostatic stellar radii, reproduce features of envelope in ation from Petrovic et al. (2006) and Gräfener et al. (2012), and are energetically unbound. This work is of particular interest for extended envelopes and winds, radiative hydrodynamic instabilities (eg. wind stagnation, clumping, etc.), and NLTE atmospheric models.

  18. The deletion of bacterial dynamin and flotillin genes results in pleiotrophic effects on cell division, cell growth and in cell shape maintenance

    Directory of Open Access Journals (Sweden)

    Dempwolff Felix

    2012-12-01

    the cell membrane, where they assemble even in the absence of any bacterial cofactor.

  19. Degradation of lucerne stem cell walls by five rumen bacterial species

    NARCIS (Netherlands)

    Jung, H.G.; Engels, F.M.; Weimer, P.J.

    2004-01-01

    The rumen bacterial strains Butyrivibrio fibrisolvens H17c, Fibrobacter succinogenes S85, Lachnospira multiparus 40, Ruminococcus albus 7 and R. flavefaciens FD-1 were compared individually and as a five-species mixture with a rumen inoculum for their ability to degrade lucerne (Medicago sativa L.)

  20. Spatial distribution of bacterial communities on volumetric and planar anodes in single-chamber air-cathode microbial fuel cells

    KAUST Repository

    Vargas, Ignacio T.

    2013-05-29

    Pyrosequencing was used to characterize bacterial communities in air-cathode microbial fuel cells across a volumetric (graphite fiber brush) and a planar (carbon cloth) anode, where different physical and chemical gradients would be expected associated with the distance between anode location and the air cathode. As expected, the stable operational voltage and the coulombic efficiency (CE) were higher for the volumetric anode than the planar anode (0.57V and CE=22% vs. 0.51V and CE=12%). The genus Geobacter was the only known exoelectrogen among the observed dominant groups, comprising 57±4% of recovered sequences for the brush and 27±5% for the carbon-cloth anode. While the bacterial communities differed between the two anode materials, results showed that Geobacter spp. and other dominant bacterial groups were homogenously distributed across both planar and volumetric anodes. This lends support to previous community analysis interpretations based on a single biofilm sampling location in these systems. © 2013 Wiley Periodicals, Inc.

  1. Effect of Structure on the Interactions between Five Natural Antimicrobial Compounds and Phospholipids of Bacterial Cell Membrane on Model Monolayers

    Directory of Open Access Journals (Sweden)

    Stella W. Nowotarska

    2014-06-01

    Full Text Available Monolayers composed of bacterial phospholipids were used as model membranes to study interactions of the naturally occurring phenolic compounds 2,5-dihydroxybenzaldehyde and 2-hydroxy-5-methoxybenzaldehyde, and the plant essential oil compounds carvacrol, cinnamaldehyde, and geraniol, previously found to be active against both Gram-positive and Gram-negative pathogenic microorganisms. The lipid monolayers consist of 1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DPPE, 1,2-dihexa- decanoyl-sn-glycero-3-phospho-(1'-rac-glycerol (DPPG, and 1,1',2,2'-tetratetradecanoyl cardiolipin (cardiolipin. Surface pressure–area (π-A and surface potential–area (Δψ-A isotherms were measured to monitor changes in the thermodynamic and physical properties of the lipid monolayers. Results of the study indicated that the five compounds modified the three lipid monolayer structures by integrating into the monolayer, forming aggregates of antimicrobial –lipid complexes, reducing the packing effectiveness of the lipids, increasing the membrane fluidity, and altering the total dipole moment in the monolayer membrane model. The interactions of the five antimicrobial compounds with bacterial phospholipids depended on both the structure of the antimicrobials and the composition of the monolayers. The observed experimental results provide insight into the mechanism of the molecular interactions between naturally-occurring antimicrobial compounds and phospholipids of the bacterial cell membrane that govern activities.

  2. Biointerface by Cell Growth on Graphene Oxide Doped Bacterial Cellulose/Poly(3,4-ethylenedioxythiophene) Nanofibers.

    Science.gov (United States)

    Chen, Chuntao; Zhang, Ting; Zhang, Qi; Chen, Xiao; Zhu, Chunlin; Xu, Yunhua; Yang, Jiazhi; Liu, Jian; Sun, Dongping

    2016-04-27

    Highly biocompatible advanced materials with excellent electroactivity are increasingly meaningful to biointerfaces and the development of biomedicine. Herein, bacterial cellulose/poly(3,4-ethylene dioxythiophene)/graphene oxide (BC/PEDOT/GO) composite nanofibers were synthesized through the in situ interfacial polymerization of PEDOT with the doping of GO. The abundant free carboxyl and hydroxy groups offer the BC/PEDOT/GO film active functional groups for surface modification. We demonstrate the use of this composite nanofiber for the electrical stimulation of PC12 neural cells as this resultant nanofiber scaffold could closely mimic the structure of the native extracellular matrix (ECM) with a promoting cell orientation and differentiation after electrical stimulation of PC12 cells. It is expected that this biocompatible BC/PEDOT/GO material will find potential applications in biological and regenerative medicine. PMID:27054801

  3. Bacterial single-cell activities along the nutrient availability gradient in a canyon-shaped reservoir: a seasonal study

    Czech Academy of Sciences Publication Activity Database

    Horňák, Karel; Jezbera, Jan; Šimek, Karel

    2010-01-01

    Roč. 60, č. 3 (2010), s. 215-225. ISSN 0948-3055. [Congress of the International Limnological Society /31./. Kapské Město, 15.08.2010-20.08.2010] R&D Projects: GA ČR(CZ) GA206/08/0015; GA ČR(CZ) GAP504/10/1534 Institutional research plan: CEZ:AV0Z60170517 Keywords : reservoir * bacterial activity * leucine and glucose incorporation * HNA and LNA cells Subject RIV: EE - Microbiology, Virology Impact factor: 2.089, year: 2010

  4. Use of bacterial and firefly luciferases as reporter genes in DEAE-dextran-mediated transfection of mammalian cells.

    Science.gov (United States)

    Pazzagli, M; Devine, J H; Peterson, D O; Baldwin, T O

    1992-08-01

    The aim of this study was to compare three different luciferase genes by placing them in a single reporter vector and expressing them in the same mammalian cell type. The luciferase genes investigated were the luc genes from the fireflies Photinus pyralis (PP) and Luciola mingrelica (LM) and the lux AB5 gene, a translational fusion of the two subunits of the bacterial luciferase from Vibrio harveyi (VH). The chloramphenicol acetyltransferase (CAT) gene was also included in this study for comparison. The performances of the assay methods of the corresponding enzymes were evaluated using reference materials and the results of the expressed enzymes following transfection were calculated using calibration curves. All of the bioluminescent assays possess high reproducibility both within and between the batches (less than 15%). The comparison of the assay methods shows that firefly luciferases have the highest detection sensitivity (0.05 and 0.08 amol for PP and LM, respectively) whereas the VH bacterial luciferase has 5 amol and CAT 100 amol. On the other hand, the transfection of the various plasmids shows that the content of the expressed enzyme within the cells is much higher for CAT than for the other luciferase genes. VH luciferase is expressed at very low levels in mammalian cells due to the relatively high temperature of growing of the mammalian cells that seems to impair the correct folding of the active enzyme. PP and LM luciferases are both expressed at picomolar level but usually 10 to 70 times less in content with respect to CAT within the transfected cells. On the basis of these results the overall improvement in sensitivity related to the use of firefly luciferases as reporter genes in mammalian cells is about 30 to 50 times with respect to that of CAT. PMID:1443530

  5. Pectin and Xyloglucan Influence the Attachment of Salmonella enterica and Listeria monocytogenes to Bacterial Cellulose-Derived Plant Cell Wall Models.

