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Sample records for high-copy t7 escherichia

  1. Augmentation of protein production by a combination of the T7 RNA polymerase system and ubiquitin fusion: Overproduction of the human DNA repair protein, ERCC1, as a ubiquitin fusion protein in Escherichia coli.

    NARCIS (Netherlands)

    M.H.M. Koken (Marcel); J.H. Odijk; M. van Duin (Mark); M.W.J. Fornerod (Maarten); D. Bootsma (Dirk); J.H.J. Hoeijmakers (Jan)

    1993-01-01

    textabstractThis article presents the development of a set of new expression vectors for overproduction of proteins in Escherichia coli. The vectors, pETUBI-ES1, 2 and 3, allow in-frame cloning of any sequence with the ubiquitin gene driven by the strong T7f10 promoter. Combination of the T7

  2. Expression of the N2 fixation gene operon of Paenibacillus sp. WLY78 under the control of the T7 promoter in Escherichia coli BL21.

    Science.gov (United States)

    Zhang, Lihong; Liu, Xiaomeng; Li, Xinxin; Chen, Sanfeng

    2015-10-01

    To investigate the transcription and translation and nitrogenase activity of the nine N2-fixing-gene (nif) operon (nifBHDKENXhesAnifX) of Paenibacillus sp. WLY78 under the control of the T7 promoter in Escherichia coli BL21 under different conditions. The Paenibacillus nif operon under the control of the T7 promoter is significantly transcribed and effectively translated in E. coli BL21 when grown in medium containing organic N compounds (yeast extract and Tryptone) or NH4+ by using RT-PCR and Western blot analysis. Transcription and translation of foreign nif genes in E. coli are not inhibited by environmental organic or inorganic N compounds or O2. However, contrary to transcription and translation, nitrogenase activity is 4% lower in the recombinant E. coli 78-32 compared to the native Paenibacillus sp. WLY78. The Paenibacillus nif operon under the control of T7 promoter enables E. coli BL21 to synthesize active nitrogenase. This study shows how the nif gene operon can be transferred to non-N2-fixing bacteria or to eukaryotic organelles.

  3. Melibiose permease and alpha-galactosidase of Escherichia coli: Identification by selective labeling using a T7 RNA polymerase/promoter expression system

    International Nuclear Information System (INIS)

    Pourcher, T.; Bassilana, M.; Sarkar, H.K.; Kaback, H.R.; Leblanc, G.

    1990-01-01

    Identification and selective labeling of the melibiose permease and alpha-galactosidase in Escherichia coli, which are encoded by the melB and melA genes, respectively, have been accomplished by selectively labeling the two gene products with a T7 RNA polymerase expression system. Following generation of a novel EcoRI restriction site in the intergenic sequence between the two genes of the mel operon by oligonucleotide-directed, site-specific mutagenesis, melA and melB were separately inserted into plasmid pT7-6 of the T7 expression system. Expression of melB was markedly enhanced by placing a strong, synthetic ribosome binding site at an optimal distance upstream from the initiation codon of melB. Expression of cloned gene products was characterized functionally and by performing autoradiographic analysis on total cell, inner membrane, and cytoplasmic proteins from cells pulse labeled with (35S)methionine in the presence of rifampicin and resolved by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The results first confirm that alpha-galactosidase is a cytoplasmic protein with an Mr of 50K; in contrast, the membrane-bound melibiose permease is identified as a protein with an apparent Mr of 39K, a value significantly higher than that of 30K previously suggested

  4. On the efficient bio-incorporation of 5-hydroxy-tryptophan in recombinant proteins expressed in Escherichia coli with T7 RNA polymerase-based vectors.

    Science.gov (United States)

    Oliveira-Souza, Wellington P; Bronze, Fellipe; Broos, Jaap; Marcondes, Marcelo F M; Oliveira, Vitor

    2017-10-21

    Biosynthetic incorporation of non-canonic amino acids is an attractive strategy to introduce new properties in recombinant proteins. Trp analogs can be incorporated in recombinant proteins replacing regular Trp during protein translation into a Trp-auxotrophic cell host. This straightforward method however, is limited to few analogs recognized and accepted by the cellular protein production machinery. 5-hydroxy-tryptophan (5OH-Trp) can be bio-incorporated using E. coli as expression host however; we have experienced very low incorporation yields - amount of protein containing regular Trp/amount of protein containing the Trp analog - during expressions of 5OH-Trp labeled proteins. Furthermore, this low incorporation yield were verified especially when the widely-used vectors based on the T7 RNA polymerase were used. Testing different 5OH-Trp incorporation protocols we verified that in these T7-based systems, the production of the T7 RNA polymerase is driven by the same elements - lac promoter/IPTG - as the target protein. Consequently, the bio-incorporation of the 5OH-Trp residues also occurs in this crucial enzyme, but, the produced T7 RNA polymerase labeled with 5OH-Trp is inactive or much less active. In the present work, we describe an efficient method to overcome this mentioned problem and bio-incorporate 5OH-Trp in proteins expressed in E. coli., using vectors based on the T7 RNA polymerase-T7 promoter. The two-step induction protocol here described showed incorporation efficiencies of 5OH-Trp higher than 90%. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Synthesis of bacteriophage-coded gene products during infection of Escherichia coli with amber mutants of T3 and T7 defective in gene 1

    DEFF Research Database (Denmark)

    Issinger, O G; Hausmann, R

    1973-01-01

    During nonpermissive infection by a T7 amber mutant in gene 1 (phage RNA polymerase-deficient), synthesis of the products of the phage genes 3 (endonuclease), 3, 5 (lysozyme), 5 (DNA polymerase), and 17 (serum blocking power) was shown to occur at about half the rate as during wild-type infection...

  6. Visualized and precise design of artificial small RNAs for regulating T7 RNA polymerase and enhancing recombinant protein folding in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Yujia Zhao

    2016-12-01

    Full Text Available Small non-coding RNAs (sRNAs have received much attention in recent years due to their unique biological properties, which can efficiently and specifically tune target gene expressions in bacteria. Inspired by natural sRNAs, recent works have proposed the use of artificial sRNAs (asRNAs as genetic tools to regulate desired gene that has been applied in several fields, such as metabolic engineering and bacterial physiology studies. However, the rational design of asRNAs is still a challenge. In this study, we proposed structure and length as two criteria to implement rational visualized and precise design of asRNAs. T7 expression system was one of the most useful recombinant protein expression systems. However, it was deeply limited by the formation of inclusion body. To settle this problem, we designed a series of asRNAs to inhibit the T7 RNA polymerase (Gene1 expression to balance the rate between transcription and folding of recombinant protein. Based on the heterologous expression of Aspergillus oryzae Li-3 glucuronidase in E. coli, the asRNA-antigene1-17bp can effectively decrease the inclusion body and increase the enzyme activity by 169.9%.

  7. Involvement of DNA gyrase in replication and transcription of bacteriophage T7 DNA

    International Nuclear Information System (INIS)

    De Wyngaert, M.A.; Hinkle, D.C.

    1979-01-01

    Growth of bacteriophage T7 is inhibited by the antibiotic coumermycin A 1 , an inhibitor of the Escherichia coli DNA gyrase. Since growth of the phage is insensitive to the antibiotic in strains containing a coumermycin-resistent DNA gyrase, this enzyme appears to be required for phage growth. We have investigated the effect of coumermycin on the kinetics of DNA, RNA, and protein synthesis during T7 infection. DNA synthesis is completely inhibited by the antibiotic. In addition, coumermycin significantly inhibits transcription of late but not early genes. Thus, E. coli DNA gyrase may play an important role in transcription as well as in replication of T7 DNA

  8. Dynamic Blue Light-Inducible T7 RNA Polymerases (Opto-T7RNAPs) for Precise Spatiotemporal Gene Expression Control.

    Science.gov (United States)

    Baumschlager, Armin; Aoki, Stephanie K; Khammash, Mustafa

    2017-11-17

    Light has emerged as a control input for biological systems due to its precise spatiotemporal resolution. The limited toolset for light control in bacteria motivated us to develop a light-inducible transcription system that is independent from cellular regulation through the use of an orthogonal RNA polymerase. Here, we present our engineered blue light-responsive T7 RNA polymerases (Opto-T7RNAPs) that show properties such as low leakiness of gene expression in the dark state, high expression strength when induced with blue light, and an inducible range of more than 300-fold. Following optimization of the system to reduce expression variability, we created a variant that returns to the inactive dark state within minutes once the blue light is turned off. This allows for precise dynamic control of gene expression, which is a key aspect for most applications using optogenetic regulation. The regulators, which only require blue light from ordinary light-emitting diodes for induction, were developed and tested in the bacterium Escherichia coli, which is a crucial cell factory for biotechnology due to its fast and inexpensive cultivation and well understood physiology and genetics. Opto-T7RNAP, with minor alterations, should be extendable to other bacterial species as well as eukaryotes such as mammalian cells and yeast in which the T7 RNA polymerase and the light-inducible Vivid regulator have been shown to be functional. We anticipate that our approach will expand the applicability of using light as an inducer for gene expression independent from cellular regulation and allow for a more reliable dynamic control of synthetic and natural gene networks.

  9. Library of synthetic transcriptional AND gates built with split T7 RNA polymerase mutants.

    Science.gov (United States)

    Shis, David L; Bennett, Matthew R

    2013-03-26

    The construction of synthetic gene circuits relies on our ability to engineer regulatory architectures that are orthogonal to the host's native regulatory pathways. However, as synthetic gene circuits become larger and more complicated, we are limited by the small number of parts, especially transcription factors, that work well in the context of the circuit. The current repertoire of transcription factors consists of a limited selection of activators and repressors, making the implementation of transcriptional logic a complicated and component-intensive process. To address this, we modified bacteriophage T7 RNA polymerase (T7 RNAP) to create a library of transcriptional AND gates for use in Escherichia coli by first splitting the protein and then mutating the DNA recognition domain of the C-terminal fragment to alter its promoter specificity. We first demonstrate that split T7 RNAP is active in vivo and compare it with full-length enzyme. We then create a library of mutant split T7 RNAPs that have a range of activities when used in combination with a complimentary set of altered T7-specific promoters. Finally, we assay the two-input function of both wild-type and mutant split T7 RNAPs and find that regulated expression of the N- and C-terminal fragments of the split T7 RNAPs creates AND logic in each case. This work demonstrates that mutant split T7 RNAP can be used as a transcriptional AND gate and introduces a unique library of components for use in synthetic gene circuits.

  10. Radiosensitivity of the induction of early enzymes by. gamma. -irradiated T7-phages

    Energy Technology Data Exchange (ETDEWEB)

    Bopp, E

    1975-01-01

    The radiosensitivity of the ability of the bacteriophage T7 to produce polymerase and lysozyme during its reproduction cycle is investigated. B-cells of Escherichia coli were infected with /sup 60/Co-..gamma..-irradiated T7 phages. From the extracts of the cells opened by ultrasonic waves, the amount of enzymes produced is determined with the aid of special enzyme tests. The fraction of inactivated phages able to produce RNA polymerase is higher than the fraction with intact DNA double strands and higher than the fraction able to inject DNA. The lowest fraction is that of inactivated phages producing lysozyme.

  11. Radiation inactivation of T7 phage

    International Nuclear Information System (INIS)

    Becker, D.; Redpath, J.L.; Grossweiner, L.I.

    1978-01-01

    The radiation inactivation of T7 phage by 25-MeV electron pulses has been measured in various media containing a wide concentration range of radical scavenging solutes and in the presence of protective and sensitizing agents. The dependence of sensitivity on pulse dose, from 1 mrad to 3.6 krad, is attributed to radical depletion via bimolecular processes. The survival data are analyzed by extending target theory to include diffusive reactions of primary and secondary radicals generated in the medium. It is concluded that OH radicals are the principal primary inactivating species and that secondary radicals from Br - , CNS - , uracil, glucose, ribose, sucrose, tyrosine, and histidine are lethal to some extent. In nutrient broth or 100 mM histidine, psoralen derivatives, Actinomycin D, and Mitomycin C are anoxic sensitizers. It is proposed that the psoralens promote the formation of non-strand break lesions as the sensitization mechanism. The target theory based on diffusional kinetics is applicable to other systems including single cells

  12. Mechanisms for the initiation of bacteriophage T7 DNA replication

    International Nuclear Information System (INIS)

    Fuller, C.W.; Beauchamp, B.B.; Engler, M.J.; Lechner, R.L.; Matson, S.W.; Tabor, S.; White, J.H.; Richardson, C.C.

    1983-01-01

    Genetic analysis of bacteriophage T7 has shown that the products of phage genes 1, 2, 3, 4, 5, and 6 are required for phage DNA synthesis in vivo. T7 RNA polymerase is the translation product of gene 1. This RNA polymerase is required for transcription of most of the phage genome, including genes 2 through 6. T7 RNA polymerase promoters consist of a highly conserved 23-bp DNA sequence. There are 17 such promoters in the T7 DNA molecule, all of which direct transcription from the same strand of the DNA. 70 references, 11 figures

  13. Investigation of specific interactions between T7 promoter and T7 RNA polymerase by force spectroscopy using atomic force microscope.

    Science.gov (United States)

    Zhang, Xiaojuan; Yao, Zhixuan; Duan, Yanting; Zhang, Xiaomei; Shi, Jinsong; Xu, Zhenghong

    2018-01-11

    The specific recognition and binding of promoter and RNA polymerase is the first step of transcription initiation in bacteria and largely determines transcription activity. Therefore, direct analysis of the interaction between promoter and RNA polymerase in vitro may be a new strategy for promoter characterization, to avoid interference due to the cell's biophysical condition and other regulatory elements. In the present study, the specific interaction between T7 promoter and T7 RNA polymerase was studied as a model system using force spectroscopy based on atomic force microscope (AFM). The specific interaction between T7 promoter and T7 RNA polymerase was verified by control experiments, and the rupture force in this system was measured as 307.2 ± 6.7 pN. The binding between T7 promoter mutants with various promoter activities and T7 RNA polymerase was analyzed. Interaction information including rupture force, rupture distance and binding percentage were obtained in vitro , and reporter gene expression regulated by these promoters was also measured according to a traditional promoter activity characterization method in vivo Using correlation analysis, it was found that the promoter strength characterized by reporter gene expression was closely correlated with rupture force and the binding percentage by force spectroscopy. These results indicated that the analysis of the interaction between promoter and RNA polymerase using AFM-based force spectroscopy was an effective and valid approach for the quantitative characterization of promoters. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  14. Intragenomic polymorphisms among high-copy loci: a genus-wide study of nuclear ribosomal DNA in Asclepias (Apocynaceae).

    Science.gov (United States)

    Weitemier, Kevin; Straub, Shannon C K; Fishbein, Mark; Liston, Aaron

    2015-01-01

    Despite knowledge that concerted evolution of high-copy loci is often imperfect, studies that investigate the extent of intragenomic polymorphisms and comparisons across a large number of species are rarely made. We present a bioinformatic pipeline for characterizing polymorphisms within an individual among copies of a high-copy locus. Results are presented for nuclear ribosomal DNA (nrDNA) across the milkweed genus, Asclepias. The 18S-26S portion of the nrDNA cistron of Asclepias syriaca served as a reference for assembly of the region from 124 samples representing 90 species of Asclepias. Reads were mapped back to each individual's consensus and at each position reads differing from the consensus were tallied using a custom perl script. Low frequency polymorphisms existed in all individuals (mean = 5.8%). Most nrDNA positions (91%) were polymorphic in at least one individual, with polymorphic sites being less frequent in subunit regions and loops. Highly polymorphic sites existed in each individual, with highest abundance in the "noncoding" ITS regions. Phylogenetic signal was present in the distribution of intragenomic polymorphisms across the genus. Intragenomic polymorphisms in nrDNA are common in Asclepias, being found at higher frequency than any other study to date. The high and variable frequency of polymorphisms across species highlights concerns that phylogenetic applications of nrDNA may be error-prone. The new analytical approach provided here is applicable to other taxa and other high-copy regions characterized by low coverage genome sequencing (genome skimming).