    Science.gov (United States)

    Tan, Michelle S F; Rahman, Sadequr; Dykes, Gary A

    2016-01-01

    Minimally processed fresh produce has been implicated as a major source of foodborne microbial pathogens globally. These pathogens must attach to the produce in order to be transmitted. Cut surfaces of produce that expose cell walls are particularly vulnerable. Little is known about the roles that different structural components (cellulose, pectin, and xyloglucan) of plant cell walls play in the attachment of foodborne bacterial pathogens. Using bacterial cellulose-derived plant cell wall models, we showed that the presence of pectin alone or xyloglucan alone affected the attachment of three Salmonella enterica strains (Salmonella enterica subsp. enterica serovar Enteritidis ATCC 13076, Salmonella enterica subsp. enterica serovar Typhimurium ATCC 14028, and Salmonella enterica subsp. indica M4) and Listeria monocytogenes ATCC 7644. In addition, we showed that this effect was modulated in the presence of both polysaccharides. Assays using pairwise combinations of S. Typhimurium ATCC 14028 and L. monocytogenes ATCC 7644 showed that bacterial attachment to all plant cell wall models was dependent on the characteristics of the individual bacterial strains and was not directly proportional to the initial concentration of the bacterial inoculum. This work showed that bacterial attachment was not determined directly by the plant cell wall model or bacterial physicochemical properties. We suggest that attachment of the Salmonella strains may be influenced by the effects of these polysaccharides on physical and structural properties of the plant cell wall model. Our findings improve the understanding of how Salmonella enterica and Listeria monocytogenes attach to plant cell walls, which may facilitate the development of better ways to prevent the attachment of these pathogens to such surfaces. PMID:26567310

  6. Identification of immunodominant regions and linear B cell epitopes of the gE envelope protein of varicella-zoster virus.

    Science.gov (United States)

    Fowler, W J; Garcia-Valcarcel, M; Hill-Perkins, M S; Murphy, G; Harper, D R; Jeffries, D J; Burns, N R; Adams, S E; Kingsman, A J; Layton, G T

    1995-12-20

    The envelope proteins of varicella-zoster virus (VZV) are highly immunogenic and one of the most abundant is glycoprotein E (gE). However, its immunodominant regions and epitopes have not been identified. In this study, using human sera from individuals with recent varicella or zoster infections, we have localized antigenic sequences of gE using recombinant hybrid Ty-virus-like particles (VLPs) carrying overlapping fragments of the gE protein. gE(1-134)-VLPs (particles carrying amino acids 1-134 of gE) and, to a lesser extent, gE(101-161)-VLPs were found to be the most antigenic when tested by Western blotting and ELISA. Other fragments of gE (spanning residues 161-623) showed weak or no antigenicity. Pepscan analysis of human sera on overlapping synthetic peptides representing residues 1-135 of gE revealed that the most antigenic region was between residues 50 and 135. Three immunodominant sequences (residues 86-105, 116-135, and, to a lesser extent, 56-75) were detected using sera from both varicella and zoster patients. All sera from varicella, but not zoster, patients reacted strongly with an epitope in peptide 66-85. Other epitopes were recognized weakly by some varicella or zoster sera. More sera need to be tested to assess the potential disease specificity of these epitopes. The neutralizing monoclonal antibody (MAb) IF-B9 reacted with residues 71-90; however, another neutralizing MAb, SG1A, which bound to both gE(1-134)-VLPs and gE(101-161)-VLPs did not bind to any peptide. The identification of immunodominant sequences of gE will help toward the development of a subunit VZV vaccine. PMID:8553555

  7. Several novel nuclear envelope transmembrane proteins identified in skeletal muscle have cytoskeletal associations.

    Science.gov (United States)

    Wilkie, Gavin S; Korfali, Nadia; Swanson, Selene K; Malik, Poonam; Srsen, Vlastimil; Batrakou, Dzmitry G; de las Heras, Jose; Zuleger, Nikolaj; Kerr, Alastair R W; Florens, Laurence; Schirmer, Eric C

    2011-01-01

    Nuclear envelopes from liver and a neuroblastoma cell line have previously been analyzed by proteomics; however, most diseases associated with the nuclear envelope affect muscle. To determine whether muscle has unique nuclear envelope proteins, rat skeletal muscle nuclear envelopes were prepared and analyzed by multidimensional protein identification technology. Many novel muscle-specific proteins were identified that did not appear in previous nuclear envelope data sets. Nuclear envelope residence was confirmed for 11 of these by expression of fusion proteins and by antibody staining of muscle tissue cryosections. Moreover, transcript levels for several of the newly identified nuclear envelope transmembrane proteins increased during muscle differentiation using mouse and human in vitro model systems. Some of these proteins tracked with microtubules at the nuclear surface in interphase cells and accumulated at the base of the microtubule spindle in mitotic cells, suggesting they may associate with complexes that connect the nucleus to the cytoskeleton. The finding of tissue-specific proteins in the skeletal muscle nuclear envelope proteome argues the importance of analyzing nuclear envelopes from all tissues linked to disease and suggests that general investigation of tissue differences in organellar proteomes might yield critical insights. PMID:20876400

  8. A channel connecting the mother cell and forespore during bacterial endospore formation

    OpenAIRE

    Meisner, Jeffrey; Xin WANG; Serrano, Monica; Henriques, Adriano O.; Moran, Charles P.

    2008-01-01

    At an early stage during Bacillus subtilis endospore development the bacterium divides asymmetrically to produce two daughter cells. The smaller cell (forespore) differentiates into the endospore, while the larger cell (mother cell) becomes a terminally differentiated cell that nurtures the developing forespore. During development the mother cell engulfs the forespore to produce a protoplast, surrounded by two bilayer membranes, which separate it from the cytoplasm of the mother cell. The act...

  9. Energy efficiency of building envelope

    OpenAIRE

    V.M. Yakubson

    2014-01-01

    November, 12-13th, in Saint-Petersburg the 7th International congress "Energy efficiency. XXI century" took place. The reports were done in breakuo groups according to the various aspects of energy efficiency challenge: HVAC systems, water supply and sewerage systems, gas supply, energy metering. One of the grourps was devoted to thermophysics of buildings and energy effective design of building envelope.

  10. Outliers In Data Envelopment Analysis

    Directory of Open Access Journals (Sweden)

    Shaik Khaleel Ahamed

    2015-06-01

    Full Text Available Data Envelopment Analysis is a linear programming technique that assigns efficiency scores to firms engaged in producing similar outputs employing similar inputs. Extremely efficient firms are potential Outliers. The method developed detects Outliers, implementing Stochastic Threshold Value, with computational ease. It is useful in data filtering in BIG DATA problems.

  11. Biosorption of metal ions by attached bacterial cells in a packed-bed bioreactor

    International Nuclear Information System (INIS)

    This work describes a simple method for the immobilization of a biosorbent. An adherent Bacillus sp. strain has been grown attached to an inert support material. This strain had the capacity to bind uranium, copper, cadmium and zinc. The desorption of these metals was quantitative at pH-values lower than 2. To study the attachment of the bacterial biomass, a laboratory-scale packed-bed bioreactor with an appropriate aeration system was developed. The colonization of the support was fast and efficient. In batch culture conditions, the biomass accumulation reached a cuasi-stationary phase after 12 h. Under optimal conditions, the attached biomass comprised around 80% of the total biomass present in the bioreactor. After the colonization phase, the packed-bed bioreactor was continuously operated to remove heavy metals from aqueous solutions. The biosorption capacity of the attached biomass was similar to that of the free bacterial suspension

  12. Comparative study of HOCl-inflicted damage to bacterial DNA ex vivo and within cells

    OpenAIRE

    Suquet, Christine; Warren, Jeffrey J.; Seth, Nimulrith; Hurst, James K.

    2009-01-01

    The prospects for using bacterial DNA as an intrinsic probe for HOCl and secondary oxidants/chlorinating agents associated with it has been evaluated using both in vitro and in vivo studies. Single-strand and double-strand breaks occurred in bare plasmid DNA that had been exposed to high levels of HOCl, although these reactions were very inefficient compared to polynucleotide chain cleavage caused by the OH•-generating reagent, peroxynitrite. Plasmid nicking was not increased when intact Esch...

  13. Nitrate reducing bacterial activity in concrete cells of nuclear waste disposal

    OpenAIRE

    Albrecht A.; Rafrafi Y.; Sablayrolles C.; Bertron A.; Kassim C.; Alquier M.; Erable B.

    2013-01-01

    International audience Leaching experiments of solid matrices (bitumen and cement pastes) have been first implemented to define the physicochemical conditions that microorganisms are likely to meet at the bitumen-concrete interface (see the paper of Bertron et al.). Of course, as might be suspected, the cement matrix imposes highly alkaline pH conditions (10 < pH < 11). The screening of a range of anaerobic denitrifying bacterial strains led us to select Halomonas desiderata as a model bac...

  14. The Salmonella enterica virulence : Its role in bacterial adaption to mammalian and protozoan cells

    OpenAIRE

    Tezcan-Merdol, Dilek

    2004-01-01

    Salmonellae are Gram-negative enteric bacteria and facultative intracellular pathogens responsible for a diversity of illnesses in a wide range of hosts, including man. Many serovars of Salmonella enterica harbor a plasmid that enhances bacterial virulence in infection models, and that seems to promote extraintestinal infection in man. Consequently, the plasmid has been referred to as the"virulence plasmid". The virulence plasmid varies in its constitution among different se...

  15. Commensal bacterial internalization by epithelial cells: An alternative portal for gut leakiness

    OpenAIRE

    Yu, Linda Chia-Hui

    2015-01-01

    Co-existing paracellular and transcellular barrier defect in intestinal epithelium was documented in inflammatory bowel disease, celiac disease, and intestinal obstruction. Mechanisms regarding tight junction disruption have been extensively studied; however, limited progress has been made in research on bacterial transcytosis. Densely packed brush border (BB), with cholesterol-based lipid rafts in the intermicrovillous membrane invagination, serves as an ultrastructural barrier to prevent di...

  16. Respiratory Viruses Augment the Adhesion of Bacterial Pathogens to Respiratory Epithelium in a Viral Species- and Cell Type-Dependent Manner

    OpenAIRE

    Avadhanula, Vasanthi; Rodriguez, Carina A.; DeVincenzo, John P.; Wang, Yan; Webby, Richard J; Ulett, Glen C.; Adderson, Elisabeth E.

    2006-01-01

    Secondary bacterial infections often complicate respiratory viral infections, but the mechanisms whereby viruses predispose to bacterial disease are not completely understood. We determined the effects of infection with respiratory syncytial virus (RSV), human parainfluenza virus 3 (HPIV-3), and influenza virus on the abilities of nontypeable Haemophilus influenzae and Streptococcus pneumoniae to adhere to respiratory epithelial cells and how these viruses alter the expression of known recept...

  17. Cell-penetrating peptides mediated protein cross-membrane delivery and its use in bacterial vector vaccine.

    Science.gov (United States)

    Ma, Jimei; Xu, Jinmei; Guan, Lingyu; Hu, Tianjian; Liu, Qin; Xiao, Jingfan; Zhang, Yuanxing

    2014-07-01

    It is an attractive strategy to develop a recombinant bacterial vector vaccine by expressing exogenous protective antigen to induce the immune response, and the main concern is how to enhance the cellular internalization of antigen produced by bacterial vector. Cell-penetrating peptides (CPPs) are short cationic/amphipathic peptides which facilitate cellular uptake of various molecular cargoes and therefore have great potentials in vector vaccine design. In this work, eleven different CPPs were fused to the C-terminus of EGFP respectively, and the resultant EGFP-CPP fusion proteins were expressed and purified to assay their cross-membrane transport in macrophage J774 A.1 cells. Among the tested CPPs, TAT showed an excellent capability to deliver the cargo protein EGFP into cytoplasm. In order to establish an efficient antigen delivery system in Escherichia coli, the EGFP-TAT synthesis circuit was combined with an in vivo inducible lysis circuit PviuA-E in E. coli to form an integrated antigen delivery system, the resultant E. coli was proved to be able to lyse upon the induction of a mimic in vivo signal and thus release intracellular EGFP-TAT intensively, which were assumed to undergo a more efficient intracellular delivery by CPP to evoke protective immune responses. Based on the established antigen delivery system, the protective antigen gene flgD from an invasive intracellular fish pathogen Edwardsiella tarda EIB202, was applied to establish an E. coli recombinant vector vaccine. This E. coli vector vaccine presented superior immune protection (RPS = 63%) under the challenge with E. tarda EIB202, suggesting that the novel antigen delivery system had great potential in bacterial vector vaccine applications. PMID:24746937

  18. Human Langerhans cells control Th cells via programmed death-ligand 1 in response to bacterial stimuli and nickel-induced contact allergy.

    Directory of Open Access Journals (Sweden)

    Manuel Hitzler

    Full Text Available Langerhans cells (LCs are suspected to initiate inflammatory immune responses to contact allergens and pathogenic bacteria. In chronic infectious diseases, programmed death ligand (PD-L 1 exhibits both inhibitory and costimulatory functions on T cell-mediated activation and tolerance. Here, we investigated the effects of contact allergens and bacterial stimuli on PD-L1 expression in LCs and the effects of altered PD-L1 expression on cytokine release of subsequently cocultured T cells. Monocyte-derived LCs (MoLCs, LCs, and skin sections of patients suffering from allergic contact dermatitis were challenged with nickel and then analyzed for PD-L1 expression by confocal laser scanning microscopy and flow cytometry. In blocking experiments, we found that the release of Th cell specific cytokines was dependent on both stimulation of LCs and inhibition of PD-L1-PD-1 interactions. Stimulation with peptidoglycan (PGN or lipopolysaccharide (LPS and blockage of PD-L1 with a specific antibody triggered the release of high levels of IL-17, IL-22, TNF-α, and IFN-γ in CD4(+T cells. If nickel was used as a stimulus, blockage of PD-L1 led to high amounts of TNF-α and IL-22. A closer look revealed PD-L1-dependent upregulation of IL-17 secretion in FACS-sorted CCR6(+/CCR4(+ T memory cells. In the presence of anti-PD-L1, PGN induced secretion of IFN-γ and IL-17 in total CCR6(+ cells, while nickel triggered secretion of IFN-γ and IL-17 exclusively in CCR6(+/CCR4(+ cells. Our findings suggest that PD-L1 on LCs plays a crucial role in type IV allergic reactions and in response to bacterial stimuli by controlling the nature of inflammatory Th cell responses.

  19. Effects of inoculation sources on the enrichment and performance of anode bacterial consortia in sensor typed microbial fuel cells

    Directory of Open Access Journals (Sweden)

    Phuong Tran

    2016-01-01

    Full Text Available Microbial fuel cells are a recently emerging technology that promises a number of applications in energy recovery, environmental treatment and monitoring. In this study, we investigated the effect of inoculating sources on the enrichment of electrochemically active bacterial consortia in sensor-typed microbial fuel cells (MFCs. Several MFCs were constructed, operated with modified artificial wastewater and inoculated with different microbial sources from natural soil, natural mud, activated sludge, wastewater and a mixture of those sources. After enrichment, the MFCs inoculated with the natural soil source generated higher and more stable currents (0.53±0.03 mA, in comparisons with the MFCs inoculated with the other sources. The results from denaturing gradient gel electrophoresis (DGGE showed that there were significant changes in bacterial composition from the original inocula to the enriched consortia. Even more interestingly, Pseudomonas sp. was found dominant in the natural soil source and also in the corresponding enriched consortium. The interactions between Pseudomonas sp. and other species in such a community are probably the key for the effective and stable performance of the MFCs.