  15. Intragenomic polymorphisms among high-copy loci: a genus-wide study of nuclear ribosomal DNA in Asclepias (Apocynaceae

    Directory of Open Access Journals (Sweden)

    Kevin Weitemier

    2015-01-01

    Full Text Available Despite knowledge that concerted evolution of high-copy loci is often imperfect, studies that investigate the extent of intragenomic polymorphisms and comparisons across a large number of species are rarely made. We present a bioinformatic pipeline for characterizing polymorphisms within an individual among copies of a high-copy locus. Results are presented for nuclear ribosomal DNA (nrDNA across the milkweed genus, Asclepias. The 18S-26S portion of the nrDNA cistron of Asclepias syriaca served as a reference for assembly of the region from 124 samples representing 90 species of Asclepias. Reads were mapped back to each individual’s consensus and at each position reads differing from the consensus were tallied using a custom perl script. Low frequency polymorphisms existed in all individuals (mean = 5.8%. Most nrDNA positions (91% were polymorphic in at least one individual, with polymorphic sites being less frequent in subunit regions and loops. Highly polymorphic sites existed in each individual, with highest abundance in the “noncoding” ITS regions. Phylogenetic signal was present in the distribution of intragenomic polymorphisms across the genus. Intragenomic polymorphisms in nrDNA are common in Asclepias, being found at higher frequency than any other study to date. The high and variable frequency of polymorphisms across species highlights concerns that phylogenetic applications of nrDNA may be error-prone. The new analytical approach provided here is applicable to other taxa and other high-copy regions characterized by low coverage genome sequencing (genome skimming.

  16. Cloning and expression of the Escherichia coli K-12 sad gene.

    OpenAIRE

    Marek, L E; Henson, J M

    1988-01-01

    The Escherichia coli K-12 sad gene, which encodes an NAD-dependent succinic semialdehyde dehydrogenase, was cloned into a high-copy-number vector. Minicells carrying a sad+ plasmid produced a 55,000-dalton peptide, the probable sad gene product.

  17. Specific structural probing of plasmid-coded ribosomal RNAs from Escherichia coli

    DEFF Research Database (Denmark)

    Aagaard, C; Rosendahl, G; Dam, M

    1991-01-01

    The preferred method for construction and in vivo expression of mutagenised Escherichia coli ribosomal RNAs (rRNAs) is via high copy number plasmids. Transcription of wild-type rRNA from the seven chromosomal rrn operons in strains harbouring plasmid-coded mutant rRNAs leads to a heterogeneous...

  18. Two mechanisms coordinate replication termination by the Escherichia coli Tus–Ter complex

    KAUST Repository

    Pandey, Manjula; Elshenawy, Mohamed; Jergic, Slobodan; Takahashi, Masateru; Dixon, Nicholas E.; Hamdan, Samir; Patel, Smita S.

    2015-01-01

    The Escherichia coli replication terminator protein (Tus) binds to Ter sequences to block replication forks approaching from one direction. Here, we used single molecule and transient state kinetics to study responses of the heterologous phage T7

  19. Thioredoxin from Escherichia coli

    International Nuclear Information System (INIS)

    Holmgren, A.; Ohlsson, I.; Grankvist, M.L.

    1978-01-01

    A competition radioimmunoassay for Escherichia coli thioredoxin using 125 I-labeled thioredoxin-S 2 and a double antibody technique was developed. The method permits determination of picomole amounts of thioredoxin in crude cell extracts and was used to study the localization of thioredoxin cell fractions. E. coli B was calculated to have approximately 10,000 copies of thioredoxin per cell mainly located in the soluble fraction after separation of the membrane and soluble fractions by gentle lysis and centrifugation. E. coli B tsnC mutants which are defective in the replication of phage T7 DNA in vivo and in vitro were examined for their content of thioredoxin. E. coli B tsnC 7004 contained no detectable level of thioredoxin in cell-free extracts examined under a variety of conditions. The results strongly suggest that tsnC 7004 is a nonsense or deletion mutant. Two other E. coli tsnC mutants, 7007 and 7008, contained detectable levels of thioredoxin in crude extracts as measured by thioredoxin reductase and gave similar immunoprecipitation reactions as the parent strain B/1. By radioimmunoassay incompletely cross-reacting material was present in both strains. These results show that tsnC 7007 and 7008 belong to a type of thioredoxin mutants with missence mutations in the thioredoxin gene affecting the function of thioredoxin as subunit in phage T7 DNA polymerase

  20. Galectin-1 as a fusion partner for the production of soluble and folded human {beta}-1,4-galactosyltransferase-T7 in E. coli

    Energy Technology Data Exchange (ETDEWEB)

    Pasek, Marta [Structural Glycobiology Section, SAIC-Frederick, Inc., Center for Cancer Research Nanobiology Program, Center for Cancer Research, NCI-Frederick, Frederick, MD 2170 (United States); Boeggeman, Elizabeth; Ramakrishnan, Boopathy [Structural Glycobiology Section, SAIC-Frederick, Inc., Center for Cancer Research Nanobiology Program, Center for Cancer Research, NCI-Frederick, Frederick, MD 2170 (United States); Basic Science Program, SAIC-Frederick, Inc., Center for Cancer Research Nanobiology Program, Center for Cancer Research, NCI-Frederick, Frederick, MD 2170 (United States); Qasba, Pradman K., E-mail: qasba@helix.nih.gov [Structural Glycobiology Section, SAIC-Frederick, Inc., Center for Cancer Research Nanobiology Program, Center for Cancer Research, NCI-Frederick, Frederick, MD 2170 (United States)

    2010-04-09

    The expression of recombinant proteins in Escherichia coli often leads to inactive aggregated proteins known as the inclusion bodies. To date, the best available tool has been the use of fusion tags, including the carbohydrate-binding protein; e.g., the maltose-binding protein (MBP) that enhances the solubility of recombinant proteins. However, none of these fusion tags work universally with every partner protein. We hypothesized that galectins, which are also carbohydrate-binding proteins, may help as fusion partners in folding the mammalian proteins in E. coli. Here we show for the first time that a small soluble lectin, human galectin-1, one member of a large galectin family, can function as a fusion partner to produce soluble folded recombinant human glycosyltransferase, {beta}-1,4-galactosyltransferase-7 ({beta}4Gal-T7), in E. coli. The enzyme {beta}4Gal-T7 transfers galactose to xylose during the synthesis of the tetrasaccharide linker sequence attached to a Ser residue of proteoglycans. Without a fusion partner, {beta}4Gal-T7 is expressed in E. coli as inclusion bodies. We have designed a new vector construct, pLgals1, from pET-23a that includes the sequence for human galectin-1, followed by the Tev protease cleavage site, a 6x His-coding sequence, and a multi-cloning site where a cloned gene is inserted. After lactose affinity column purification of galectin-1-{beta}4Gal-T7 fusion protein, the unique protease cleavage site allows the protein {beta}4Gal-T7 to be cleaved from galectin-1 that binds and elutes from UDP-agarose column. The eluted protein is enzymatically active, and shows CD spectra comparable to the folded {beta}4Gal-T1. The engineered galectin-1 vector could prove to be a valuable tool for expressing other proteins in E. coli.

  1. Escherichia Coli

    Science.gov (United States)

    Goodsell, David S.

    2009-01-01

    Diverse biological data may be used to create illustrations of molecules in their cellular context. I describe the scientific results that support a recent textbook illustration of an "Escherichia coli cell". The image magnifies a portion of the bacterium at one million times, showing the location and form of individual macromolecules. Results…

  2. Elucidating the pH-Dependent Structural Transition of T7 Bacteriophage Endolysin.

    Science.gov (United States)

    Sharma, Meenakshi; Kumar, Dinesh; Poluri, Krishna Mohan

    2016-08-23

    Bacteriophages are the most abundant and diverse biological entities on earth. Bacteriophage endolysins are unique peptidoglycan hydrolases and have huge potential as effective enzybiotics in various infectious models. T7 bacteriophage endolysin (T7L), also known as N-acetylmuramoyl-l-alanine amidase or T7 lysozyme, is a 17 kDa protein that lyses a range of Gram-negative bacteria by hydrolyzing the amide bond between N-acetylmuramoyl residues and the l-alanine of the peptidoglycan layer. Although the activity profiles of several of the T7 family members have been known for many years, the molecular basis for their pH-dependent differential activity is not clear. In this study, we explored the pH-induced structural, stability, and activity characteristics of T7L by applying a variety of biophysical techniques and protein nuclear magnetic resonance (NMR) spectroscopy. Our studies established a reversible structural transition of T7L below pH 6 and the formation of a partially denatured conformation at pH 3. This low-pH conformation is thermally stable and exposed its hydrophobic pockets. Further, NMR relaxation measurements and structural analysis unraveled that T7L is highly dynamic in its native state and a network of His residues are responsible for the observed pH-dependent conformational dynamics and transitions. As bacteriophage chimeric and engineered endolysins are being developed as novel therapeutics against multiple drug resistance pathogens, we believe that our results are of great help in designing these entities as broadband antimicrobial and/or antibacterial agents.

  3. Thioredoxin suppresses microscopic hopping of T7 DNA polymerase on duplex DNA

    NARCIS (Netherlands)

    Etson, Candice M.; Hamdan, Samir M.; Richardson, Charles C.; Oijen, Antoine M. van; Richardson, Charles C.

    2010-01-01

    The DNA polymerases involved in DNA replication achieve high processivity of nucleotide incorporation by forming a complex with processivity factors. A model system for replicative DNA polymerases, the bacteriophage T7 DNA polymerase (gp5), encoded by gene 5, forms a tight, 1:1 complex with

  4. Genome Sequence of Streptomyces viridosporus Strain T7A ATCC 39115, a Lignin-Degrading Actinomycete

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Jennifer R. [Brown University; Goodwin, Lynne A. [Los Alamos National Laboratory (LANL); Teshima, Hazuki [Los Alamos National Laboratory (LANL); Detter, J. Chris [U.S. Department of Energy, Joint Genome Institute; Tapia, Roxanne [Los Alamos National Laboratory (LANL); Han, Cliff [Los Alamos National Laboratory (LANL); Huntemann, Marcel [U.S. Department of Energy, Joint Genome Institute; Wei, Chia-Lin [Los Alamos National Laboratory (LANL); Han, James [U.S. Department of Energy, Joint Genome Institute; Chen, Amy [U.S. Department of Energy, Joint Genome Institute; Kyrpides, Nikos C [U.S. Department of Energy, Joint Genome Institute; Mavromatis, K [U.S. Department of Energy, Joint Genome Institute; Szeto, Ernest [U.S. Department of Energy, Joint Genome Institute; Markowitz, Victor [U.S. Department of Energy, Joint Genome Institute; Ivanova, N [U.S. Department of Energy, Joint Genome Institute; Mikhailova, Natalia [U.S. Department of Energy, Joint Genome Institute; Ovchinnikova, Galina [U.S. Department of Energy, Joint Genome Institute; Pagani, Ioanna [U.S. Department of Energy, Joint Genome Institute; Pati, Amrita [U.S. Department of Energy, Joint Genome Institute; Woyke, Tanja [U.S. Department of Energy, Joint Genome Institute; Pitluck, Sam [U.S. Department of Energy, Joint Genome Institute; Peters, Lin [U.S. Department of Energy, Joint Genome Institute; Nolan, Matt [U.S. Department of Energy, Joint Genome Institute; Land, Miriam L [ORNL; Sello, Jason K. [Brown University

    2013-01-01

    We announce the availability of the genome sequence of Streptomyces viridosporus strain T7A ATCC 39115, a plant biomass- degrading actinomycete. This bacterium is of special interest because of its capacity to degrade lignin, an underutilized compo- nent of plants in the context of bioenergy. It has a full complement of genes for plant biomass catabolism.

  5. A small-angle scattering study of bacteriophage T7 using synchrotron radiation

    International Nuclear Information System (INIS)

    Feigin, L.A.; Svergun, D.I.; Dembo, A.T.; Ronto, G.; Toth, K.

    1989-01-01

    Structure transitions in the bacterial virus T7, due to an environment of varying ionic strength, are investigated by means of synchrotron radiation small-angle scattering. Effects of radiation damages and kinetics of the structure transition are separated. Time dependencies of the structural parameters and distribution functions are obtained and characteristic features of the structure rearrangements are described. (orig.)

  6. Bacteriophage T7 structure according to the data of small-angle X-ray scattering

    Energy Technology Data Exchange (ETDEWEB)

    Rol' bin, Yu A; Svergun, D I; Feigin, L A; Gashpar, Sh; Ronto, D [AN SSSR, Moscow. Inst. Kristallografii

    1980-01-01

    An attempt is made to obtain complete data on the form, sizes, weight and hydration of the T7 bacteriophage cultivated on E.coli cells and the peculiarities of phage DNA structure using the method of small-angle scattering.

  7. Two modes of interaction of the single-stranded DNA-binding protein of bacteriophage T7 with the DNA polymerase-thioredoxin complex

    KAUST Repository

    Ghosh, Sharmistha; Hamdan, Samir; Richardson, Charles C.

    2010-01-01

    The DNA polymerase encoded by bacteriophage T7 has low processivity. Escherichia coli thioredoxin binds to a segment of 76 residues in the thumb subdomain of the polymerase and increases the processivity. The binding of thioredoxin leads to the formation of two basic loops, loops A and B, located within the thioredoxin-binding domain (TBD). Both loops interact with the acidic C terminus of the T7 helicase. A relatively weak electrostatic mode involves the C-terminal tail of the helicase and the TBD, whereas a high affinity interaction that does not involve the C-terminal tail occurs when the polymerase is in a polymerization mode. T7 gene 2.5 single-stranded DNA-binding protein (gp2.5) also has an acidic C-terminal tail. gp2.5 also has two modes of interaction with the polymerase, but both involve the C-terminal tail of gp2.5. An electrostatic interaction requires the basic residues in loops A and B, and gp2.5 binds to both loops with similar affinity as measured by surface plasmon resonance. When the polymerase is in a polymerization mode, the C terminus of gene 2.5 protein interacts with the polymerase in regions outside the TBD.gp2.5 increases the processivity of the polymerase-helicase complex during leading strand synthesis. When loop B of the TBD is altered, abortive DNA products are observed during leading strand synthesis. Loop B appears to play an important role in communication with the helicase and gp2.5, whereas loop A plays a stabilizing role in these interactions. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. Two modes of interaction of the single-stranded DNA-binding protein of bacteriophage T7 with the DNA polymerase-thioredoxin complex

    KAUST Repository

    Ghosh, Sharmistha

    2010-04-06

    The DNA polymerase encoded by bacteriophage T7 has low processivity. Escherichia coli thioredoxin binds to a segment of 76 residues in the thumb subdomain of the polymerase and increases the processivity. The binding of thioredoxin leads to the formation of two basic loops, loops A and B, located within the thioredoxin-binding domain (TBD). Both loops interact with the acidic C terminus of the T7 helicase. A relatively weak electrostatic mode involves the C-terminal tail of the helicase and the TBD, whereas a high affinity interaction that does not involve the C-terminal tail occurs when the polymerase is in a polymerization mode. T7 gene 2.5 single-stranded DNA-binding protein (gp2.5) also has an acidic C-terminal tail. gp2.5 also has two modes of interaction with the polymerase, but both involve the C-terminal tail of gp2.5. An electrostatic interaction requires the basic residues in loops A and B, and gp2.5 binds to both loops with similar affinity as measured by surface plasmon resonance. When the polymerase is in a polymerization mode, the C terminus of gene 2.5 protein interacts with the polymerase in regions outside the TBD.gp2.5 increases the processivity of the polymerase-helicase complex during leading strand synthesis. When loop B of the TBD is altered, abortive DNA products are observed during leading strand synthesis. Loop B appears to play an important role in communication with the helicase and gp2.5, whereas loop A plays a stabilizing role in these interactions. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Genomic characteristics of vB_PpaP_PP74, a T7-like Autographivirinae bacteriophage infecting a potato pathogen of the newly proposed species Pectobacterium parmentieri.

    Science.gov (United States)

    Kabanova, Anastasia; Shneider, Mikhail; Bugaeva, Eugenia; Ha, Vo Thi Ngoc; Miroshnikov, Kirill; Korzhenkov, Aleksei; Kulikov, Eugene; Toschakov, Stepan; Ignatov, Alexander; Miroshnikov, Konstantin

    2018-02-08

    Bacteriophage vB_PpaP_PP74 (PP74) is a novel virulent phage that infects members of the species Pectobacterium parmentieri, a newly established species of soft-rot-causing bacteria in the family Pectobacteriaceae, derived from potato-specific Pectobacterium wasabiae. vB_PpaP_PP74 was identified as a member of the family Podoviridae by transmission electron microscopy. The phage has a 39,790-bp dsDNA genome containing 50 open reading frames (ORFs). Because of the absence of genes encoding toxins or lysogeny factors, PP74 may be considered a candidate phage for pathogen biocontrol applications. The genome layout is similar to genomes of T7-like phages within the subfamily Autographivirinae, and therefore, functions can be attributed to most of ORFs. However, the closest nucleotide sequence homologs of phage PP74 are unclassified Escherichia phages. Based on phylogenetic analysis, vB_PpaP_PP74 is a sensu lato T7-like phage, but it forms a distant subgenus group together with homologous enterobacterial phages.