  20. Marking 100 years since Rudolf Höber’s discovery of the insulating envelope surrounding cells and of the beta-dispersion exhibited by tissue

    Directory of Open Access Journals (Sweden)

    Ronald Pethig

    2012-11-01

    Full Text Available Between 1910 and 1913 Rudolf Höber presented proof that the interiors of red blood cells and muscle cells contain conducting electrolytes, and that each conducting core is contained within an insulating membrane.  He did this by demonstrating, in a series of remarkable electrical experiments, that the conductivity of compacted cell samples at low frequencies (~150 Hz was about ten-times less than the value obtained at ~5 MHz.  On perforation of the membrane, the low-frequency conductivity increased to a value approaching that exhibited at MHz frequencies. Apart from representing a major milestone in the development of cell biology and electrophysiology, Höber’s work was the first description of what we now call the dielectric b-dispersion exhibited by cell suspensions and fresh tissue.

  1. Bacterial intermediate filaments

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Cabeen, M.; Jacobs-Wagner, C.

    2009-01-01

    controls the length of the structure by favoring lateral insertion of crescentin subunits over bipolar longitudinal extension when the structure ends reach the cell poles. The crescentin structure is stably anchored to the cell envelope, and this cellular organization requires MreB function, identifying a...... new function for MreB and providing a parallel to the role of actin in IF assembly and organization in metazoan cells. Additionally, analysis of an MreB localization mutant suggests that cell wall insertion during cell elongation normally occurs along two helices of opposite handedness, each...

  2. Regulation of DMBT1 via NOD2 and TLR4 in intestinal epithelial cells modulates bacterial recognition and invasion

    DEFF Research Database (Denmark)

    Rosenstiel, Philip; Sina, Christian; End, Caroline;

    2007-01-01

    -kappaB activation and cytokine secretion in vitro. Thus, DMBT1 may play an important role in the first line of mucosal defense conferring immune exclusion of bacterial cell wall components. Dysregulated intestinal DMBT1 expression due to mutations in the NOD2/CARD15 gene may be part of the complex pathophysiology......Mucosal epithelial cell layers are constantly exposed to a complex resident microflora. Deleted in malignant brain tumors 1 (DMBT1) belongs to the group of secreted scavenger receptor cysteine-rich proteins and is considered to be involved in host defense by pathogen binding. This report describes...... intracellular pathogen receptor NOD2 via NF-kappaB activation. DMBT1 is strongly up-regulated in the inflamed intestinal mucosa of Crohn's disease patients with wild-type, but not with mutant NOD2. We show that DMBT1 inhibits cytoinvasion of Salmonella enterica and LPS- and muramyl dipeptide-induced NF...

  3. Dynamics of extended AGB star envelopes

    CERN Document Server

    Dreyer, C; Sedlmayr, E

    2010-01-01

    The dust formed in extended circumstellar envelopes of long-period variables and Miras has a strong influence on the envelope dynamics. A radiatively driven instability caused by the formation of dust leads to the development of an autonomous dynamics characterised by a set of distinct frequencies. We study the interplay between the envelope's internal dynamics and an external excitation by a pulsating star.

  4. Codelivery of Envelope Protein in Alum with MVA Vaccine Induces CXCR3-Biased CXCR5+ and CXCR5- CD4 T Cell Responses in Rhesus Macaques.

    Science.gov (United States)

    Iyer, Smita S; Gangadhara, Sailaja; Victor, Blandine; Gomez, Rosy; Basu, Rahul; Hong, Jung Joo; Labranche, Celia; Montefiori, David C; Villinger, Francois; Moss, Bernard; Amara, Rama Rao

    2015-08-01

    The goal of an HIV vaccine is to generate robust and durable protective Ab. Vital to this goal is the induction of CD4(+) T follicular helper (TFH) cells. However, very little is known about the TFH response to HIV vaccination and its relative contribution to magnitude and quality of vaccine-elicited Ab titers. In this study, we investigated these questions in the context of a DNA/modified vaccinia virus Ankara SIV vaccine with and without gp140 boost in aluminum hydroxide in rhesus macaques. In addition, we determined the frequency of vaccine-induced CD4(+) T cells coexpressing chemokine receptor, CXCR5 (facilitates migration to B cell follicles) in blood and whether these responses were representative of lymph node TFH responses. We show that booster modified vaccinia virus Ankara immunization induced a distinct and transient accumulation of proliferating CXCR5(+) and CXCR5(-) CD4 T cells in blood at day 7 postimmunization, and the frequency of the former but not the latter correlated with TFH and B cell responses in germinal centers of the lymph node. Interestingly, gp140 boost induced a skewing toward CXCR3 expression on germinal center TFH cells, which was strongly associated with longevity, avidity, and neutralization potential of vaccine-elicited Ab response. However, CXCR3(+) cells preferentially expressed the HIV coreceptor CCR5, and vaccine-induced CXCR3(+)CXCR5(+) cells showed a moderate positive association with peak viremia following SIV251 infection. Taken together, our findings demonstrate that vaccine regimens that elicit CXCR3-biased TFH cell responses favor Ab persistence and avidity but may predispose to higher acute viremia in the event of breakthrough infections. PMID:26116502

  5. Bacterial hydrodynamics

    CERN Document Server

    Lauga, Eric

    2015-01-01

    Bacteria predate plants and animals by billions of years. Today, they are the world's smallest cells yet they represent the bulk of the world's biomass, and the main reservoir of nutrients for higher organisms. Most bacteria can move on their own, and the majority of motile bacteria are able to swim in viscous fluids using slender helical appendages called flagella. Low-Reynolds-number hydrodynamics is at the heart of the ability of flagella to generate propulsion at the micron scale. In fact, fluid dynamic forces impact many aspects of bacteriology, ranging from the ability of cells to reorient and search their surroundings to their interactions within mechanically and chemically-complex environments. Using hydrodynamics as an organizing framework, we review the biomechanics of bacterial motility and look ahead to future challenges.

  6. An Envelope Glycoprotein of the Human Endogenous Retrovirus HERV-W Is Expressed in the Human Placenta and Fuses Cells Expressing the Type D Mammalian Retrovirus Receptor

    OpenAIRE

    Blond, Jean-Luc; Lavillette, Dimitri; Cheynet, Valérie; Bouton, Olivier; Oriol, Guy; Chapel-Fernandes, Sylvie; Mandrand, Bernard; Mallet, François; Cosset, François-Loïc

    2000-01-01

    A new human endogenous retrovirus (HERV) family, termed HERV-W, was recently described (J.-L. Blond, F. Besème, L. Duret, O. Bouton, F. Bedin, H. Perron, B. Mandrand, and F. Mallet, J. Virol. 73:1175–1185, 1999). HERV-W mRNAs were found to be specifically expressed in placenta cells, and an env cDNA containing a complete open reading frame was recovered. In cell-cell fusion assays, we demonstrate here that the product of the HERV-W env gene is a highly fusogenic membrane glycoprotein. Transfe...

  7. Conformational instability governed by disulfide bonds partitions the dominant from subdominant helper T-cell responses specific for HIV-1 envelope glycoprotein gp120

    OpenAIRE

    Nguyen, Hong-Nam P.; Steede, N. Kalaya; Robinson, James E.; Landry, Samuel J.

    2015-01-01

    Most individuals infected with human immunodeficiency virus type 1 (HIV-1) generate a CD4+ T-cell response that is dominated by a few epitopes. Immunodominance may be counterproductive because a broad CD4+ T-cell response is associated with reduced viral load. Previous studies indicated that antigen three-dimensional structure controls antigen processing and presentation and therefore CD4+ T-cell epitope dominance. Dominant epitopes occur adjacent to the V1-V2, V3, and V4 loops because proteo...

  8. Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria.

    OpenAIRE

    Mistou, Michel-Yves; Sutcliffe, Iain; van Sorge, Nina

    2016-01-01

    The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall te...

  9. Comparison of quantitative and qualitative antibody-producing cell responses to lipopolysaccharide in cell walls of the bacterial form and in membranes of the protoplast L-form of Proteus mirabilis.

    OpenAIRE

    Karch, H; Nixdorff, K

    1980-01-01

    Membranes of the stable protoplast L-form of Proteus mirabilis strain VI were highly immunogenic carriers of lipopolysaccharide when compared with the immune responses to lipopolysaccharide contained in cell walls of the bacterial form of this organism.

  10. Bacterial cell wall preservation during organic matter diagenesis in sediments off Peru

    DEFF Research Database (Denmark)

    Lomstein, Bente Aagaard; Niggemann, Jutta; Jørgensen, Bo Barker;

    evaluated from the percentage of carbon and nitrogen present as amino acid carbon and nitrogen, the ratio between protein precursors and their non-protein degradation products, compositional changes in the amino acid spectra and the percentage of carbon and nitrogen present as amino sugar carbon and...... nitrogen. The study clearly demonstrated a strong bacterial imprint in organic matter during early diagenesis. Hence, the key players in organic matter mineralization became an increasingly import component of refractory organic matter with ongoing degradation. Session #:053 Date: 01-27-09 Time: 11...

  11. Identification of novel bacterial histidine biosynthesis inhibitors using docking, ensemble rescoring, and whole-cell assays

    DEFF Research Database (Denmark)

    Henriksen, Signe Teuber; Liu, J.; Estiu, G.;

    2010-01-01

    . aureus histidine biosynthesis pathway, which is predicted to be essential for bacterial biomass productions. Virtual screening of a library of similar to 10(6) compounds identified 49 potential inhibitors of three enzymes of this pathway. Eighteen representative compounds were directly tested on three S....... aureus-and two Escherichia coli strains in standard disk inhibition assays. Thirteen compounds are inhibitors of some or all of the S. aureus strains, while 14 compounds weakly inhibit growth in one or both E. coli strains. The high hit rate obtained from a fast virtual screen demonstrates the...

  12. The herpes simplex virus UL20 protein functions in glycoprotein K (gK intracellular transport and virus-induced cell fusion are independent of UL20 functions in cytoplasmic virion envelopment

    Directory of Open Access Journals (Sweden)

    Kousoulas Konstantin G

    2007-11-01

    Full Text Available Abstract The HSV-1 UL20 protein (UL20p and glycoprotein K (gK are both important determinants of cytoplasmic virion morphogenesis and virus-induced cell fusion. In this manuscript, we examined the effect of UL20 mutations on the coordinate transport and Trans Golgi Network (TGN localization of UL20p and gK, virus-induced cell fusion and infectious virus production. Deletion of 18 amino acids from the UL20p carboxyl terminus (UL20 mutant 204t inhibited intracellular transport and cell-surface expression of both gK and UL20, resulting in accumulation of UL20p and gK in the endoplasmic reticulum (ER in agreement with the inability of 204t to complement UL20-null virus replication and virus-induced cell fusion. In contrast, less severe carboxyl terminal deletions of either 11 or six amino acids (UL20 mutants 211t and 216t, respectively allowed efficient UL20p and gK intracellular transport, cell-surface expression and TGN colocalization. However, while both 211t and 216t failed to complement for infectious virus production, 216t complemented for virus-induced cell fusion, but 211t did not. These results indicated that the carboxyl terminal six amino acids of UL20p were crucial for infectious virus production, but not involved in intracellular localization of UL20p/gK and concomitant virus-induced cell fusion. In the amino terminus of UL20, UL20p mutants were produced changing one or both of the Y38 and Y49 residues found within putative phosphorylation sites. UL20p tyrosine-modified mutants with both tyrosine residues changed enabled efficient intracellular transport and TGN localization of UL20p and gK, but failed to complement for either infectious virus production, or virus-induced cell fusion. These results show that UL20p functions in cytoplasmic envelopment are separable from UL20 functions in UL20p intracellular transport, cell surface expression and virus-induced cell fusion.

  13. Chemical Models of Collapsing Envelopes

    CERN Document Server

    Bergin, E A

    1999-01-01

    We discuss recent models of chemical evolution in the developing and collapsing protostellar envelopes associated with low-mass star formation. In particular, the effects of depletion of gas-phase molecules onto grain surfaces is considered. We show that during the middle to late evolutionary stages, prior to the formation of a protostar, various species selectively deplete from the gas phase. The principal pattern of selective depletions is the depletion of sulfur-bearing molecules relative to nitrogen-bearing species: NH3 and N2H+. This pattern is shown to be insensitive to the details of the dynamics and marginally sensitive to whether the grain mantle is dominated by polar or non-polar molecules. Based on these results we suggest that molecular ions are good tracers of collapsing envelopes. The effects of coupling chemistry and dynamics on the resulting physical evolution are also examined. Particular attention is paid to comparisons between models and observations.

  14. Bacterial mitosis: partitioning protein ParA oscillates in spiral-shaped structures and positions plasmids at mid-cell

    DEFF Research Database (Denmark)

    Ebersbach, Gitte; Gerdes, Kenn; Charbon, Gitte Ebersbach

    2004-01-01

    with a single plasmid focus, the focus located preferentially at mid-cell. In cells with two foci, these located at quarter-cell positions. In the absence of ParB and parC1/parC2, ParA-GFP formed stationary helices extending from one end of the nucleoid to the other. In the presence of ParB and parC1/parC2, ParA-GFP......The par2 locus of Escherichia coli plasmid pB171 encodes oscillating ATPase ParA, DNA binding protein ParB and two cis-acting DNA regions to which ParB binds (parC1 and parC2). Three independent techniques were used to investigate the subcellular localization of plasmids carrying par2. In cells...... oscillated in spiral-shaped structures. Amino acid substitutions in ParA simultaneously abolished ParA spiral formation, oscillation and either plasmid localization or plasmid separation at mid-cell. Therefore, our results suggest that ParA spirals position plasmids at the middle of the bacterial nucleoid...

  15. Profiling of bacterial cells and cell-surface proteins of plant-associated bacteria by standard analytical techniques

    Czech Academy of Sciences Publication Activity Database

    Moravcová, Dana; Horká, Marie; Šalplachta, Jiří; Kubesová, Anna; Vykydalová, Marie; Kahle, Vladislav

    Latvia: Latvian Institute of Organic Synthesis, 2011 - (Kažoka, H.). s. 94 [Nordic Separation Science Society Conference /6./. 24.09.2011-27.09.2011, Riga] R&D Projects: GA AV ČR IAAX00310701; GA MV VG20112015021 Institutional research plan: CEZ:AV0Z40310501 Keywords : bacterial profiling * Rhizobium * MALDI-TOF MS Subject RIV: CB - Analytical Chemistry, Separation http://www.nosss.eu

  16. Data Envelopment Analysis: An Overview

    OpenAIRE

    Subhash C. Ray

    2014-01-01

    Over the past decades Data Envelopment Analysis (DEA) has emerged as an important nonparametric method of evaluating performance of decision making units through benchmarking. Although developed primarily for measuring technical efficiency, DEA is now applied extensively for measuring scale efficiency, cost efficiency, and profit efficiency as well. This paper integrates the different DEA models commonly applied in empirical research with their underlying theoretical foundations in neoclassic...

  17. An X-ray Absorption Fine Structure study of Au adsorbed onto the non-metabolizing cells of two soil bacterial species

    Energy Technology Data Exchange (ETDEWEB)

    Song, Zhen; Kenney, Janice P.L.; Fein, Jeremy B.; Bunker, Bruce A. (Notre)

    2015-02-09

    Gram-positive and Gram-negative bacterial cells can remove Au from Au(III)-chloride solutions, and the extent of removal is strongly pH dependent. In order to determine the removal mechanisms, X-ray Absorption Fine Structure (XAFS) spectroscopy experiments were conducted on non-metabolizing biomass of Bacillus subtilis and Pseudomonas putida with fixed Au(III) concentrations over a range of bacterial concentrations and pH values. X-ray Absorption Near Edge Structure (XANES) and Extended X-ray Absorption Fine Structure (EXAFS) data on both bacterial species indicate that more than 90% of the Au atoms on the bacterial cell walls were reduced to Au(I). In contrast to what has been observed for Au(III) interaction with metabolizing bacterial cells, no Au(0) or Au-Au nearest neighbors were observed in our experimental systems. All of the removed Au was present as adsorbed bacterial surface complexes. For both species, the XAFS data suggest that although Au-chloride-hydroxide aqueous complexes dominate the speciation of Au in solution, Au on the bacterial cell wall is characterized predominantly by binding of Au atoms to sulfhydryl functional groups and amine and/or carboxyl functional groups, and the relative importance of the sulfhydryl groups increases with increasing pH and with decreasing Au loading. The XAFS data for both microorganism species suggest that adsorption is the first step in the formation of Au nanoparticles by bacteria, and the results enhance our ability to account for the behavior of Au in bacteria-bearing geologic systems.