  10. Visualization of uncorrelated, tandem symmetry mismatches in the internal genome packaging apparatus of bacteriophage T7.

    Science.gov (United States)

    Guo, Fei; Liu, Zheng; Vago, Frank; Ren, Yue; Wu, Weimin; Wright, Elena T; Serwer, Philip; Jiang, Wen

    2013-04-23

    Motor-driven packaging of a dsDNA genome into a preformed protein capsid through a unique portal vertex is essential in the life cycle of a large number of dsDNA viruses. We have used single-particle electron cryomicroscopy to study the multilayer structure of the portal vertex of the bacteriophage T7 procapsid, the recipient of T7 DNA in packaging. A focused asymmetric reconstruction method was developed and applied to selectively resolve neighboring pairs of symmetry-mismatched layers of the portal vertex. However, structural features in all layers of the multilayer portal vertex could not be resolved simultaneously. Our results imply that layers with mismatched symmetries can join together in several different relative orientations, and that orientations at different interfaces assort independently to produce structural isomers, a process that we call combinatorial assembly isomerism. This isomerism explains rotational smearing in previously reported asymmetric reconstructions of the portal vertex of T7 and other bacteriophages. Combinatorial assembly isomerism may represent a new regime of structural biology in which globally varying structures assemble from a common set of components. Our reconstructions collectively validate previously proposed symmetries, compositions, and sequential order of T7 portal vertex layers, resolving in tandem the 5-fold gene product 10 (gp10) shell, 12-fold gp8 portal ring, and an internal core stack consisting of 12-fold gp14 adaptor ring, 8-fold bowl-shaped gp15, and 4-fold gp16 tip. We also found a small tilt of the core stack relative to the icosahedral fivefold axis and propose that this tilt assists DNA spooling without tangling during packaging.

  11. Specificity of binding to four-way junctions in DNA by bacteriophage T7 endonuclease I.

    OpenAIRE

    Parsons, C A; West, S C

    1990-01-01

    T7 endonuclease I binds specifically to four-way junctions in duplex DNA and promotes their resolution into linear duplexes. Under conditions in which the nuclease activity is blocked by the absence of divalent cations, the enzyme forms a distinct protein-DNA complex with the junction, as detected by gel retardation and filter binding assays. The formation of this complex is structure-specific and contrasts with the short-lived binding complexes formed on linear duplex DNA. The binding comple...

  12. Nucleotide Selectivity at a Preinsertion Checkpoint of T7 RNA Polymerase Transcription Elongation.

    Science.gov (United States)

    E, Chao; Duan, Baogen; Yu, Jin

    2017-04-20

    Nucleotide selection is crucial for transcription fidelity control, in particular, for viral T7 RNA polymerase (RNAP) lack of proofreading activity. It has been recognized that multiple kinetic checkpoints exist prior to full nucleotide incorporation. In this work, we implemented intensive atomistic molecular dynamics (MD) simulations to quantify how strong the nucleotide selection is at the initial checkpoint of an elongation cycle of T7 RNAP. The incoming nucleotides bind into a preinsertion site where a critical tyrosine residue locates nearby to assist the nucleotide selection. We calculated the relative binding free energy between a noncognate nucleotide and a cognate one at a preinsertion configuration via alchemical simulations, showing that a small selection free energy or the binding free energy difference (∼3 k B T) exists between the two nucleotides. Indeed, another preinsertion configuration favored by the noncognate nucleotides was identified, which appears to be off path for further nucleotide insertion and additionally assists the nucleotide selection. By chemical master equation (CME) approach, we show that the small selection free energy at the preinsertion site along with the off-path noncognate nucleotide filtering can help substantially to reduce the error rate and to maintain the elongation rate high in the T7 RNAP transcription.

  13. Construction of a T7 Human Lung Cancer cDNA Library

    Directory of Open Access Journals (Sweden)

    Wentao YUE

    2008-10-01

    Full Text Available Background and objective Currently, only a limited numbers of tumor markers for non small lung cancer (NSCLC diagnosis, new biomarker, such as serum autoantibody may improve the early detection of lung cancer. Our objective is construction human lung squamous carcinoma and adenocarcinoma T7 phage display cDNA library from the tissues of NSCLC patients. Methods mRNA was isolated from a pool of total RNA extract from NSCLC tissues obtained from 5 adenocarcinomas and 5 squamous carcinomas, and then mRNA was reverse transcribed into double stranded cDNA. After digestion, the cDNA was inserted into T7Select 10-3 vector. The phage display cDNA library was constructed by package reaction in vitro and plate proliferation. Plaque assay and PCR were used to evaluate the library.Results Two T7 phage display cDNA library were established. Plaque assay show the titer of lung squamas carcinoma library was 1.8×106 pfu, and the adenocarcinoma library was 5×106 pfu. The phage titer of the amplified library were 3.2×1010 pfu/mL and 2.5×1010 pfu/mL. PCR amplification of random plaque show insert ratio were 100% (24/24 in adenocarcinoma library and 95.8% in human lung squamas carcinoma library (23/24. Insert range from 300 bp to 1 500 bp. Conclusion Two phage display cDNA library from NSCLC were constructed.

  14. In vitro recombination of bacteriophage T7 DNA damaged by uv radiation

    International Nuclear Information System (INIS)

    Masker, W.E.; Kuemmerle, N.B.

    1980-01-01

    A system capable of in vitro packaging of exogenous bacteriophage T7 DNA has been used to monitor the biological activity of DNA replicated in vitro. This system has been used to follow the effects of uv radiation on in vitro replication and recombination. During the in vitro replication process, a considerable exchange of genetic information occurs between T7 DNA molecules present in the reaction mixture. This in vitro recombination is reflected in the genotype of the T7 phage produced after in vitro encapsulation; depending on the genetic markers selected, recombinants can comprise nearly 20% of the total phage production. When uv-irradiated DNA is incubated in this system, the amount of in vitro synthesis is reduced and the total amount of viable phage produced after in vitro packaging is diminished. In vitro recombination rates are also lower when the participating DNA molecules have been exposed to uv. However, biochemical and genetic measurements confirmed that there is little or no transfer of pyrimidine dimers from irradiated DNA into undamaged molecules

  15. Assessment of the Effects of Various UV Sources on Inactivation and Photoproduct Induction in Phage T7 Dosimeter

    NARCIS (Netherlands)

    Fekete, A.; Vink, A.A.; Gaspar, S.; Berces, A.; Modos, K.; Ronto, Gy.; Roza, L.

    1998-01-01

    The correlation between the biologically effective dose (BED) of a phage T7 biological dosimeter and the induction of cyclobutane pyrimidine dimers (CPD) and (6-4) photoproducts ((6-4)PD) in the phage DNA was determined using seven various UV sources. The BED is the inactivation rate of phage T7

  16. Dark and photoreactivity of 4'-aminomethyl-4,5',8-trimethylpsoralen with T7 phage.

    Science.gov (United States)

    Tóth, K; Csik, G; Rontó, G

    1990-04-15

    The dark and photoreactions of 4'-aminomethyl-4,5',8-trimethylpsoralen (AMT) with T7 phage were investigated from biological and structural points of view. The dark reaction leads to the structural destabilization of the double helix of the DNA as is shown by optical melting measurements. The genotoxicity of AMT in the dark is comparable with that of known genotoxic drugs as determined by phage inactivation. The photoreaction with UVA light leads to the formation of mono- and di-adducts depending on the wavelength and dose used. Mono- and di-adducts influence DNA stability differently; biologically both types of adducts are genotoxic as measured by action spectra.

  17. A Novel Three-Colour Fluorescence in Situ Hybridization Approach for the Detection of t(7;12)(q36;p13) in Acute Myeloid Leukaemia Reveals New Cryptic Three Way Translocation t(7;12;16)

    Energy Technology Data Exchange (ETDEWEB)

    Naiel, Abdulbasit [Leukaemia and Chromosome Research Laboratory, Division of Biosciences, Brunel University, London, Middlesex UB8 3PH (United Kingdom); Vetter, Michael [MetaSystems, Altlussheim 68804 (Germany); Plekhanova, Olga [Regional Children’s Hospital N 1, Ekaterinburg 620149 (Russian Federation); Fleischman, Elena; Sokova, Olga [N.N. Blokhin Russian Cancer Research Center Russian Academy of Medical Science, Moscow 115478 (Russian Federation); Tsaur, Grigory [Regional Children’s Hospital N 1, Ekaterinburg 620149 (Russian Federation); Research Institute of Medical Cell Technologies, Ekaterinburg 620149 (Russian Federation); Harbott, Jochen [Oncogenetic Laboratory, Department of Paediatric Haematology and Oncology, Justus Liebig University, Giessen 35392 (Germany); Tosi, Sabrina, E-mail: sabrina.tosi@brunel.ac.uk [Leukaemia and Chromosome Research Laboratory, Division of Biosciences, Brunel University, London, Middlesex UB8 3PH (United Kingdom)

    2013-03-11

    The t(7;12)(q36;p13) translocation is a recurrent chromosome abnormality that involves the ETV6 gene on chromosome 12 and has been identified in 20–30% of infant patients with acute myeloid leukaemia (AML). The detection of t(7;12) rearrangements relies on the use of fluorescence in situ hybridization (FISH) because this translocation is hardly visible by chromosome banding methods. Furthermore, a fusion transcript HLXB9-ETV6 is found in approximately 50% of t(7;12) cases, making the reverse transcription PCR approach not an ideal screening method. Considering the report of few cases of variant translocations harbouring a cryptic t(7;12) rearrangement, we believe that the actual incidence of this abnormality is higher than reported to date. The clinical outcome of t(7;12) patients is believed to be poor, therefore an early and accurate diagnosis is important in the clinical management and treatment. In this study, we have designed and tested a novel three-colour FISH approach that enabled us not only to confirm the presence of the t(7;12) in a number of patients studied previously, but also to identify a cryptic t(7;12) as part of a complex rearrangement. This new approach has proven to be an efficient and reliable method to be used in the diagnostic setting.

  18. An intermediate state of T7 RNA polymerase provides another pathway of nucleotide selection

    International Nuclear Information System (INIS)

    Wang Zhan-Feng; Liu Yu-Ru; Wang Peng-Ye; Xie Ping

    2017-01-01

    Phage T7 RNA polymerase is a single-subunit transcription enzyme, transcribing template DNA to RNA. Nucleoside triphosphate (NTP) selection and translocation are two critical steps of the transcription elongation. Here, using all-atom molecular dynamics simulations, we found that between pre- and post-translocation states of T7 RNA polymerase an intermediate state exists, where the O helix C-terminal residue tyrosine 639, which plays important roles in translocation, locates between its pre- and post-translocation positions and the side chain of the next template DNA nucleotide has moved into the active site. NTP selection in this intermediate state was studied, revealing that the selection in the intermediate state can be achieved relying on the effect of Watson–Crick interaction between NTP and template DNA nucleotide, effect of stability of the components near the active site such as the nascent DNA–RNA hybrid and role of tyrosine 639. This indicates that another NTP-selection pathway can also exist besides the main pathway where NTP selection begins at the post-translocation state upon the entry of NTP. (paper)

  19. Bypass of a Nick by the Replisome of Bacteriophage T7*

    Science.gov (United States)

    Zhu, Bin; Lee, Seung-Joo; Richardson, Charles C.

    2011-01-01

    DNA polymerase and DNA helicase are essential components of DNA replication. The helicase unwinds duplex DNA to provide single-stranded templates for DNA synthesis by the DNA polymerase. In bacteriophage T7, movement of either the DNA helicase or the DNA polymerase alone terminates upon encountering a nick in duplex DNA. Using a minicircular DNA, we show that the helicase·polymerase complex can bypass a nick, albeit at reduced efficiency of 7%, on the non-template strand to continue rolling circle DNA synthesis. A gap in the non-template strand cannot be bypassed. The efficiency of bypass synthesis depends on the DNA sequence downstream of the nick. A nick on the template strand cannot be bypassed. Addition of T7 single-stranded DNA-binding protein to the complex stimulates nick bypass 2-fold. We propose that the association of helicase with the polymerase prevents dissociation of the helicase upon encountering a nick, allowing the helicase to continue unwinding of the duplex downstream of the nick. PMID:21701044

  20. Bypass of a nick by the replisome of bacteriophage T7.

    Science.gov (United States)

    Zhu, Bin; Lee, Seung-Joo; Richardson, Charles C

    2011-08-12

    DNA polymerase and DNA helicase are essential components of DNA replication. The helicase unwinds duplex DNA to provide single-stranded templates for DNA synthesis by the DNA polymerase. In bacteriophage T7, movement of either the DNA helicase or the DNA polymerase alone terminates upon encountering a nick in duplex DNA. Using a minicircular DNA, we show that the helicase · polymerase complex can bypass a nick, albeit at reduced efficiency of 7%, on the non-template strand to continue rolling circle DNA synthesis. A gap in the non-template strand cannot be bypassed. The efficiency of bypass synthesis depends on the DNA sequence downstream of the nick. A nick on the template strand cannot be bypassed. Addition of T7 single-stranded DNA-binding protein to the complex stimulates nick bypass 2-fold. We propose that the association of helicase with the polymerase prevents dissociation of the helicase upon encountering a nick, allowing the helicase to continue unwinding of the duplex downstream of the nick.

  1. Evaluation of the plasticity and damage parameters of aluminium 7075 T7

    International Nuclear Information System (INIS)

    Elgueta Vergara, Marcelo

    2004-01-01

    Mechanical damage occurs when, because of deformations, the initial crystalline network of a material is modified and suffers deterioration or progressive mechanical damage, until a macro crack occurs and then, the rupture or final fracture of the material. This mechanical damage is physically represented by the presence of cavities, decohesions, microfissures, etc. The mechanical damage, considered to be the progressive deterioration of the matter when submitted to deformations, is a phenomenon that, compared with the rupture of the material, has only recently begun to be studied. This work defines the parameters of plasticity, the laws of isotropic and kinematic hardening and the one dimensional law of mechanical damage of an aluminum alloy 7075 T7, widely used in the aeronautics industry. Measurements of deformations were taken with extensometry stamps, extensometries and a digital imagery correlation technique, in tests under a monotonous as well as a cyclic load (CW)

  2. High density growth of T7 expression strains with auto-induction option

    Energy Technology Data Exchange (ETDEWEB)

    Studier, F. William (Stony Brook, NY)

    2010-07-20

    A bacterial growth medium for promoting auto-induction of transcription of cloned DNA in cultures of bacterial cells grown batchwise is disclosed. The transcription is under the control of a lac repressor. Also disclosed is a bacterial growth medium for improving the production of a selenomethionine-containing protein or polypeptide in a bacterial cell, the protein or polypeptide being produced by recombinant DNA techniques from a lac or T7lac promoter, the bacterial cell encoding a vitamin B12-dependent homocysteine methylase. Finally, disclosed is a bacterial growth medium for suppressing auto-induction of expression in cultures of bacterial cells grown batchwise, said transcription being under the control of lac repressor.

  3. Catalyze and chemical inhibition of the R7T7 glass kinetics

    International Nuclear Information System (INIS)

    Gin, St.; Advocat, Th.

    1997-01-01

    This article highlights some phenomena likely to modify the glass alteration kinetics and/or the nature of the alteration products according to the chemical composition of the leaching water. It discusses experimental results showing that in neutral and basic media, the presence of inorganic anions or organic acid (simple carboxylic acid and humic acid) has relatively little effect on the long-term glass matrix alterability Actinide mobility appears to be more dependent on the concentration of complexing agents in the leaching solution. The behavior of phosphate ions, which may inhibit or catalyze the R7T7 glass alteration kinetics depending on the experimental conditions, is discussed; the gel microstructure can be related to the glass alteration kinetics by detailed examination of the alteration products. (authors)

  4. R7T7 glass alteration in the presence of mortar: effect of the cement grade

    International Nuclear Information System (INIS)

    Andriambololona, Z.; Godon, N.; Vernaz, E.