  18. Nitrate reducing bacterial activity in concrete cells of nuclear waste disposal

    International Nuclear Information System (INIS)

    Leaching experiments of solid matrices (bitumen and cement pastes) have been first implemented to define the physicochemical conditions that microorganisms are likely to meet at the bitumen-concrete interface (see the paper of Bertron et al.). Of course, as might be suspected, the cement matrix imposes highly alkaline pH conditions (10 < pH <11). The screening of a range of anaerobic denitrifying bacterial strains led us to select Halomonas desiderata as a model bacterium capable of catalyzing the reaction of nitrate reduction in these extreme conditions of pH. The denitrifying activity of Halomonas desiderata was quantified in batch bioreactor in the presence of solid matrices and / or leachate from bitumen and cement matrices. Denitrification was relatively fast in the presence of cement matrix (<100 hours) and 2 to 3 times slower in the presence of bituminous matrix. Overall, the presence of solid cement promoted the kinetics of denitrification. The observation of solid surfaces at the end of the experiment revealed the presence of a biofilm of Halomonas desiderata on the cement paste surface. These attached bacteria showed a denitrifying activity comparable to planktonic bacterial culture. On the other side, no colonization of bitumen could be highlighted as either by SEM or epifluorescence microscopy. Now, we are currently developing a continuous experimental bioreactor which should allow us a more rational understanding of the bitumen-cement-microbe interactions. (authors)

  19. Nitrate reducing bacterial activity in concrete cells of nuclear waste disposal

    Directory of Open Access Journals (Sweden)

    Albrecht A.

    2013-07-01

    Full Text Available Leaching experiments of solid matrices (bitumen and cement pastes have been first implemented to define the physicochemical conditions that microorganisms are likely to meet at the bitumen-concrete interface (see the paper of Bertron et al.. Of course, as might be suspected, the cement matrix imposes highly alkaline pH conditions (10 bacterial strains led us to select Halomonas desiderata as a model bacterium capable of catalyzing the reaction of nitrate reduction in these extreme conditions of pH. The denitrifying activity of Halomonas desiderata was quantified in batch bioreactor in the presence of solid matrices and / or leachate from bitumen and cement matrices. Denitrification was relatively fast in the presence of cement matrix (<100 hours and 2 to 3 times slower in the presence of bituminous matrix. Overall, the presence of solid cement promoted the kinetics of denitrification. The observation of solid surfaces at the end of the experiment revealed the presence of a biofilm of Halomonas desiderata on the cement paste surface. These attached bacteria showed a denitrifying activity comparable to planktonic bacterial culture. On the other side, no colonization of bitumen could be highlighted as either by SEM or epifluorescence microscopy. Now, we are currently developing a continuous experimental bioreactor which should allow us a more rational understanding of the bitumen-cement-microbe interactions.

  20. Bacterial Wall Components such as Lipothecoid Acid, Peptidoglycan, Liposaccharide and Lipid A Stimulate Cell Proliferation in Intestinal Epithelial Cells

    OpenAIRE

    Olaya, Jaime H.; Neopikhanov, Vadim; Söderman, Charlotte; Uribe, Andrés

    2011-01-01

    Earlier studies indicate that the microflora contains mitogens to intestinal epithelial cells. Our aim is to examine whether cell wall components of both Gram-negative and positive bacteria influence cell proliferation in small intestinal and colonic epithelial cells. A human colonic epithelial cell line from adenocarcinoma (IEC-6) and a nontransformed small intestinal cell line from germ-free rats (LS-123) were incubated with (a) lipothecoid acid from Streptococcus faecalis at 1.56–50 ...

  1. Moxifloxacin in Preventing Bacterial Infections in Patients Who Have Undergone Donor Stem Cell Transplant

    Science.gov (United States)

    2013-03-14

    Breast Cancer; Chronic Myeloproliferative Disorders; Gestational Trophoblastic Tumor; Infection; Leukemia; Lymphoma; Multiple Myeloma and Plasma Cell Neoplasm; Myelodysplastic Syndromes; Myelodysplastic/Myeloproliferative Neoplasms; Neuroblastoma; Ovarian Cancer; Testicular Germ Cell Tumor

  2. Differential survival of solitary and aggregated bacterial cells promotes aggregate formation on leaf surfaces

    Science.gov (United States)

    Monier, J.-M.; Lindow, S. E.

    2003-01-01

    The survival of individual Pseudomonas syringae cells was determined on bean leaf surfaces maintained under humid conditions or periodically exposed to desiccation stress. Cells of P. syringae strain B728a harboring a GFP marker gene were visualized by epifluorescence microscopy, either directly in situ or after recovery from leaves, and dead cells were identified as those that were stained with propidium iodide in such populations. Under moist, conducive conditions on plants, the proportion of total live cells was always high, irrespective of their aggregated state. In contrast, the proportion of the total cells that remained alive on leaves that were periodically exposed to desiccation stress decreased through time and was only ≈15% after 5 days. However, the fraction of cells in large aggregates that were alive on such plants in both condition was much higher than more solitary cells. Immediately after inoculation, cells were randomly distributed over the leaf surface and no aggregates were observed. However, a very aggregated pattern of colonization was apparent within 7 days, and >90% of the living cells were located in aggregates of 100 cells or more. Our results strongly suggest that, although conducive conditions favor aggregate formation, such cells are much more capable of tolerating environmental stresses, and the preferential survival of cells in aggregates promotes a highly clustered spatial distribution of bacteria on leaf surfaces. PMID:14665692

  3. Single-cell level based approach to investigate bacterial metabolism during batch industrial fermentation

    DEFF Research Database (Denmark)

    Nierychlo, Marta; Larsen, Poul; Eriksen, Niels T.;

    Escherichia coli fermentations have been studied for decades, but most results are based on average measurements of the whole populations of cells, whilst averaged data can mask the distribution of activities at the sub-population or single-cell level. A population of genetically identical cells ...

  4. Variations of both bacterial community and extracellular polymers: the inducements of increase of cell hydrophobicity from biofloc to aerobic granule sludge.

    Science.gov (United States)

    Guo, Feng; Zhang, Sheng-Hua; Yu, Xin; Wei, Bo

    2011-06-01

    To investigate the inducements of increase of cell hydrophobicity from aerobic biofloc (ABF) and granular sludge (AGS), in this study, as the first time the hydrophilic and hydrophobic bacterial communities were analyzed independently. Meanwhile, the effect of extracellular polymers (EPS) on the cell hydrophobicity is also studied. Few Bacteroidetes were detected (1.35% in ABF and 3.84% in AGS) in hydrophilic bacteria, whereas they are abundant in the hydrophobic cells (47.8% and 43% for ABF and AGS, respectively). The main species of Bacteroidetes changed from class Sphingobacteria to Flavobacteria in AGS. On the other hand, EPS is directly responsible to cell hydrophobicity. For AGS, cell hydrophobicity was sharply decreased after EPS extraction. Both quantity and property of the extracellular protein are related to hydrophobicity. Our results showed the variation of cell hydrophobicity was resulted from variations of both bacterial population and EPS. PMID:21482465

  5. Synchrony in human, mouse and bacterial cell cultures--a comparison

    Science.gov (United States)

    Helmstetter, Charles E.; Thornton, Maureen; Romero, Ana; Eward, K. Leigh

    2003-01-01

    Growth characteristics of synchronous human MOLT-4, human U-937 and mouse L1210 cultures produced with a new minimally-disturbing technology were compared to each other and to synchronous Escherichia coli B/r. Based on measurements of cell concentrations during synchronous growth, synchrony persisted in similar fashion for all cells. Cell size and DNA distributions in the mammalian cultures also progressed synchronously and reproducibly for multiple cell cycles. The results demonstrate that unambiguous multi-cycle synchrony, critical for verifying the absence of significant growth imbalances induced by the synchronization procedure, is feasible with these cell lines, and possibly others.

  6. Procalcitonin neutralizes bacterial LPS and reduces LPS-induced cytokine release in human peripheral blood mononuclear cells

    Directory of Open Access Journals (Sweden)

    Matera Giovanni

    2012-05-01

    Full Text Available Abstract Background Procalcitonin (PCT is a polypeptide with several cationic aminoacids in its chemical structure and it is a well known marker of sepsis. It is now emerging that PCT might exhibit some anti-inflammatory effects. The present study, based on the evaluation of the in vitro interaction between PCT and bacterial lipopolisaccharide (LPS, reports new data supporting the interesting and potentially useful anti-inflammatory activity of PCT. Results PCT significantly decreased (p Salmonella typhimurium (rough chemotype and Escherichia coli (smooth chemotype. Subsequently, the in vitro effects of PCT on LPS-induced cytokine release were studied in human peripheral blood mononuclear cells (PBMC. When LPS was pre-incubated for 30 minutes with different concentrations of PCT, the release of interleukin-10 (IL-10 and tumor necrosis factor alpha (TNFα by PBMC decreased in a concentration-dependent manner after 24 hours for IL-10 and 4 hours for TNFα. The release of monocyte chemotactic protein-1 (MCP-1 exhibited a drastic reduction at 4 hours for all the PCT concentrations assessed, whereas such decrease was concentration-dependent after 24 hours. Conclusions This study provides the first evidence of the capability of PCT to directly neutralize bacterial LPS, thus leading to a reduction of its major inflammatory mediators.

  7. Conformational instability governed by disulfide bonds partitions the dominant from subdominant helper T-cell responses specific for HIV-1 envelope glycoprotein gp120

    Science.gov (United States)

    Nguyen, Hong-Nam P.; Steede, N. Kalaya; Robinson, James E.; Landry, Samuel J.

    2015-01-01

    Most individuals infected with human immunodeficiency virus type 1 (HIV-1) generate a CD4+ T-cell response that is dominated by a few epitopes. Immunodominance may be counterproductive because a broad CD4+ T-cell response is associated with reduced viral load. Previous studies indicated that antigen three-dimensional structure controls antigen processing and presentation and therefore CD4+ T-cell epitope dominance. Dominant epitopes occur adjacent to the V1-V2, V3, and V4 loops because proteolytic antigen processing in the loops promotes presentation of adjacent sequences. In this study, three gp120 (strain JR-FL) variants were constructed, in which deletions of single outer-domain disulfide bonds were expected to introduce local conformational flexibility and promote presentation of additional CD4+ T-cell epitopes. Following mucosal immunization of C57BL/6 mice with wild-type or variant gp120 lacking the V3-flanking disulfide bond, the typical pattern of dominant epitopes was observed, suggesting that the disulfide bond posed no barrier to antigen presentation. In mice that lacked gamma interferoninducible lysosomal thioreductase (GILT), proliferative responses to the typically dominant epitopes of gp120 were selectively depressed, and the dominance pattern was rearranged. Deletion of the V3-flanking disulfide bond or one of the V4-flanking disulfide bonds partially restored highly proliferative responses to the typically dominant epitopes. These results reveal an acute dependence of dominant CD4+ T-cell responses on the native gp120 conformation. PMID:25944298

  8. Genomewide screening for fusogenic human endogenous retrovirus envelopes identifies syncytin 2, a gene conserved on primate evolution

    OpenAIRE

    Blaise, Sandra; de Parseval, Nathalie; Bénit, Laurence; Heidmann, Thierry

    2003-01-01

    Screening human sequence databases for endogenous retroviral elements with coding envelope genes has revealed 16 candidate genes that we assayed for their fusogenic properties. All 16 genes were cloned in a eukaryotic expression vector and assayed for cell–cell fusion by using a large panel of mammalian cells in transient transfection assays. Fusion was observed for two human endogenous retrovirus (HERV) envelopes, the previously characterized HERV-W envelope, also called syncytin, and a prev...

  9. Lung CD8+ T cells in COPD have increased expression of bacterial TLRs

    Directory of Open Access Journals (Sweden)

    Freeman Christine M

    2013-02-01

    Full Text Available Abstract Background Toll-like receptors (TLRs on T cells can modulate their responses, however, the extent and significance of TLR expression by lung T cells, NK cells, or NKT cells in chronic obstructive pulmonary disease (COPD is unknown. Methods Lung tissue collected from clinically-indicated resections (n = 34 was used either: (a to compare the expression of TLR1, TLR2, TLR2/1, TLR3, TLR4, TLR5, TLR6 and TLR9 on lung CD8+ T cells, CD4+ T cells, NK cells and NKT cells from smokers with or without COPD; or (b to isolate CD8+ T cells for culture with anti-CD3ε without or with various TLR ligands. We measured protein expression of IFN-γ, TNF-α, IL-13, perforin, granzyme A, granzyme B, soluble FasL, CCL2, CCL3, CCL4, CCL5, CCL11, and CXCL9 in supernatants. Results All the lung subsets analyzed demonstrated low levels of specific TLR expression, but the percentage of CD8+ T cells expressing TLR1, TLR2, TLR4, TLR6 and TLR2/1 was significantly increased in COPD subjects relative to those without COPD. In contrast, from the same subjects, only TLR2/1 and TLR2 on lung CD4+ T cells and CD8+ NKT cells, respectively, showed a significant increase in COPD and there was no difference in TLR expression on lung CD56+ NK cells. Production of the Tc1 cytokines IFN-γ and TNF-α by lung CD8+ T cells were significantly increased via co-stimulation by Pam3CSK4, a specific TLR2/1 ligand, but not by other agonists. Furthermore, this increase in cytokine production was specific to lung CD8+ T cells from patients with COPD as compared to lung CD8+ T cells from smokers without COPD. Conclusions These data suggest that as lung function worsens in COPD, the auto-aggressive behavior of lung CD8+ T cells could increase in response to microbial TLR ligands, specifically ligands against TLR2/1.

  10. Periodic growth of bacterial colonies

    Science.gov (United States)

    Yamazaki, Yoshihiro; Ikeda, Takemasa; Shimada, Hirotoshi; Hiramatsu, Fumiko; Kobayashi, Naoki; Wakita, Jun-ichi; Itoh, Hiroto; Kurosu, Sayuri; Nakatsuchi, Michio; Matsuyama, Tohey; Matsushita, Mitsugu

    2005-06-01

    The formation of concentric ring colonies by bacterial species Bacillus subtilis and Proteus mirabilis has been investigated experimentally, focusing our attention on the dependence of local cell density upon the bacterial motility. It has been confirmed that these concentric ring colonies reflect the periodic change of the bacterial motility between motile cell state and immotile cell state. We conclude that this periodic change is macroscopically determined neither by biological factors (i.e., biological clock) nor by chemical factors (chemotaxis as inhibitor). And our experimental results strongly suggest that the essential factor for the change of the bacterial motility during concentric ring formation is the local cell density.

  11. Dispersed cells represent a distinct stage in the transition from bacterial biofilm to planktonic lifestyles

    DEFF Research Database (Denmark)

    Chua, Song Lin; Liu, Yang; Yam, Joey Kuok Hoong;

    2014-01-01

    Bacteria assume distinct lifestyles during the planktonic and biofilm modes of growth. Increased levels of the intracellular messenger c-di-GMP determine the transition from planktonic to biofilm growth, while a reduction causes biofilm dispersal. It is generally assumed that cells dispersed from...... biofilms immediately go into the planktonic growth phase. Here we use single-nucleotide resolution transcriptomic analysis to show that the physiology of dispersed cells from Pseudomonas aeruginosa biofilms is highly different from those of planktonic and biofilm cells. In dispersed cells, the expression...... of the small regulatory RNAs RsmY and RsmZ is downregulated, whereas secretion genes are induced. Dispersed cells are highly virulent against macrophages and Caenorhabditis elegans compared with planktonic cells. In addition, they are highly sensitive towards iron stress, and the combination of a...