    1991-01-01

    R7T7 glass alteration was investigated in the presence of four mortars prepared from four different cement grades: 'CPA' Portland cement (mortar M1), CPA with pozzolana additive (M2), CPA with amorphous silica additive (M3) and 'CLK' blast furnace slag cement (M4). Glass specimens were also altered in Volvic mineral water and in a cement effluent. Glass corrosion in the cement media was greater than in Volvic water, but well below what could be expected from the high pH (approx 12.5). The relatively low alteration was probably related to the protective action of the calcium-enriched gel layer that formed at the glass surface. The glass corrosion rate was 2 to 3 times lower with cement containing pozzolana or silica gel additives or with CLK cement than with CPA cement alone. 8 refs., 8 figs

  5. Towards observing the encounter of the T7 DNA replication fork with a lesion site at the Single molecule level

    KAUST Repository

    Shirbini, Afnan

    2017-01-01

    and established the T7 leading strand synthesis at the single molecule level. I also optimized various control experiments to remove any interference from the nonspecific interactions of the DNA with the surface. My work established the foundation to image

  6. The Roles of Tidal Evolution and Evaporative Mass Loss in the Origin of CoRoT-7 b

    Science.gov (United States)

    Jackson, Brian; Miller, Neil; Barnes, Rory; Raymond, Sean N.; Fortney, Jonathan J.; Greenberg, Richard

    2010-01-01

    CoRoT-7 b is the first confirmed rocky exoplanet, but, with an orbital semimajor axis of 0.0172 au, its origins may be unlike any rocky planet in our Solar System. In this study, we consider the roles of tidal evolution and evaporative mass loss in CoRoT-7 b's history, which together have modified the planet's mass and orbit. If CoRoT-7 b has always been a rocky body, evaporation may have driven off almost half its original mass, but the mass loss may depend sensitively on the extent of tidal decay of its orbit. As tides caused CoRoT-7 b's orbit to decay, they brought the planet closer to its host star, thereby enhancing the mass loss rate. Such a large mass loss also suggests the possibility that CoRoT-7 b began as a gas giant planet and had its original atmosphere completely evaporated. In this case, we find that CoRoT-7 b's original mass probably did not exceed 200 Earth masses (about two-third of a Jupiter mass). Tides raised on the host star by the planet may have significantly reduced the orbital semimajor axis, perhaps causing the planet to migrate through mean-motion resonances with the other planet in the system, CoRoT-7 c. The coupling between tidal evolution and mass loss may be important not only for CoRoT-7 b but also for other close-in exoplanets, and future studies of mass loss and orbital evolution may provide insight into the origin and fate of close-in planets, both rocky and gaseous.

  7. Use of T7 RNA polymerase to direct expression of outer Surface Protein A (OspA) from the Lyme disease Spirochete, Borrelia burgdorferi

    Science.gov (United States)

    Dunn, John J.; Lade, Barbara N.

    1991-01-01

    The OspA gene from a North American strain of the Lyme disease Spirochete, Borrelia burgdorferi, was cloned under the control of transciption and translation signals from bacteriophage T7. Full-length OspA protein, a 273 amino acid (31kD) lipoprotein, is expressed poorly in Escherichia coli and is associated with the insoluble membrane fraction. In contrast, a truncated form of OspA lacking the amino-terminal signal sequence which normally would direct localization of the protein to the outer membrane is expressed at very high levels (less than or equal to 100 mg/liter) and is soluble. The truncated protein was purified to homogeneity and is being tested to see if it will be useful as an immunogen in a vaccine against Lyme disease. Circular dichroism and fluorescence spectroscopy was used to characterize the secondary structure and study conformational changes in the protein. Studies underway with other surface proteins from B burgdorferi and a related spirochete, B. hermsii, which causes relapsing fever, leads us to conclude that a strategy similar to that used to express the truncated OspA can provide a facile method for producing variations of Borrelia lipoproteins which are highly expressed in E. coli and soluble without exposure to detergents.

  8. Alteration of R7T7-type nuclear glass in deep geological storage conditions

    International Nuclear Information System (INIS)

    Combarieu, G. de

    2007-02-01

    This PhD thesis is aimed to study the alteration of SON68 glass, French inactive glass of R7T7-type, in contact with near field materials of a deep geological storage (French concept from ANDRA) which are mainly metallic iron and Callovo-Oxfordian clay. Therefore, experiments involving a 'glass-iron-clay' system at lab-scale have been carried out. Interactions between glass, iron and clay have been characterised from submicron to millimeter scale by means of SEM, TEM, XRD and XAS and Raman spectroscopies in terms of chemistry and crystal-chemistry. In the mean time, a conceptual model of glass alteration has been developed to account for most of the experimental observations and known mechanisms of alteration. The model has been then transposed within the transport-chemistry code HYTEC, together with developed models of clay and iron corrosion, to simulate the experiments described above. This work is thus a contribution to the understanding of iron corrosion in Callovo-Oxfordian clay and subsequent glass alteration in the newly formed corrosion products, the whole process being considered as a lab-scale model of a deep geological storage of radioactive wastes. (author)

  9. Mechanism of sequence-specific template binding by the DNA primase of bacteriophage T7

    KAUST Repository

    Lee, Seung-Joo

    2010-03-28

    DNA primases catalyze the synthesis of the oligoribonucleotides required for the initiation of lagging strand DNA synthesis. Biochemical studies have elucidated the mechanism for the sequence-specific synthesis of primers. However, the physical interactions of the primase with the DNA template to explain the basis of specificity have not been demonstrated. Using a combination of surface plasmon resonance and biochemical assays, we show that T7 DNA primase has only a slightly higher affinity for DNA containing the primase recognition sequence (5\\'-TGGTC-3\\') than for DNA lacking the recognition site. However, this binding is drastically enhanced by the presence of the cognate Nucleoside triphosphates (NTPs), Adenosine triphosphate (ATP) and Cytosine triphosphate (CTP) that are incorporated into the primer, pppACCA. Formation of the dimer, pppAC, the initial step of sequence-specific primer synthesis, is not sufficient for the stable binding. Preformed primers exhibit significantly less selective binding than that observed with ATP and CTP. Alterations in subdomains of the primase result in loss of selective DNA binding. We present a model in which conformational changes induced during primer synthesis facilitate contact between the zinc-binding domain and the polymerase domain. The Author(s) 2010. Published by Oxford University Press.

  10. Physicochemical properties and long-term behavior of french R7T7 nuclear waste glass

    International Nuclear Information System (INIS)

    Vernaz, E.

    1990-01-01

    The French R7T7 nuclear glass composition was carefully selected to allow incorporation of some thirty different oxides found in fission product solutions. The resulting glass exhibits very low crystallization, and its physical and chemical properties are very similar to those of standard industrial glasses. Nuclear glasses have been shown to withstand α doses corresponding to several hundred thousand years under repository conditions. Predicting the long-term behavior of fission product glasses subjected to aqueous corrosion is no doubt the most difficult aspect of the problem. Predictions are necessarily based on mathematical models. A substantial research effort has been undertaken to identify all the basic corrosion mechanisms liable to control long-term alteration. These mechanisms are now relatively well understood, and provide the basis for developing the indispensable models. Realistic storage conditions exist under which glass alteration occurs at a very slow rate, and can fulfill its role as the first containment barrier for several tens of thousands of years

  11. Altered minor-groove hydrogen bonds in DNA block transcription elongation by T7 RNA polymerase.

    Science.gov (United States)

    Tanasova, Marina; Goeldi, Silvan; Meyer, Fabian; Hanawalt, Philip C; Spivak, Graciela; Sturla, Shana J

    2015-05-26

    DNA transcription depends upon the highly efficient and selective function of RNA polymerases (RNAPs). Modifications in the template DNA can impact the progression of RNA synthesis, and a number of DNA adducts, as well as abasic sites, arrest or stall transcription. Nonetheless, data are needed to understand why certain modifications to the structure of DNA bases stall RNA polymerases while others are efficiently bypassed. In this study, we evaluate the impact that alterations in dNTP/rNTP base-pair geometry have on transcription. T7 RNA polymerase was used to study transcription over modified purines and pyrimidines with altered H-bonding capacities. The results suggest that introducing wobble base-pairs into the DNA:RNA heteroduplex interferes with transcriptional elongation and stalls RNA polymerase. However, transcriptional stalling is not observed if mismatched base-pairs do not H-bond. Together, these studies show that RNAP is able to discriminate mismatches resulting in wobble base-pairs, and suggest that, in cases of modifications with minor steric impact, DNA:RNA heteroduplex geometry could serve as a controlling factor for initiating transcription-coupled DNA repair. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. MIRO Observation of Comet C/2002 T7 (LINEAR) Water Line Spectrum

    Science.gov (United States)

    Lee, Seungwon; Frerking, Margaret; Hofstadter, Mark; Gulkis, Samuel; von Allmen, Paul; Crovisier, Jaques; Biver, Nicholas; Bockelee-Morvan, Dominique

    2011-01-01

    Comet C/2002 T7 (LINEAR) was observed with the Microwave Instrument for Rosetta Orbiter (MIRO) on April 30, 2004, between 5 hr and 16 hr UT. The comet was 0.63AU distance from the Sun and 0.68AU distance from the MIRO telescope at the time of the observations. The water line involving the two lowest rotational levels at 556.936 GHz is observed at 557.070 GHz due to a large Doppler frequency shift. The detected water line spectrum is interpreted using a non local thermal equilibrium (Non-LTE) molecular excitation and radiative transfer model. Several synthetic spectra are calculated with various coma profiles that are plausible for the comet at the time of observations. The coma profile is modeled with three characteristic parameters: outgassing rate, a constant expansion velocity, and a constant gas temperature. The model calculation result shows that for the distant line observation where contributions from a large coma space is averaged, the combination of the outgassing rate and the gas expansion velocity determines the line shape while the gas temperature has a negligible effect. The comparison between the calculated spectra and the MIRO measured spectrum suggests that the outgassing rate of the comet is about 2.0x1029 molecules/second and its gas expansion velocity about 1.2 km/s at the time of the observations.

  13. Alteration of 'R7T7' type nuclear glasses: statistical approach, experimental validation, local evolution model

    International Nuclear Information System (INIS)

    Thierry, F.

    2003-02-01

    The aim of this work is to propose an evolution of nuclear (R7T7-type) glass alteration modeling. The first part of this thesis is about development and validation of the 'r(t)' model. This model which predicts the decrease of alteration rates in confined conditions is based upon a coupling between a first-order dissolution law and a diffusion barrier effect of the alteration gel layer. The values and the uncertainties regarding the main adjustable parameters of the model (α, Dg and C*) have been determined from a systematic study of the available experimental data. A program called INVERSION has been written for this purpose. This work lead to characterize the validity domain of the 'r(t)' model and to parametrize it. Validation experiments have been undertaken, confirming the validity of the parametrization over 200 days. A new model is proposed in the second part of this thesis. It is based on an inhibition of glass dissolution reaction by silicon coupled with a local description of silicon retention in the alteration gel layer. This model predicts the evolutions of boron and silicon concentrations in solution as well as the concentrations and retention profiles in the gel layer. These predictions have been compared to measurements of retention profiles by the secondary ion mass spectrometry (SIMS) method. The model has been validated on fractions of gel layer which reactivity present low or moderate disparities. (author)

  14. C-terminal phenylalanine of bacteriophage T7 single-stranded DNA-binding protein is essential for strand displacement synthesis by T7 DNA polymerase at a nick in DNA.

    Science.gov (United States)

    Ghosh, Sharmistha; Marintcheva, Boriana; Takahashi, Masateru; Richardson, Charles C

    2009-10-30

    Single-stranded DNA-binding protein (gp2.5), encoded by gene 2.5 of bacteriophage T7, plays an essential role in DNA replication. Not only does it remove impediments of secondary structure in the DNA, it also modulates the activities of the other replication proteins. The acidic C-terminal tail of gp2.5, bearing a C-terminal phenylalanine, physically and functionally interacts with the helicase and DNA polymerase. Deletion of the phenylalanine or substitution with a nonaromatic amino acid gives rise to a dominant lethal phenotype, and the altered gp2.5 has reduced affinity for T7 DNA polymerase. Suppressors of the dominant lethal phenotype have led to the identification of mutations in gene 5 that encodes the T7 DNA polymerase. The altered residues in the polymerase are solvent-exposed and lie in regions that are adjacent to the bound DNA. gp2.5 lacking the C-terminal phenylalanine has a lower affinity for gp5-thioredoxin relative to the wild-type gp2.5, and this affinity is partially restored by the suppressor mutations in DNA polymerase. gp2.5 enables T7 DNA polymerase to catalyze strand displacement DNA synthesis at a nick in DNA. The resulting 5'-single-stranded DNA tail provides a loading site for T7 DNA helicase. gp2.5 lacking the C-terminal phenylalanine does not support this event with wild-type DNA polymerase but does to a limited extent with T7 DNA polymerase harboring the suppressor mutations.

  15. C-terminal Phenylalanine of Bacteriophage T7 Single-stranded DNA-binding Protein Is Essential for Strand Displacement Synthesis by T7 DNA Polymerase at a Nick in DNA*

    Science.gov (United States)

    Ghosh, Sharmistha; Marintcheva, Boriana; Takahashi, Masateru; Richardson, Charles C.

    2009-01-01

    Single-stranded DNA-binding protein (gp2.5), encoded by gene 2.5 of bacteriophage T7, plays an essential role in DNA replication. Not only does it remove impediments of secondary structure in the DNA, it also modulates the activities of the other replication proteins. The acidic C-terminal tail of gp2.5, bearing a C-terminal phenylalanine, physically and functionally interacts with the helicase and DNA polymerase. Deletion of the phenylalanine or substitution with a nonaromatic amino acid gives rise to a dominant lethal phenotype, and the altered gp2.5 has reduced affinity for T7 DNA polymerase. Suppressors of the dominant lethal phenotype have led to the identification of mutations in gene 5 that encodes the T7 DNA polymerase. The altered residues in the polymerase are solvent-exposed and lie in regions that are adjacent to the bound DNA. gp2.5 lacking the C-terminal phenylalanine has a lower affinity for gp5-thioredoxin relative to the wild-type gp2.5, and this affinity is partially restored by the suppressor mutations in DNA polymerase. gp2.5 enables T7 DNA polymerase to catalyze strand displacement DNA synthesis at a nick in DNA. The resulting 5′-single-stranded DNA tail provides a loading site for T7 DNA helicase. gp2.5 lacking the C-terminal phenylalanine does not support this event with wild-type DNA polymerase but does to a limited extent with T7 DNA polymerase harboring the suppressor mutations. PMID:19726688

  16. Development and application of a T7 RNA polymerase-dependent expression system for antibiotic production improvement in Streptomyces.

    Science.gov (United States)

    Wei, Junhong; Tian, Jinjin; Pan, Guoqing; Xie, Jie; Bao, Jialing; Zhou, Zeyang

    2017-06-01

    To develop a reliable and easy to use expression system for antibiotic production improvement of Streptomyces. A two-compound T7 RNA polymerase-dependent gene expression system was developed to fulfill this demand. In this system, the T7 RNA polymerase coding sequence was optimized based on the codon usage of Streptomyces coelicolor. To evaluate the functionality of this system, we constructed an activator gene overexpression strain for enhancement of actinorhodin production. By overexpression of the positive regulator actII-ORF4 with this system, the maximum actinorhodin yield of engineered strain was 15-fold higher and the fermentation time was decreased by 48 h. The modified two-compound T7 expression system improves both antibiotic production and accelerates the fermentation process in Streptomyces. This provides a general and useful strategy for strain improvement of important antibiotic producing Streptomyces strains.

  17. Aqueous corrosion of silicate glasses. Analogy between volcanic glasses and the French nuclear waste glass R7T7

    International Nuclear Information System (INIS)

    Goldschmidt, F.

    1991-01-01

    The behaviour of borosilicate glasses upon aqueous corrosion is controlled for long periods of time (>10,000 years) by processes which are not directly accessible by means of laboratory experiments. The analogical approach consists here to compare leaching performances between the french nuclear waste glass R7T7 and natural volcanic glasses, basaltic and rhyolitic ones. The three glasses were leached in the same conditions; open system, 90 deg C, initial pH of 9.7. Basaltic and R7T7 glasses having the same kinetic of dissolution, the basaltic glass was chosen as the best analogue. (author). refs., figs., tabs

  18. Early phyllosilicates formed by alteration of R7T7 glass in water at 250 deg C

    International Nuclear Information System (INIS)

    Vernaz, E.; Dussossoy, J.L.; Caurel, J.; Crovisier, J.L.