  12. Bacterial DNA activates cell mediated immune response and nitric oxide overproduction in peritoneal macrophages from patients with cirrhosis and ascites

    OpenAIRE

    Francés, R; Muñoz, C.; Zapater, P; Uceda, F; Gascón, I; Pascual, S.; Pérez-Mateo, M; J. Such

    2004-01-01

    Background and aims: Translocation of intestinal bacteria to ascitic fluid is probably the first step in the development of episodes of spontaneous bacterial peritonitis in patients with cirrhosis. We have recently reported the detection of bacterial DNA in blood and ascitic fluid from patients with advanced cirrhosis, what we consider as molecular evidence of bacterial translocation. Several studies have shown the immunogenic role of bacterial DNA in vitro, and we hypothesised that the prese...

  13. Microfluidic-Based Droplet and Cell Manipulations Using Artificial Bacterial Flagella

    Directory of Open Access Journals (Sweden)

    Yun Ding

    2016-02-01

    Full Text Available Herein, we assess the functionality of magnetic helical microswimmers as basic tools for the manipulation of soft materials, including microdroplets and single cells. Their ability to perform a range of unit operations is evaluated and the operational challenges associated with their use are established. In addition, we also report on interactions observed between the head of such helical swimmers and the boundaries of droplets and cells and discuss the possibilities of assembling an artificial swimming microorganism or a motorized cell.

  14. Tyrosine protein kinase inhibitors block invasin-promoted bacterial uptake by epithelial cells.

    OpenAIRE

    Rosenshine, I.; Duronio, V; Finlay, B B

    1992-01-01

    The ability to enter into (invade) mammalian cells is an essential virulence determinant of many pathogenic bacteria and intracellular parasites. These organisms are internalized by host cells upon attachment to their surface. However, the mechanisms used by intracellular parasites to induce internalization into host cells have not been defined. We found that the protein kinase inhibitor staurosporine blocks invasion by some pathogenic bacteria, including Yersinia enterocolitica and Yersinia ...

  15. Inhibition of Salmonella typhimurium Invasion by Host Cell Expression of Secreted Bacterial Invasion Proteins

    OpenAIRE

    Carlson, Steve A.; Jones, Bradley D.

    1998-01-01

    Pathogenic Salmonella species initiate infection of a host by inducing their own uptake into intestinal epithelial cells. An invasive phenotype is conferred to this pathogen by a number of proteins that are components of a type III secretion system. During the invasion process, the bacteria utilize this secretion system to release proteins that enter the host cell and apparently interact with unknown host cell components that induce alterations in the actin cytoskeleton. To investigate the ro...

  16. Bacterial cellulose-polyaniline nano-biocomposite: A porous media hydrogel bioanode enhancing the performance of microbial fuel cell

    Science.gov (United States)

    Mashkour, Mehrdad; Rahimnejad, Mostafa; Mashkour, Mahdi

    2016-09-01

    Microbial fuel cells (MFCs) are one of the possible renewable energy supplies which microorganisms play an active role in bio-oxidize reactions of a substrate such as glucose. Electrode materials and surface modifications are highly effective tools in enhancing MFCs' Performance. In this study, new composite anodes are fabricated. Bacterial cellulose (BC) is used as continuous phase and polyaniline (PANI) as dispersed one which is synthesized by in situ chemical oxidative polymerization on BC's fibers. With hydrogel nature of BC as a novel feature and polyaniline conductivity there meet the favorable conditions to obtain an active microbial biofilm on anode surface. Maximum power density of 117.76 mW/m2 in current density of 617 mA/m2 is achieved for BC/PANI anode. The amounts demonstrate a considerable enhancement compared with graphite plate (1 mW/m2 and 10 mA/m2).

  17. Intra-Species Bacterial Quorum Sensing Studied at Single Cell Level in a Double Droplet Trapping System

    Directory of Open Access Journals (Sweden)

    Wilhelm T. S. Huck

    2013-05-01

    Full Text Available In this paper, we investigated the intra-species bacterial quorum sensing at the single cell level using a double droplet trapping system. Escherichia coli transformed to express the quorum sensing receptor protein, LasR, were encapsulated in microdroplets that were positioned adjacent to microdroplets containing the autoinducer, N-(3-oxododecanoyl-L-homoserine lactone (OdDHL. Functional activation of the LasR protein by diffusion of the OdDHL across the droplet interface was measured by monitoring the expression of green fluorescent protein (GFP from a LasR-dependent promoter. A threshold concentration of OdDHL was found to induce production of quorum-sensing associated GFP by E. coli. Additionally, we demonstrated that LasR-dependent activation of GFP expression was also initiated when the adjacent droplets contained single E. coli transformed with the OdDHL synthase gene, LasI, representing a simple quorum sensing circuit between two droplets.

  18. Enhanced production of murine interferon gamma by T cells generated in response to bacterial infection

    OpenAIRE

    1982-01-01

    Spleen cell cultures derived from animals infected 6 d earlier with Listeria monocytogenes produced 10-20-fold more murine interferon gamma (MuIFN gamma) than spleen cells from nonimmune mice in response to stimulation with T cell mitogens. A striking temporal association was found between the enhanced synthesis of MuIFN gamma and the development of anti-Listeria immunity in that both the potential for increased MuIFN gamma production and the generation of Listeria-protective T cells develope...

  19. Cellular and molecular remodelling of a host cell for vertical transmission of bacterial symbionts

    Science.gov (United States)

    Luan, Jun-Bo; Shan, Hong-Wei; Isermann, Philipp; Huang, Jia-Hsin; Lammerding, Jan; Liu, Shu-Sheng; Douglas, Angela E.

    2016-01-01

    Various insects require intracellular bacteria that are restricted to specialized cells (bacteriocytes) and are transmitted vertically via the female ovary, but the transmission mechanisms are obscure. We hypothesized that, in the whitefly Bemisia tabaci, where intact bacteriocytes (and not isolated bacteria) are transferred to oocytes, the transmission mechanism would be evident as cellular and molecular differences between the nymph (pre-adult) and adult bacteriocytes. We demonstrate dramatic remodelling of bacteriocytes at the developmental transition from nymph to adulthood. This transition involves the loss of cell–cell adhesion, high division rates to constant cell size and onset of cell mobility, enabling the bacteriocytes to crawl to the ovaries. These changes are accompanied by cytoskeleton reorganization and changes in gene expression: genes functioning in cell–cell adhesion display reduced expression and genes involved in cell division, cell motility and endocytosis/exocytosis have elevated expression in adult bacteriocytes, relative to nymph bacteriocytes. This study demonstrates, for the first time, how developmentally orchestrated remodelling of gene expression and correlated changes in cell behaviour underpin the capacity of bacteriocytes to mediate the vertical transmission and persistence of the symbiotic bacteria on which the insect host depends. PMID:27358364

  20. A Stochastic Single-Molecule Event Triggers Phenotype Switching of a Bacterial Cell

    Science.gov (United States)

    Xie, Sunney; Choi, Paul; Cai, Long

    2009-03-01

    By monitoring fluorescently labeled lactose permease with single-molecule sensitivity, we investigated the molecular mechanism of how an Escherichia coli cell with the lac operon switches from one phenotype to another. At intermediate inducer concentrations, a population of genetically identical cells exhibits two phenotypes: induced cells with highly fluorescent membranes and uninduced cells with a small number of membrane-bound permeases. We found that this basal-level expression results from partial dissociation of the tetrameric lactose repressor from one of its operators on looped DNA. In contrast, infrequent events of complete dissociation of the repressor from DNA result in large bursts of permease expression that trigger induction of the lac operon. Hence, a stochastic single-molecule event determines a cell's phenotype.