    1991-01-01

    T7T7 glass samples have been altered during periods of one and seven days at 250 deg C in distilled water. Ultramicrotomic thin sections were performed in the outermost part of the glass for a study under analytical transmission electron microscope. After one day the alteration layer is about 5 μm thick. It is made up of sheeted silicated particles, tentatively identified as septechlorite (7 A intervals) mixed with an intercrystalline amorphous matrix of different composition. After seven days the alteration layer is about 11 μm thick. It is made up of sheeted silicated particles of smectite type (10 A intervals). No 7 A particles were found in the seven days sample. This study illustrates the capacity of poorly crystalline phyllosilicates to adapt to the chemical changes in solution. 15 refs., 4 tabs., 2 plates

  19. R7T7-type HLW glass alteration under irradiation. Study of the residual alteration rate regime

    International Nuclear Information System (INIS)

    Rolland, Severine

    2012-01-01

    In France, fission products and minor actinides remaining after reprocessing of spent nuclear fuel are confined in a borosilicate glass matrix, named R7T7, for disposal in a geological repository. However, in these conditions, after several thousand years, water could arrive in contact with glass and be radio-lysed. In this work, we investigated the irradiation influence and especially the influence of the energy deposition on the residual glass alteration rate regime in pure water. Two types of leaching tests have been carried out. The first were performed on radioactive glass and the second on a SON68 glass (nonradioactive surrogate of R7T7 glass) under external irradiation γ. (author) [fr

  20. Towards observing the encounter of the T7 DNA replication fork with a lesion site at the Single molecule level

    KAUST Repository

    Shirbini, Afnan

    2017-05-01

    Single-molecule DNA flow-stretching assays have been a powerful approach to study various aspects on the mechanism of DNA replication for more than a decade. This technique depends on flow-induced force on a bead attached to a surface-tethered DNA. The difference in the elastic property between double-strand DNA (long) and single-strand DNA (short) at low regime force allows the observation of the beads motion when the dsDNA is converted to ssDNA by the replisome machinery during DNA replication. Here, I aim to develop an assay to track in real-time the encounter of the bacteriophage T7 replisome with abasic lesion site inserted on the leading strand template. I optimized methods to construct the DNA substrate that contains the abasic site and established the T7 leading strand synthesis at the single molecule level. I also optimized various control experiments to remove any interference from the nonspecific interactions of the DNA with the surface. My work established the foundation to image the encounter of the T7 replisome with abasic site and to characterize how the interactions between the helicase and the polymerase could influence the polymerase proofreading ability and its direct bypass of this highly common DNA damage type.

  1. Promoter binding, initiation, and elongation by bacteriophage T7 RNA polymerase. A single-molecule view of the transcription cycle.

    Science.gov (United States)

    Skinner, Gary M; Baumann, Christoph G; Quinn, Diana M; Molloy, Justin E; Hoggett, James G

    2004-01-30

    A single-molecule transcription assay has been developed that allows, for the first time, the direct observation of promoter binding, initiation, and elongation by a single RNA polymerase (RNAP) molecule in real-time. To promote DNA binding and transcription initiation, a DNA molecule tethered between two optically trapped beads was held near a third immobile surface bead sparsely coated with RNAP. By driving the optical trap holding the upstream bead with a triangular oscillation while measuring the position of both trapped beads, we observed the onset of promoter binding, promoter escape (productive initiation), and processive elongation by individual RNAP molecules. After DNA template release, transcription re-initiation on the same DNA template is possible; thus, multiple enzymatic turnovers by an individual RNAP molecule can be observed. Using bacteriophage T7 RNAP, a commonly used RNAP paradigm, we observed the association and dissociation (k(off)= 2.9 s(-1)) of T7 RNAP and promoter DNA, the transition to the elongation mode (k(for) = 0.36 s(-1)), and the processive synthesis (k(pol) = 43 nt s(-1)) and release of a gene-length RNA transcript ( approximately 1200 nt). The transition from initiation to elongation is much longer than the mean lifetime of the binary T7 RNAP-promoter DNA complex (k(off) > k(for)), identifying a rate-limiting step between promoter DNA binding and promoter escape.

  2. Binding of Mn-deoxyribonucleoside Triphosphates to the Active Site of the DNA Polymerase of Bacteriophage T7

    Energy Technology Data Exchange (ETDEWEB)

    B Akabayov; C Richardson

    2011-12-31

    Divalent metal ions are crucial as cofactors for a variety of intracellular enzymatic activities. Mg{sup 2+}, as an example, mediates binding of deoxyribonucleoside 5'-triphosphates followed by their hydrolysis in the active site of DNA polymerase. It is difficult to study the binding of Mg{sup 2+} to an active site because Mg{sup 2+} is spectroscopically silent and Mg{sup 2+} binds with low affinity to the active site of an enzyme. Therefore, we substituted Mg{sup 2+} with Mn{sup 2+}:Mn{sup 2+} that is not only visible spectroscopically but also provides full activity of the DNA polymerase of bacteriophage T7. In order to demonstrate that the majority of Mn{sup 2+} is bound to the enzyme, we have applied site-directed titration analysis of T7 DNA polymerase using X-ray near edge spectroscopy. Here we show how X-ray near edge spectroscopy can be used to distinguish between signal originating from Mn{sup 2+} that is free in solution and Mn{sup 2+} bound to the active site of T7 DNA polymerase. This method can be applied to other enzymes that use divalent metal ions as a cofactor.

  3. WATER PRODUCTION IN COMETS 2001 Q4 (NEAT) AND 2002 T7 (LINEAR) DETERMINED FROM SOHO/SWAN OBSERVATIONS

    International Nuclear Information System (INIS)

    Combi, M. R.; Lee, Y.; Maekinen, J. T. T.; Bertaux, J.-L.; Quemerais, E.

    2009-01-01

    The SWAN all-sky camera on the Solar and Heliospheric Observatory (SOHO) spacecraft detected the hydrogen Lyman-alpha (Lyα) comae of comets 2001 Q4 NEAT and 2002 T7 LINEAR for large portions of their perihelion apparitions in 2003 and 2004. C/2001 Q4 NEAT was observed from 2003 September 14 through 2004 November 2, covering heliocentric distances from 3.23 AU before perihelion to 2.75 AU after, and C/2002 T7 LINEAR was observed from 2003 December 4 through 2004 August 6, covering heliocentric distances from 2.52 AU before perihelion to 2.09 AU after. We combined the full set of comet specific and full-sky observations and used our time-resolved model (TRM), which enables us to extract continuous values of the daily-average value of the water production rate throughout most of this entire period. The average power-law fit to the production rate variation of C/2001 Q4 NEAT with heliocentric distance, r, gives 3.5 x 10 29 r -1.7 and that for C/2002 T7 LINEAR gives 4.6 x 10 29 r -2.0 . Both comets show roughly a factor of 2 asymmetry in activity about perihelion, being more active before perihelion. C/2001 Q4 NEAT showed a production rate outburst about 30 days before perihelion (2004 April 15) and then a large extended increase above the nominal trend from 50 to 70 days after perihelion (2004 July 5-July 25).

  4. Experimental investigation of the effects of aqueous species on the dissolution kinetics of R7T7 glass; Etude experimentale de l`influence d`especes aqueuses sur la cinetique de dissolution du verre nucleaire R7T7

    Energy Technology Data Exchange (ETDEWEB)

    Gin, S

    1994-10-01

    This contribution to the study of aqueous corrosion of the French ``R7T7`` reference nuclear containment glass includes a bibliographic survey of prior investigations, highlighting the problems encountered in interpreting the interactions in systems containing clay materials in contact with the glass. An experimental methodology is proposed to investigate the effects of inorganic aqueous species separately from those of a few organic acids on the dissolution mechanisms and kinetics of R7T7 glass at 90 deg. C. The experimental results discussed support the idea that several glass network forming elements may have a kinetically limiting role. The most likely hypothesis to account for the absence of saturation conditions with respect to the glass in certain clay media involves the formation of complexes with kinetically limiting metallic elements such as aluminum released by glass corrosion. This work contributes to a better understanding of the basic mechanisms of nuclear glass dissolution in a geological repository environment. It facilitates the interpretation of glass alteration studies in realistic or actual solutions and may contribute to specifying near field chemical barriers in the form of additives (amorphous silica, aluminum hydroxides or phosphates) around the glass disposal package to enhance the stability of the glass matrix. (author). 148 refs., 40 figs., 32 tabs., 1 append.

  5. The role of DNA-protein interaction in the UV damage of T7 bacteriophage at high fluences

    International Nuclear Information System (INIS)

    Fekete, A.; Ronto, G.

    1980-01-01

    The influence of higher fluences (0.5-10 kJm -2 ) and that of phage protein coat on the UV (lambda = 254 nm) damage of T7 DNA were studied by UV difference spectroscopy. Beside the pyrimidine dimers and adducts produced also in isolated DNA in the case of intact phages and fluences exceeding 0.5 kJ m -2 other photoproducts, probably DNA-protein cross-links were identified as well. Phages deprived of their protein coat by a thermal treatment show similar UV damage to that of isolated DNA. (author)

  6. Escherichia coli pathotypes

    Science.gov (United States)

    Escherichia coli strains are important commensals of the intestinal tract of humans and animals; however, pathogenic strains, including diarrhea-inducing E. coli and extraintestinal pathogenic E. coli. Intestinal E. coli pathotypes may cause a dehydrating watery diarrhea, or more severe diseases su...

  7. Aqueous corrosion mechanisms of the nuclear glass R7T7. Experimental approach. Kinetics and thermodynamic simulation

    International Nuclear Information System (INIS)

    Advocat, T.

    1991-01-01

    An inactive borosilicate glass made of about 30 oxides is studied. The composition was developed by the CEA for encapsulation of calcinated fission product solutions from reprocessing. Hydration energy of glass is first calculated for 8 glasses and results are compared to experimental data. Dissolution of R7T7 glass is examined at 90 0 C in a large range of pH and S/V ratios (glass surface/solution volume). In dilute media (S/V = 0.1 cm -1 ) dissolution is selective at acid pH and stoichiometric at basic pH. In alkaline media dissolution rate increases with pH. Corrosion products, generally amorphous or badly crystallized are observed on glass surface. For high S/V ratios (4, 20, 80 and 200 cm -1 ) the very low dissolution rate is explained by saturation. Orthosilic acid controls corrosion kinetics. A kinetic equation is proposed taking into account pH, S/V ratio and dissolved silica concentration. Geochemical consequences of R7T7 dissolution are modelled at 100 0 C and 90 0 C. Affinity of dissolution reaction depends upon many factors (pH, silica concentration, nature and crystallinity of secondary phases. Reaction affinity is not constant for the long-term [fr

  8. R7T7 glass alteration mechanism in an aqueous closed system: understanding and modelling the long term alteration kinetic

    International Nuclear Information System (INIS)

    Chave, T.

    2007-10-01

    The long term alteration rate of the French R7T7 nuclear glass has been investigated since many years because it will define the overall resistance of the radionuclide containment matrix. Recent studies have shown that the final rate remains constant or is slightly decreasing with time. It never reaches zero. Though this residual rate is very low, only 5 nm per year at 50 C, it would be the dominant alteration phenomenon in a geological repository. Two mechanisms are suggested for explaining such behaviour: diffusion in solution of elements from glass through an amorphous altered layer and precipitation of neo-formed phases. The diffusion processes are in agreement with a solid state diffusion mechanism and can lead to secondary phase precipitation due to solution concentration increases. Observed phases are mainly phyllosilicates and zeolites, in specific conditions. Phyllosilicates are expected to maintain the residual kinetic rate whereas alteration resumption could be observed in presence of zeolites at very high pH or temperature (10.5 at 90 C or temperature above 150 C). Both diffusion and neo-formed phase precipitation have been investigated in order to better understand their impact on the residual alteration rate and have then been modelled by a calculation code, coupling chemistry and transport, in order to be able to better anticipate the long term behaviour of the glass R7T7 in an aqueous closed system. (author)

  9. Interaction of bacteriophage T4 and T7 single-stranded DNA-binding proteins with DNA

    International Nuclear Information System (INIS)

    Shokri, Leila; Williams, Mark C; Rouzina, Ioulia

    2009-01-01

    Bacteriophages T4 and T7 are well-studied model replication systems, which have allowed researchers to determine the roles of many proteins central to DNA replication, recombination and repair. Here we summarize and discuss the results from two recently developed single-molecule methods to determine the salt-dependent DNA-binding kinetics and thermodynamics of the single-stranded DNA (ssDNA)-binding proteins (SSBs) from these systems. We use these methods to characterize both the equilibrium double-stranded DNA (dsDNA) and ssDNA binding of the SSBs T4 gene 32 protein (gp32) and T7 gene 2.5 protein (gp2.5). Despite the overall two-orders-of-magnitude weaker binding of gp2.5 to both forms of DNA, we find that both proteins exhibit four-orders-of-magnitude preferential binding to ssDNA relative to dsDNA. This strong preferential ssDNA binding as well as the weak dsDNA binding is essential for the ability of both proteins to search dsDNA in one dimension to find available ssDNA-binding sites at the replication fork

  10. Conjugation in Escherichia coli

    Science.gov (United States)

    Boyer, Herbert

    1966-01-01

    Boyer, Herbert (Yale University, New Haven, Conn.). Conjugation in Escherichia coli. J. Bacteriol. 91:1767–1772. 1966.—The sex factor of Escherichia coli K-12 was introduced into an E. coli B/r strain by circumventing the host-controlled modification and restriction incompatibilities known to exist between these closely related strains. The sexual properties of the constructed F+ B strain and its Hfr derivatives were examined. These studies showed that the E. coli strain B/r F+ and Hfr derivatives are similar to the E. coli strain K-12 F+ and Hfr derivatives. However, the site of sex factor integration was found to be dependent on the host genome. PMID:5327905

  11. Effects of deposited nuclear and electronic energy on the hardness of R7T7-type containment glass

    Energy Technology Data Exchange (ETDEWEB)

    Peuget, S. [Commissariat a l' Energie Atomique, CEA Marcoule, DEN/DTCD/SECM/LMPA, Batiment 166, BP 17171, F-30207 Bagnols-sur-Ceze Cedex (France)]. E-mail: sylvain.peuget@cea.fr; Noel, P.-Y. [Commissariat a l' Energie Atomique, CEA Marcoule, DEN/DTCD/SECM/LMPA, Batiment 166, BP 17171, F-30207 Bagnols-sur-Ceze Cedex (France); Loubet, J.-L. [Laboratoire de Tribologie et Dynamique des Systemes, UMR CNRS 5513, Ecole Centrale de Lyon 36, avenue Guy de Collongue, 69134 Ecully Cedex (France); Pavan, S. [Laboratoire de Tribologie et Dynamique des Systemes, UMR CNRS 5513, Ecole Centrale de Lyon 36, avenue Guy de Collongue, 69134 Ecully Cedex (France); Nivet, P. [Commissariat a l' Energie Atomique, CEA Marcoule, DEN/DTCD/SECM/LMPA, Batiment 166, BP 17171, F-30207 Bagnols-sur-Ceze Cedex (France); Chenet, A. [Commissariat a l' Energie Atomique, CEA Marcoule, DEN/DTCD/SECM/LMPA, Batiment 166, BP 17171, F-30207 Bagnols-sur-Ceze Cedex (France)

    2006-05-15

    The effects of elastic and inelastic interactions induced by cumulative alpha decay on the hardness of R7T7-type nuclear containment glass were investigated on actinide-doped glass specimens and by external irradiation of inactive glass by light and heavy ions. Vickers microindentation and nanoindentation hardness measurements showed that in the deposited energy range investigated (below 3 x 10{sup 22} keV/cm{sup 3}) inelastic effects have no influence on the plastic response of the glass. Conversely, identical hardness variations versus the nuclear energy deposited in the material were observed on curium-doped glass and on glass irradiated by ion bombardment. The observed hardness variation stabilized after the deposited energy reached about 3 x 10{sup 2} keV{sub nucl}/cm{sup 3}. These findings indicate that the change in the plastic response of the glass is a consequence of ballistic effects.

  12. Experimental alteration of R7T7 glass in salt brines at 90 deg C and 150 deg C

    International Nuclear Information System (INIS)

    Godon, N.; Vernaz, E.; Gin, S.; Beaufort, D.; Thomassin, J.H.

    1991-01-01

    Static experiments have been developed to investigate the R7T7 glass corrosion in four natural salt brines (brines 1 and 3: pure halite, brines 2 and 4: high Mg, K fluid inclusions rich halite), at 90 deg C and 150 deg C with 0.7 cm -1 S/V ratio and at 11 different running times. Analysis of brines after alteration (pHmeter and ICP) added to a detailed study of the crystalline phases developed at the interface glass-brine (XRD,SEM and Microprobe), showed that the influence of the compositional difference is more important on the nature of the secondary phases formed than on the corrosion rate of the glass. After 91 days of alteration at 150 deg C stady states to be reached (after 40 days at 90 deg C). A long term experiment (1 year) is necessary to confirm this hypothesis. 7 refs., 7 figs., 2 tabs

  13. Corrosion mechanisms and behaviour of actinides in the 'R7T7' nuclear glass

    International Nuclear Information System (INIS)

    Fillet, Sylvie

    1987-01-01

    This research thesis reports the study of aqueous corrosion of the R7T7 nuclear glass and of the identified corrosion mechanisms in conditions of static lixiviation which are close to that expected during long term storage in a geological environment. More specifically, this work aims at assessing the durability of this glass which has been selected for the vitrification of solutions from pressurized water reactors. The main glass alteration phenomena have been studied. The first part addresses the study of the alteration of the glassy matrix, and aims at identifying corrosion mechanisms in various lixiviation conditions (high temperature, saturation). The second part addresses the action of different materials present in the environment on the glassy matrix by simulating as well as possible a storage case. Based on the obtained results, a mathematical model is developed to predict the glass behaviour on the long term. Finally, the glass confinement power with respect to actinides is studied [fr

  14. Alteration of R7T7-type nuclear glass in deep geological storage conditions; Alteration du verre de confinement de dechets type R7T7 en condition de stockage geologique

    Energy Technology Data Exchange (ETDEWEB)

    Combarieu, G de

    2007-02-15

    This PhD thesis is aimed to study the alteration of SON68 glass, French inactive glass of R7T7-type, in contact with near field materials of a deep geological storage (French concept from ANDRA) which are mainly metallic iron and Callovo-Oxfordian clay. Therefore, experiments involving a 'glass-iron-clay' system at lab-scale have been carried out. Interactions between glass, iron and clay have been characterised from submicron to millimeter scale by means of SEM, TEM, XRD and XAS and Raman spectroscopies in terms of chemistry and crystal-chemistry. In the mean time, a conceptual model of glass alteration has been developed to account for most of the experimental observations and known mechanisms of alteration. The model has been then transposed within the transport-chemistry code HYTEC, together with developed models of clay and iron corrosion, to simulate the experiments described above. This work is thus a contribution to the understanding of iron corrosion in Callovo-Oxfordian clay and subsequent glass alteration in the newly formed corrosion products, the whole process being considered as a lab-scale model of a deep geological storage of radioactive wastes. (author)

  15. Alteration of R7T7-type nuclear glass in deep geological storage conditions; Alteration du verre de confinement de dechets type R7T7 en condition de stockage geologique

    Energy Technology Data Exchange (ETDEWEB)

    Combarieu, G. de

    2007-02-15

    This PhD thesis is aimed to study the alteration of SON68 glass, French inactive glass of R7T7-type, in contact with near field materials of a deep geological storage (French concept from ANDRA) which are mainly metallic iron and Callovo-Oxfordian clay. Therefore, experiments involving a 'glass-iron-clay' system at lab-scale have been carried out. Interactions between glass, iron and clay have been characterised from submicron to millimeter scale by means of SEM, TEM, XRD and XAS and Raman spectroscopies in terms of chemistry and crystal-chemistry. In the mean time, a conceptual model of glass alteration has been developed to account for most of the experimental observations and known mechanisms of alteration. The model has been then transposed within the transport-chemistry code HYTEC, together with developed models of clay and iron corrosion, to simulate the experiments described above. This work is thus a contribution to the understanding of iron corrosion in Callovo-Oxfordian clay and subsequent glass alteration in the newly formed corrosion products, the whole process being considered as a lab-scale model of a deep geological storage of radioactive wastes. (author)

  16. The DNA-mimic antirestriction proteins ArdA ColIB-P9, Arn T4, and Ocr T7 as activators of H-NS-dependent gene transcription.

    Science.gov (United States)

    Melkina, Olga E; Goryanin, Ignatiy I; Zavilgelsky, Gennadii B

    2016-11-01

    The antirestriction proteins ArdA ColIb-P9, Arn T4 and Ocr T7 specifically inhibit type I and type IV restriction enzymes and belong to the family of DNA-mimic proteins because their three-dimensional structure is similar to the double-helical B-form DNA. It is proposed that the DNA-mimic proteins are able to bind nucleoid protein H-NS and alleviate H-NS-silencing of the transcription of bacterial genes. Escherichia coli lux biosensors were constructed by inserting H-NS-dependent promoters into a vector, thereby placing each fragment upstream of the promoterless Photorhabdus luminescens luxCDABE operon. It was demonstrated that the DNA-mimic proteins ArdA, Arn and Ocr activate the transcription of H-NS-dependent promoters of the lux operon of marine luminescent bacteria (mesophilic Aliivibrio fischeri and psychrophilic Aliivibrio logei), and the dps gene from E. coli. It was also demonstrated that the ArdA antirestriction protein, the genes of which are located on transmissive plasmids ColIb-P9, R64, PK101, decreases levels of H-NS silencing of the PluxC promoter during conjugation in the recipient bacteria. Copyright © 2016 Elsevier GmbH. All rights reserved.

  17. Alteration of French waste glass matrix of R7T7 type in deep geological disposal conditions

    International Nuclear Information System (INIS)

    Combarieu, G. de

    2007-02-01

    The Geological disposal is a possible option for safe and long term management of long lived and highly radioactive wastes. In order to predict the release of radionuclides in the environment, the comprehensive knowledge of glass dissolution rates as well as the properties of near- and far-field in which migration will occur is necessary. This thesis is aimed to describe the alteration of SON68 glass, inactive analog of French R7T7 glass, in contact with disposal materials: metallic iron and Callovo-Oxfordian argilite. Therefore, original experiments have been carried out on a laboratory scaled system involving 'glass-iron-argilite' interactions. The transformations of chemistry and crystal-chemistry are investigated with multi-scale probing tools: SEM, TEM, XRD, XRF, EXAFS and Raman spectroscopies. In the same time, the glass alteration is modeled to obtain a source term in good agreement with the major phenomena observed in common experiments. As an end, geochemical models of iron and argilite transformations are also developed and set together in the transport-chemistry code HYTEC to simulate chemical reactions (iron corrosion, argilite evolution, and glass alteration). Simulations and comparison with experiments have improved the overall knowledge of the glass-iron-clay system. (author)

  18. Sequence and structural characterization of great salt lake bacteriophage CW02, a member of the T7-like supergroup.

    Science.gov (United States)

    Shen, Peter S; Domek, Matthew J; Sanz-García, Eduardo; Makaju, Aman; Taylor, Ryan M; Hoggan, Ryan; Culumber, Michele D; Oberg, Craig J; Breakwell, Donald P; Prince, John T; Belnap, David M

    2012-08-01

    Halophage CW02 infects a Salinivibrio costicola-like bacterium, SA50, isolated from the Great Salt Lake. Following isolation, cultivation, and purification, CW02 was characterized by DNA sequencing, mass spectrometry, and electron microscopy. A conserved module of structural genes places CW02 in the T7 supergroup, members of which are found in diverse aquatic environments, including marine and freshwater ecosystems. CW02 has morphological similarities to viruses of the Podoviridae family. The structure of CW02, solved by cryogenic electron microscopy and three-dimensional reconstruction, enabled the fitting of a portion of the bacteriophage HK97 capsid protein into CW02 capsid density, thereby providing additional evidence that capsid proteins of tailed double-stranded DNA phages have a conserved fold. The CW02 capsid consists of bacteriophage lambda gpD-like densities that likely contribute to particle stability. Turret-like densities were found on icosahedral vertices and may represent a unique adaptation similar to what has been seen in other extremophilic viruses that infect archaea, such as Sulfolobus turreted icosahedral virus and halophage SH1.

  19. Chandra Snapshot Spectral Imaging of Comets C/2001 Q4 (NEAT) and C/2002 T7 (LINEAR)

    Science.gov (United States)

    Lisse, Carey

    2003-09-01

    The highly favorable perigee passage of the very bright comets C/2001 Q4 (NEAT) and C/2002 T7 (LINEAR) in late May 2004 provides an opportunity to study cometary x-ray emission in conjunction with the new CHIPS spectroscopic mission. In 10 ksec of on-target time for each comet, ACIS-S will obtain snapshot images of the comets in the heart of the CHIPS 0.05 0.150 keV spectroscopic monitoring period in late-May 2004. The combined observations have the potential of directly detecting for the first time the ultra-soft emission due to Mg, S, Si, and Fe predicted by McCammon et al. (2002) from soft x-ray background measurements and by Kharchenko et al. (2000, 2003) from models of solar wind minor ion charge exchange emission. New work by Wegmann, Dennerl, and Lisse (2004) allows a determination of the neutral gas production rate from the spatial scale of the emission, and an independent determination of the solar wind minor ion flux density using the x-ray surface brightness.

  20. Calcium carbonate mediates higher lignin peroxidase activity in the culture supernatant of Streptomyces Viridosporus T7A

    Directory of Open Access Journals (Sweden)

    J. M. B. MACEDO

    1999-06-01

    Full Text Available Lignin peroxidase (LiP production has been extensively studied due to the potential use of this enzyme in environmental pollution control. Important aspects of the production of the enzyme by S. viridosporus T7A which have been studied include the improvement of yield and enzyme stabilization. In experiments performed in agitated flasks containing culture media composed of yeast extract as the source of nitrogen, mineral salts and different carbon sources, the use of glucose resulted in the highest values for LiP activity (350 U/L, specific LiP activity (450 U/g and productivity (7 U/L/h. As the profile obtained with glucose-containing medium suggested enzyme instability, the effect of calcium carbonate was evaluated. The addition of CaCO3 in two different concentrations, 0.5% and 5.0%, resulted in higher values of maximum LiP activity, 600 and 900 U/L, respectively. The presence of this salt also anticipated enzyme activity peaks and allowed the detection of higher enzyme activities in the extracellular medium for longer periods of time. These results indicate a positive effect of calcium carbonate on LiP production, which is extremely relevant for industrial processes.

  1. Pyrovanadolysis: a Pyrophosphorolysis-like Reaction Mediated by Pyrovanadate MN2plus and DNA Polymerase of Bacteriophage T7

    Energy Technology Data Exchange (ETDEWEB)

    B Akabayov; A Kulczyk; S Akabayov; C Thiele; L McLaughlin; B Beauchamp; C Richardson

    2011-12-31

    DNA polymerases catalyze the 3'-5'-pyrophosphorolysis of a DNA primer annealed to a DNA template in the presence of pyrophosphate (PP{sub i}). In this reversal of the polymerization reaction, deoxynucleotides in DNA are converted to deoxynucleoside 5'-triphosphates. Based on the charge, size, and geometry of the oxygen connecting the two phosphorus atoms of PP{sub i}, a variety of compounds was examined for their ability to carry out a reaction similar to pyrophosphorolysis. We describe a manganese-mediated pyrophosphorolysis-like activity using pyrovanadate (VV) catalyzed by the DNA polymerase of bacteriophage T7. We designate this reaction pyrovanadolysis. X-ray absorption spectroscopy reveals a shorter Mn-V distance of the polymerase-VV complex than the Mn-P distance of the polymerase-PP{sub i} complex. This structural arrangement at the active site accounts for the enzymatic activation by Mn-VV. We propose that the Mn{sup 2+}, larger than Mg{sup 2+}, fits the polymerase active site to mediate binding of VV into the active site of the polymerase. Our results may be the first documentation that vanadium can substitute for phosphorus in biological processes.

  2. Development of a T7 Phage Display Library to Detect Sarcoidosis and Tuberculosis by a Panel of Novel Antigens

    Directory of Open Access Journals (Sweden)

    Harvinder Talwar

    2015-04-01

    Full Text Available Sarcoidosis is a granulomatous inflammatory disease, diagnosed through tissue biopsy of involved organs in the absence of other causes such as tuberculosis (TB. No specific serologic test is available to diagnose and differentiate sarcoidosis from TB. Using a high throughput method, we developed a T7 phage display cDNA library derived from mRNA isolated from bronchoalveolar lavage (BAL cells and leukocytes of sarcoidosis patients. This complex cDNA library was biopanned to obtain 1152 potential sarcoidosis antigens and a microarray was constructed to immunoscreen two different sets of sera from healthy controls and sarcoidosis. Meta-analysis identified 259 discriminating sarcoidosis antigens, and multivariate analysis identified 32 antigens with a sensitivity of 89% and a specificity of 83% to classify sarcoidosis from healthy controls. Additionally, interrogating the same microarray platform with sera from subjects with TB, we identified 50 clones that distinguish between TB, sarcoidosis and healthy controls. The top 10 sarcoidosis and TB specific clones were sequenced and homologies were searched in the public database revealing unique epitopes and mimotopes in each group. Here, we show for the first time that immunoscreenings of a library derived from sarcoidosis tissue differentiates between sarcoidosis and tuberculosis antigens. These novel biomarkers can improve diagnosis of sarcoidosis and TB, and may aid to develop or evaluate a TB vaccine.

  3. DNA Recognition by the DNA Primase of Bacteriophage T7: A Structure Function Study of the Zinc-Binding Domain

    International Nuclear Information System (INIS)

    Akabayov, B.; Lee, S.; Akabayov, S.; Rekhi, S.; Zhu, B.; Richardson, C.

    2009-01-01

    Synthesis of oligoribonucleotide primers for lagging-strand DNA synthesis in the DNA replication system of bacteriophage T7 is catalyzed by the primase domain of the gene 4 helicase-primase. The primase consists of a zinc-binding domain (ZBD) and an RNA polymerase (RPD) domain. The ZBD is responsible for recognition of a specific sequence in the ssDNA template whereas catalytic activity resides in the RPD. The ZBD contains a zinc ion coordinated with four cysteine residues. We have examined the ligation state of the zinc ion by X-ray absorption spectroscopy and biochemical analysis of genetically altered primases. The ZBD of primase engaged in catalysis exhibits considerable asymmetry in coordination to zinc, as evidenced by a gradual increase in electron density of the zinc together with elongation of the zinc-sulfur bonds. Both wild-type primase and primase reconstituted from purified ZBD and RPD have a similar electronic change in the level of the zinc ion as well as the configuration of the ZBD. Single amino acid replacements in the ZBD (H33A and C36S) result in the loss of both zinc binding and its structural integrity. Thus the zinc in the ZBD may act as a charge modulation indicator for the surrounding sulfur atoms necessary for recognition of specific DNA sequences.

  4. Myxoma virus M-T7, a secreted homolog of the interferon-gamma receptor, is a critical virulence factor for the development of myxomatosis in European rabbits.

    Science.gov (United States)

    Mossman, K; Nation, P; Macen, J; Garbutt, M; Lucas, A; McFadden, G

    1996-01-01

    Myxoma virus is a leporipoxvirus of New World rabbits (Sylvilagus sp.) that induces a rapidly lethal infection known as myxomatosis in the European rabbit (Oryctolagus cuniculus). Like all poxviruses, myxoma virus encodes a plethora of proteins to circumvent or inhibit a variety of host antiviral immune mechanisms. M-T7, the most abundantly secreted protein of myxoma virus-infected cells, was originally identified as an interferon-gamma receptor homolog (Upton, Mossman, and McFadden, Science 258, 1369-1372, 1992). Here, we demonstrate that M-T7 is dispensable for virus replication in cultured cells but is a critical virulence factor for virus pathogenesis in European rabbits. Disruption of both copies of the M-T7 gene in myxoma virus was achieved by the deletion of 372 bp of M-T7 coding sequences, replacement with a selectable marker, p7.5Ecogpt, and selection of a recombinant virus (vMyxlac-T7gpt) resistant to mycophenolic acid. vMyxlac-T7gpt expressed no detectable M-T7 protein and infected cells supernatants were devoid of any detectable interferon-gamma binding activities. Immunohistochemical staining with anti-beta-galactosidase and anti-CD43 antibodies demonstrated that in vMyxlac-T7gpt-infected rabbits the loss of M-T7 not only caused a dramatic reduction in disease symptoms and viral dissemination to secondary sites, but also dramatically influenced host leukocyte behavior. Notably, primary lesions in wild-type virus infections were generally underlayed by large masses of inflammatory cells that did not effectively migrate into the dermal sites of viral replication, whereas in vMyxlac-T7gpt infections this apparent block to leukocyte influx was relieved. A second major phenotypic distinction noted for the M-T7 knockout virus was the extensive activation of lymphocytes in secondary immune organs, particularly the spleen and lymph nodes, by Day 4 of the infection. This is in stark contrast to infection by wild-type myxoma virus, which results in relatively

  5. Vanillin production using Escherichia coli cells over-expressing isoeugenol monooxygenase of Pseudomonas putida.

    Science.gov (United States)

    Yamada, Mamoru; Okada, Yukiyoshi; Yoshida, Toyokazu; Nagasawa, Toru

    2008-04-01

    The isoeugenol monooxygenase gene of Pseudomonas putida IE27 was inserted into an expression vector, pET21a, under the control of the T7 promoter. The recombinant plasmid was introduced into Escherichia coli BL21(DE3) cells, containing no vanillin-degrading activity. The transformed E. coli BL21(DE3) cells produced 28.3 g vanillin/l from 230 mM isoeugenol, with a molar conversion yield of 81% at 20 degrees C after 6 h. In the reaction system, no accumulation of undesired by-products, such as vanillic acid or acetaldehyde, was observed.

  6. Geochemical modelling of the long-term dissolution behaviour of the French nuclear glass R7T7

    International Nuclear Information System (INIS)

    Michaux, L.; Mouche, E.; Petit, J.-C.; Fritz, B.

    1992-01-01

    The long-term dissolution behaviour of the French nuclear reference glass R7T7 was studied by means of the geochemical code DISSOL. New experimental data which support some of the assumptions of DISSOL are presented: namely, that the dissolution is congruent and that the altered layer can be considered as an assemblage of secondary phases. At 100 o C the main results of modelling are that the altered layer is essentially formed of a pure siliceous phase (amorphous silica or chalcedony) associated with smectites and zeolites. This sequence of secondary minerals is closely linked to the chemical composition of the glass. For high degrees of reaction, corresponding to high B concentration, the ionic strength reaches 1 and the pH varies from 9 to 10 depending on the CO 2 fugacity; B,Li and Na are essentially found in solution and their concentrations depend on the amount of dissolved glass. By contrast Fe,Al and Zn have low solution concentrations which are controlled by solubility products of secondary minerals. Silicon and Ca have an intermediate behaviour which depends on the choice of selected secondary minerals. The total volume of the secondary phases is always lower than that of the corresponding dissolved glass. The results of modelling compared to static leaching experimental results show only minor differences which can be explained by kinetic control or colloid formation. It is concluded that the altered layer is not a barrier to diffusion. The consequences of this work for actinide solubility are also discussed. (author)

  7. Enterohemorrhagic Escherichia coli (EHEC

    Directory of Open Access Journals (Sweden)

    Abdullah Kilic

    2011-08-01

    Full Text Available Escherichia coli is a bacterium that is commonly found in the gut of humans and warm-blooded animals. Most strains of E. coli are harmless for human. E. coli O157:H7 is the most common member of a group of pathogenic E. coli strains known variously as enterohaemorrhagic, verocytotoxin-producing, or Shiga-toxin-producing organisms. EHEC bacterium is the major cause of haemorrhagic colitis and haemolytic uraemic syndrome. The reservoir of this pathogen appears to be mainly cattle and other ruminants such as camels. It is transmitted to humans primarily through consumption of contaminated foods. [TAF Prev Med Bull 2011; 10(4.000: 387-388

  8. Molecular subtypes in stage II-III colon cancer defined by genomic instability: early recurrence-risk associated with a high copy-number variation and loss of RUNX3 and CDKN2A.

    Directory of Open Access Journals (Sweden)

    Marianne Berg

    Full Text Available We sought to investigate various molecular subtypes defined by genomic instability that may be related to early death and recurrence in colon cancer.We sought to investigate various molecular subtypes defined by instability at microsatellites (MSI, changes in methylation patterns (CpG island methylator phenotype, CIMP or copy number variation (CNV in 8 genes. Stage II-III colon cancers (n = 64 were investigated by methylation-specific multiplex ligated probe amplification (MS-MLPA. Correlation of CNV, CIMP and MSI, with mutations in KRAS and BRAFV600E were assessed for overlap in molecular subtypes and early recurrence risk by uni- and multivariate regression.The CIMP phenotype occurred in 34% (22/64 and MSI in 27% (16/60 of the tumors, with noted CIMP/MSI overlap. Among the molecular subtypes, a high CNV phenotype had an associated odds ratio (OR for recurrence of 3.2 (95% CI 1.1-9.3; P = 0.026. Losses of CACNA1G (OR of 2.9, 95% CI 1.4-6.0; P = 0.001, IGF2 (OR of 4.3, 95% CI 1.1-15.8; P = 0.007, CDKN2A (p16 (OR of 2.0, 95% CI 1.1-3.6; P = 0.024, and RUNX3 (OR of 3.4, 95% CI 1.3-8.7; P = 0.002 were associated with early recurrence, while MSI, CIMP, KRAS or BRAF V600E mutations were not. The CNV was significantly higher in deceased patients (CNV in 6 of 8 compared to survivors (CNV in 3 of 8. Only stage and loss of RUNX3 and CDKN2A were significant in the multivariable risk-model for early recurrence.A high copy number variation phenotype is a strong predictor of early recurrence and death, and may indicate a dose-dependent relationship between genetic instability and outcome. Loss of tumor suppressors RUNX3 and CDKN2A were related to recurrence-risk and warrants further investigation.

  9. An anti-tumor protein produced by Trichinella spiralis and identified by screening a T7 phage display library, induces apoptosis in human hepatoma H7402 cells

    Science.gov (United States)

    Trichinella spiralis infection confers effective resistance to tumor cell expansion. In this study, a T7 phage cDNA display library was constructed to express genes encoded by T. spiralis. Organic phase multi-cell screening was used to sort through candidate proteins in a transfected human chronic m...

  10. Catabolic fate of Streptomyces viridosporus T7A-Produced, acid precipitable polymeric lignin upon incubation with ligninolytic Streptomyces species and Phanerochaete chrysosporium

    International Nuclear Information System (INIS)

    Pometto, A.L. III; Crawford, D.L.

    1986-01-01

    Degradation of ground and hot-water-extracted corn stover (Zea mays) lignocellulose by Streptomyces viridosporus T7A generates a water-soluble lignin degradation intermediate termed acid-precipitable polymeric lignin (APPL). The further catabolism of T7A-APPL by S. viridosporus T7A, S. badius 252, and S. setonii75Vi2 was followed for 3 weeks. APPL catabolism by Phanerochaete chrysosporium was followed in stationary cultures in a low-nitrogen medium containing 1% (wt/vol) glucose and 0.05% (wt/vol) T7A-APPL. Metabolism of the APPL was followed by turbidometric assay (600 nm) and by direct measurement of APPL recoverable from the medium. Accumulation and disappearance of soluble low-molecular-weight products of APPL catabolism were followed by gas-liquid chromatography and by high-pressure liquid chromatography, utilizing a diode array detector. Mineralization of a [ 14 C-lignin]APPL was also followed. The percent 14 C recovered as 14 CO 2 , 14 C-APPL, 14 C-labeled water-soluble products, and cell mass-associated radioactivity, were determined for each microorganism after 1 and 3 weeks of incubation in bubbler tube cultures at 37 0 C. P. chrysosporium evolved the most 14 CO 2 , and S. viridosporus gave the greatest decrease in recoverable 14 C-APPL. The results show that S. badius was not able to significantly degrade the APPL, while the other microorganisms demonstrated various APPL-degrading abilities

  11. ANIMAL ENTEROTOXIGENIC ESCHERICHIA COLI

    Science.gov (United States)

    Dubreuil, J. Daniel; Isaacson, Richard E.; Schifferli, Dieter M.

    2016-01-01

    Enterotoxigenic Escherichia coli (ETEC) is the most common cause of E. coli diarrhea in farm animals. ETEC are characterized by the ability to produce two types of virulence factors; adhesins that promote binding to specific enterocyte receptors for intestinal colonization and enterotoxins responsible for fluid secretion. The best-characterized adhesins are expressed in the context of fimbriae, such as the F4 (also designated K88), F5 (K99), F6 (987P), F17 and F18 fimbriae. Once established in the animal small intestine, ETEC produces enterotoxin(s) that lead to diarrhea. The enterotoxins belong to two major classes; heat-labile toxin that consist of one active and five binding subunits (LT), and heat-stable toxins that are small polypeptides (STa, STb, and EAST1). This chapter describes the disease and pathogenesis of animal ETEC, the corresponding virulence genes and protein products of these bacteria, their regulation and targets in animal hosts, as well as mechanisms of action. Furthermore, vaccines, inhibitors, probiotics and the identification of potential new targets identified by genomics are presented in the context of animal ETEC. PMID:27735786

  12. PART I. ESCHERICHIA COLI

    Directory of Open Access Journals (Sweden)

    Sanaa Mahdi Oraibi

    2016-11-01

    Full Text Available The presence of Escherichia coli in the air of facilities involved in management and composting of post-slaughter poultry wastes in selected plants of West Western Pomerania region was studied. Measurements were made on four dates in a variety of weather conditions during the year. The study was conducted at 5 objects that differ in the type of waste and the degree of preparation for composting. These were: chemical treatment and preliminary processing plant, liquid wastes reservoir, platform for preparation of materials for composting, storage of biological sediments, and composting facility. Measurement of bacteria count was carried out in accordance with the applicable procedures on selective chromogenic TBX medium. The assays revealed the presence of E. coli at all test objects, but not always on all measurement dates. It has been shown that the presence of E. coli was from 20 to 3047 CFU∙m-3 of air, although the largest quantities were most frequently detected in the air of the building for post-slaughter waste pre-treatment in chemical treatment plant.

  13. The complete sequence of marine bacteriophage VpV262 infecting vibrio parahaemolyticus indicates that an ancestral component of a T7 viral supergroup is widespread in the marine environment

    International Nuclear Information System (INIS)

    Hardies, Stephen C.; Comeau, Andre M.; Serwer, Philip; Suttle, Curtis A.

    2003-01-01

    The 46,012-bp sequence of the marine bacteriophage VpV262 infecting the bacterium Vibrio parahaemolyticus is reported. The VpV262 sequence reveals that it is a distant relative of marine Roseophage SIO1, and an even more distant relative of coliphage T7. VpV262 and SIO1 appear to represent a widespread marine phage group that lacks an RNA polymerase gene and is ancestral to the T7-like phages. We propose that this group together with the T7-like phages be designated as the T7 supergroup. The ancestral head structure gene module for the T7 supergroup was reconstructed by using sensitive biased Psi-blast searches supplemented by statistical support derived from gene order. In the early and replicative segments, these phages have participated in extensive interchange with the viral gene pool. VpV262 carries a different replicative module than SIO1 and the T7-like phages

  14. Hydrotalcite formed by alteration of R7T7 nuclear waste glass and basaltic glass in salt brine at 190 degrees C

    International Nuclear Information System (INIS)

    Abdelouas, A.; Crovisier, J.L.; Lutze, W.; Mueller, R.; Bernotat, W.

    1994-01-01

    The R7T7 and synthetic basaltic glasses were submitted to corrosion in a saline MgCl 2 dominated solution at 190 degrees C. For both glasses, the early alteration product is a hydrotalcite-like compound in which HPO 4 2- , SO 4 2- and Cl - substitutes to CO 3 2- . The measured d 003 spacing is 7.68 angstrom for the hydrotalcite formed from R7T7 glass and 7.62 angstrom for the hydrotalcite formed from basaltic glass which reflect the high aluminium content. Chemical microanalyses show that the hydrotalcite is subsequently covered by a silica-rich gel which evolves into saponite after few months

  15. The complete genomic sequence of lytic bacteriophage gh-1 infecting Pseudomonas putida--evidence for close relationship to the T7 group

    International Nuclear Information System (INIS)

    Kovalyova, Irina V.; Kropinski, Andrew M.

    2003-01-01

    The genome of the lytic Pseudomonas putida bacteriophage gh-1 is linear double-stranded DNA containing 37,359 bp with 216-bp direct terminal repeats. Like other members of the T7 group, the gh-1 genome contains regions of high homology to T7 interspersed with nonhomologous regions that contain small open reading frames of unknown function. The genome shares 31 genes in common with other members of the T7 group, including RNA polymerase, and an additional 12 unique putative genes. A major difference between gh-1 and other members of this group is the absence of any open reading frames between the left direct terminal repeat and gene 1. Sequence analysis of the gh-1 genome also revealed the presence of 10 putative phage promoters with a consensus sequence similar to the promoters of T3 and phiYeO3-12 (consensus: TAAAAACCCTCACTRTGGCHSCM). P. putida mutants resistant to gh-1 were demonstrated to have an altered lipopolysaccharide structure, indicating that members of this group use lipopolysaccharide as their cellular receptor

  16. 76 FR 20542 - Escherichia coli

    Science.gov (United States)

    2011-04-13

    ... beef, Escherichia coli and coliphages were found in chicken, fresh pork, fresh oyster, fresh mushrooms, lettuce, chicken pot pie, biscuit dough, deli loaf, deli roasted turkey, and package roasted chicken... surfaces, and in foods such as ground beef, pork sausage, chicken, oysters, cheese, fresh mushrooms, and...

  17. ESCHERICHIA COLI AND STAPHYLOCOCCUS AUREUS

    African Journals Online (AJOL)

    DR. AMINU

    ABSTRACT. The bio-effects of the ethanol extracts from the leaf and stem of Momordica charantia were studied with the view to ascertain the medical usefulness ascribed to the plant by the locals. The plant parts, stem and leaf, revealed remarkable activity against Escherichia coli and Staphlococcus aureus. The leaves ...

  18. Conjugal Pairing in Escherichia Coli

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 13; Issue 8. Conjugal Pairing in Escherichia Coli. Joshua Lederberg. Classics Volume 13 Issue 8 August 2008 pp 793-794. Fulltext. Click here to view fulltext PDF. Permanent link: https://www.ias.ac.in/article/fulltext/reso/013/08/0793-0794 ...

  19. Escherichia coli as a probiotic?

    NARCIS (Netherlands)

    Jansen, GJ; Wildeboer-Veloo, ACM; van der Waaij, D; Degener, JE

    1998-01-01

    The influence of oral treatment with a suspension of non-pathogenic Escherichia coli cells (commercially available as: Symbioflor II(R)) on the morphological composition of the gut microflora and on the systemic humoral immune response (the IgG-, IgA- and IgM-isotype) against the bacterial cells in

  20. Percutaneous electric neurostimulation of dermatome T7 improves glycemic profile in obese and type 2 diabetic patients. A randomized clinical study.

    Science.gov (United States)

    Ruiz-Tovar, Jaime; Llavero, Carolina; Ortega, Irene; Diez, María; Zubiaga, Lorea; Calpena, Rafael

    2015-01-01

    Endocrine pancreatic segregation is regulated by the autonomic nervous system. The parasympathetic system stimulates insulin production by the beta cells and inhibits the adrenergic discharge by the sympathetic nervous system. The aim of this study was to evaluate the effect of percutaneous neurostimulation (PENS) of dermatome T7, generating a somato-autonomic reflex, whose efferent pathway are the vagal branches that specifically stimulate the pancreas. The effect of this treatment on glycemia, insulin secretion and insulin resistance was investigated. A prospective randomized clinical trial was performed. Patients with Body Mass Index>30kg/m(2) and diagnosis of diabetes mellitus treated with Metformin were included. Patients were divided into 2 groups: Patients undergoing PENS of dermatome T7 (12 sessions of 30minutes weekly) associated with a 1,200 Kcal/day diet (Group 1) and patients following only a 1,200Kcal/day diet (Group 2). All the patients underwent a blood sample extraction before the treatment and 7 days after finishing it. 60 patients were included: 30 in each group. After finishing the treatment, in Group 1 a significant decrease in glycemia (Mean decrease of 62,1mg/dl; P=.024) and HOMA (Mean decrease 1.37; P=.014) was observed. In Group 2, no significant differences between pre and post-treatment values were observed. PENS of dermatome T7 associated with a 1,200Kcal/day diet achieves a greater reduction in glycemia and insulin resistance than with diet exclusively after 3 months of treatment. Copyright © 2014 AEC. Publicado por Elsevier España, S.L.U. All rights reserved.

  1. AML with t(7;12)(q36;p13) is associated with infancy and trisomy 19. Data from NOPHO-AML and review of the literature

    DEFF Research Database (Denmark)

    Espersen, Anne Dorte Lerche; Noren-Nyström, Ulrika; Abrahamsson, Jonas

    2018-01-01

    The t(7;12)(q36;p13) (MNX1/ETV6) is not included in the WHO classification but has been described in up to 30% of acute myeloid leukemia (AML) in children ...). A literature review identified 35 patients with this translocation, published between 2000 and 2015. Outcome data were available in 22 cases. The NOPHO-AML (Nordic Society for Pediatric Hematology and Oncology) database contained 651 patients with AML from 1993 to 2014 and seven (1.1%) had the translocation...

  2. Mechanisms and kinetics laws of inactive R7T7 reference glass dissolution in water at 90 deg C: initial dissolution rate measurements

    International Nuclear Information System (INIS)

    Advocat, T.; Ghaleb, D.; Vernaz, E.

    1993-02-01

    The initial dissolution rate of inactive R7T7 reference glass was measured at 90 deg C in dilute aqueous solutions first at unspecified pH, then with imposed pH values. In distilled water, R7T7 glass corrosion initially involved preferential extraction of boron and network modifier elements (Li, Na, Ca) as long as the solution pH remained acid. When the solution pH became alkaline, glass dissolution was stoichiometric. These two mechanisms were confirmed by dissolution tests in aqueous solutions at imposed pH values under acid and alkaline conditions. The initial dissolution rate r 0 in mole.cm -3 .s -1 also increased significantly in alkaline media when the pH of the aqueous phase increased: in slightly acid media, selective glass dissolution formed a residual, de-alkalinized, hydrated glass that was characterized by transmission electron microscopy and secondary ion mass spectrometry. Under steady-state dissolution conditions, the initial glass corrosion rate (in mole.cm -3 .s -1 ) was: in acid and alkaline media, amorphous and crystallized alteration products formed after complete dissolution of the silicated glass network. The first products formed consisted mainly of Zr, Rare Earths, Fe and Al. (author). 67 refs., 29 figs., 26 tabs., 21 plates

  3. R7T7 nuclear waste glass behavior in moist clay: role of the clay mass/glass surface area ratio

    International Nuclear Information System (INIS)

    Godon, N.; Vernaz, E.

    1989-01-01

    R7T7 glass alteration was investigated in the presence of various moist clays. In contact with smectite 4a, selected in France as a potential engineered barrier material, the glass was significantly corroded: after 6 months the glass corrosion rate was practically the same as the initial alteration rate in double-distilled water. Substantially lower alteration was observed in contact with bentonite 6 activated by sodium carbonate. Smectite 4a consumes silicon released by glass corrosion, and thus retards the rise to high silicon concentrations in solution at which the glass corrosion rate diminishes. Glass can therefore in some cases be altered as much in moist clay as in water with high renewal rates. Other experiments with smaller quantities of smectite 4a showed that the phenomenon becomes less important in time: its duration is proportional to the ratio between the clay mass (C) and the glass surface area (SA). Comparing the results of studies at different C/SA ratios indicates that low glass corrosion rates are obtained more slowly at higher C/SA ratios. Tests with 239 Pu-doped R7T7 glass also showed that the radionuclide retention factor in the alteration film at the glass surface is only 6, compared with a factor of nearly 50 in double-distilled water

  4. Hydrothermal alteration of the glass R7T7. Glass dissolution kinetics at 150 and 2500, role of neo-formed phases

    International Nuclear Information System (INIS)

    Caurel, J.

    1990-01-01

    The glass R7T7 is chosen in France for vitrification of solution from reprocessing. Safety requires the knowledge of R7T7 long term behavior in deep geologic formations. Temperature dependence of leaching between 50 and 300 0 C is studied by static tests for 7 days. An activation energy of 30kJ/Mole is calculated between 50; 75 or 100 0 C and 250 0 C. Results suggest similar corrosion mechanisms between 90-100 and 250 0 C by a complete change between 250 and 275 0 C. Glass corrosion kinetics at 150 0 C and 250 0 C between 1 day and 1 year evidence the precipitation of aluminosilicates and formation of thick amorphous gels progressively enriched with silica. Glass dissolution at 150 0 C and 250 0 C is simulated with the geochemical DISSOL code. Results suggest that dissolution kinetics are controlled by activity of H 4 SiO 4 in solution only. Silica contained into the gel controls corrosion kinetics different from 0. Even if the nature of dissolution mechanisms does not seem modified between 150 and 250 0 C, sample cracking at 250 0 C induces an increase of dissolved glass that does not allow a direct comparison of corrosion kinetics between 150 and 250 0 C [fr

  5. Effects of MgO on short and long term stabilities in water of R7T7 and M7 nuclear waste glasses

    Energy Technology Data Exchange (ETDEWEB)

    Advocat, T; Vernaz, E; Dussossoy, J [CEA Centre d` Etudes de la Vallee du Rhone, 30 - Marcoule (France). Dept. des Procedes de Retraitement; Crovisier, J L [Centre National de la Recherche Scientifique (CNRS), 67 - Strasbourg (France)

    1993-12-31

    Magnesium oxide was added to the standard R7T7 and M7 glass compositions developed for containment of fission product solutions. M7 differs from the R7T7 reference glass mainly by a larger proportion of network modifier elements and a correspondingly lower proportion of glass network forming elements. The percentage of fission products, simulated in this study by inactive elements, was the same in both cases. Increasing the MgO content of the glass compositions by 2 to 5 wt% resulted in significant variations in the aqueous leaching resistance at 90 and 100 deg C. Experimental findings demonstrated that the initial dissolution rate measured at 100 deg C in a Soxhlet apparatus was proportional to the MgO content and inversely proportional to the network former content (mainly SiO2). This was confirmed by a glass hydration model based on the thermodynamic stability of the glass matrix components. Aqueous corrosion tests were also conducted at 90 deg C under static conditions at various SA/V ratios to simulate the progress of the reaction. Under these conditions, the glass dissolution rate diminished more slowly in time when the initial magnesium content was high and the network former content was low. This may be due primarily to a variation in the glass silica solubility limit related to the glass composition; it may also be related to the formation of secondary silica and magnesia alteration products controlling the glass dissolution reaction affinity.

  6. Systematic comparison of co-expression of multiple recombinant thermophilic enzymes in Escherichia coli BL21(DE3).

    Science.gov (United States)

    Chen, Hui; Huang, Rui; Zhang, Y-H Percival

    2017-06-01

    The precise control of multiple heterologous enzyme expression levels in one Escherichia coli strain is important for cascade biocatalysis, metabolic engineering, synthetic biology, natural product synthesis, and studies of complexed proteins. We systematically investigated the co-expression of up to four thermophilic enzymes (i.e., α-glucan phosphorylase (αGP), phosphoglucomutase (PGM), glucose 6-phosphate dehydrogenase (G6PDH), and 6-phosphogluconate dehydrogenase (6PGDH)) in E. coli BL21(DE3) by adding T7 promoter or T7 terminator of each gene for multiple genes in tandem, changing gene alignment, and comparing one or two plasmid systems. It was found that the addition of T7 terminator after each gene was useful to decrease the influence of the upstream gene. The co-expression of the four enzymes in E. coli BL21(DE3) was demonstrated to generate two NADPH molecules from one glucose unit of maltodextrin, where NADPH was oxidized to convert xylose to xylitol. The best four-gene co-expression system was based on two plasmids (pET and pACYC) which harbored two genes. As a result, apparent enzymatic activities of the four enzymes were regulated to be at similar levels and the overall four-enzyme activity was the highest based on the formation of xylitol. This study provides useful information for the precise control of multi-enzyme-coordinated expression in E. coli BL21(DE3).

  7. Characterization, Genome Sequence, and Analysis of Escherichia Phage CICC 80001, a Bacteriophage Infecting an Efficient L-Aspartic Acid Producing Escherichia coli.

    Science.gov (United States)

    Xu, Youqiang; Ma, Yuyue; Yao, Su; Jiang, Zengyan; Pei, Jiangsen; Cheng, Chi

    2016-03-01

    Escherichia phage CICC 80001 was isolated from the bacteriophage contaminated medium of an Escherichia coli strain HY-05C (CICC 11022S) which could produce L-aspartic acid. The phage had a head diameter of 45-50 nm and a tail of about 10 nm. The one-step growth curve showed a latent period of 10 min and a rise period of about 20 min. The average burst size was about 198 phage particles per infected cell. Tests were conducted on the plaques, multiplicity of infection, and host range. The genome of CICC 80001 was sequenced with a length of 38,810 bp, and annotated. The key proteins leading to host-cell lysis were phylogenetically analyzed. One protein belonged to class II holin, and the other two belonged to the endopeptidase family and N-acetylmuramoyl-L-alanine amidase family, respectively. The genome showed the sequence identity of 82.7% with that of Enterobacteria phage T7, and carried ten unique open reading frames. The bacteriophage resistant E. coli strain designated CICC 11021S was breeding and its L-aspartase activity was 84.4% of that of CICC 11022S.

  8. Effects of β-irradiation in multicomponent glasses simulating the matrix of the French nuclear waste glass (R7T7)

    International Nuclear Information System (INIS)

    Boizot, B.; Ghaleb, D.; Petite, G.

    2001-01-01

    4-, 5- and 6-oxide components alumino-borosilicate glasses, with compositions closed to the matrix of the french nuclear glass 'R7T7' have been irradiated with electrons (β) at 2.5 MeV with a Van de Graff accelerator. These glasses have been studied after irradiation with different spectroscopic methods: Electron Paramagnetic Resonance for the study of defects, Raman Micro-spectroscopy for the study of amorphous network evolution under irradiation, and by 11 B MAS NMR. The results of these studies are presented here. It shows in particular a great sensibility to the irradiation conditions like dose rate and irradiation temperature, who are therefore important parameters for the representativeness of such experiments. (authors)

  9. Partitioning of the rare earths and actinides between R7T7 nuclear glass alteration products and solution according to disposal conditions

    International Nuclear Information System (INIS)

    Menard, O.

    1995-01-01

    The alteration of nuclear glass by water is liable to release radionuclides into the environment. Determining the release kinetics of these elements and their aqueous chemical forms are therefore essential steps in establishing the safety of a geological repository site. Leach tests were conducted with a nonradioactive specimen of the French ''R7T7'' light water containment glass spiked with U and Th, and with two R7T7 specimens spiked with 237 Np and 239 Pu, respectively. The alteration solution compositions were representative of deep groundwater and contained carbonate, sulfate, phosphate, fluorine and chlorine ions. The release of U, Th, Np and Pu, as well as of the rare earths La, Ce and Nd were monitored by ICP mass spectrometry and by α spectrometry. Scanning and transmission electron microscopic examination of the nonradioactive altered glass surfaces was also performed to assess the partitioning balance for the rare earths, U and Th between the glass alteration products and solution. The mobility of these elements depends on two competing mechanisms. The rare earths and thorium are incorporated in the alteration products (gel); the retention process is assumed to involve chemisorption or coprecipitation, enhanced in the gel layer by the presence of phosphate ions in particular. Conversely, the aqueous species in the alteration solutions (mainly anions) form complexes with the actinides and rare earths; this phenomenon is particularly evident with U and Np. The presence of carbonate ions favors this mobility. Plutonium differs from U and Np in that it is adsorbed mainly on colloids formed by glass dissolution, the principal factors governing its chemical evolution in solution. (author). refs., 122 figs., 185 tabs

  10. Enteroaggregative Escherichia coli in Daycare

    DEFF Research Database (Denmark)

    Hebbelstrup Jensen, Betina; Stensvold, Christen R.; Struve, Carsten

    2016-01-01

    Enteroaggregative Escherichia coli (EAEC) has been associated with persistent diarrhea, reduced growth acceleration, and failure to thrive in children living in developing countries and with childhood diarrhea in general in industrialized countries. The clinical implications of an EAEC carrier...... and answered a questionnaire regarding gastrointestinal symptoms and exposures. Exposures included foreign travel, consumption of antibiotics, and contact with a diseased animal. In the capital area of Denmark, a total of 179 children aged 0-6 years were followed in a cohort study, in the period between 2009...

  11. Asymptomatic bacteriuria Escherichia coli strains

    DEFF Research Database (Denmark)

    Hancock, Viktoria; Nielsen, E.M.; Klemm, Per

    2006-01-01

    Urinary tract infections (UTIs) affect millions of people each year. Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU) in humans. Persons affected by ABU may carry a particular E. coli strain for extended periods of time without any symptoms. In contrast...... to uropathogenic E. coli (UPEC) that cause symptomatic UTI, very little is known about the mechanisms by which these strains colonize the urinary tract. Here, we have investigated the growth characteristics in human urine as well as adhesin repertoire of nine ABU strains; the ability of ABU strains to compete...

  12. Identification of the Serratia marcescens hemolysin determinant by cloning into Escherichia coli

    International Nuclear Information System (INIS)

    Braun, V.; Neuss, B.; Ruan, Y.; Schiebel, E.; Schoeffler, H.; Jander, G.

    1987-01-01

    A cosmid bank of Serratia marcescens was established from which DNA fragments were cloned into the plasmid pBR322, which conferred the chromosomally encoded hemolytic activity to Escherichia coli K-12. By transposon mutagenesis with Tn1000 and Tn5 IS50/sub L/::phoA (TnphoA), the coding region was assigned to a DNA fragment, designated hyl, comprising approximately 7 kilobases. Two proteins with molecular weights of 61,000 (61K protein) and 160,000 (160K protein) were expressed by the pBR322 derivatives and by a plasmid which contained the hly genes under the control of a phage T7 promoter and the T7 RNA polymerase. When strongly overexpressed the 160K protein was released by E. coli cells into the extracellular medium concomitant with hemolytic activity. The genes encoding the 61K and the 160K proteins were transcribed in the same direction. Mutants expressing a 160K protein truncated at the carboxyl-terminal end were partially hemolytic. Hemolysis was progressively inhibited by saccharides with increasing molecular weights from maltotriose (M/sub r/ 504) to maltoheptaose (M/sub r/ 1152) and as totally abolished by dextran 4 (M/sub r/ 4000). This result and the observed influx of [ 14 C]sucrose into erythrocytes in the presence of hemolytic E. coli transformants under osmotically protective conditions suggest the formation of defined transmembrane channels by the hemolysin

  13. Two mechanisms coordinate replication termination by the Escherichia coli Tus-Ter complex.

    Science.gov (United States)

    Pandey, Manjula; Elshenawy, Mohamed M; Jergic, Slobodan; Takahashi, Masateru; Dixon, Nicholas E; Hamdan, Samir M; Patel, Smita S

    2015-07-13

    The Escherichia coli replication terminator protein (Tus) binds to Ter sequences to block replication forks approaching from one direction. Here, we used single molecule and transient state kinetics to study responses of the heterologous phage T7 replisome to the Tus-Ter complex. The T7 replisome was arrested at the non-permissive end of Tus-Ter in a manner that is explained by a composite mousetrap and dynamic clamp model. An unpaired C(6) that forms a lock by binding into the cytosine binding pocket of Tus was most effective in arresting the replisome and mutation of C(6) removed the barrier. Isolated helicase was also blocked at the non-permissive end, but unexpectedly the isolated polymerase was not, unless C(6) was unpaired. Instead, the polymerase was blocked at the permissive end. This indicates that the Tus-Ter mechanism is sensitive to the translocation polarity of the DNA motor. The polymerase tracking along the template strand traps the C(6) to prevent lock formation; the helicase tracking along the other strand traps the complementary G(6) to aid lock formation. Our results are consistent with the model where strand separation by the helicase unpairs the GC(6) base pair and triggers lock formation immediately before the polymerase can sequester the C(6) base. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  14. Two mechanisms coordinate replication termination by the Escherichia coli Tus–Ter complex

    KAUST Repository

    Pandey, Manjula

    2015-07-13

    The Escherichia coli replication terminator protein (Tus) binds to Ter sequences to block replication forks approaching from one direction. Here, we used single molecule and transient state kinetics to study responses of the heterologous phage T7 replisome to the Tus–Ter complex. The T7 replisome was arrested at the non-permissive end of Tus–Ter in a manner that is explained by a composite mousetrap and dynamic clamp model. An unpaired C(6) that forms a lock by binding into the cytosine binding pocket of Tus was most effective in arresting the replisome and mutation of C(6) removed the barrier. Isolated helicase was also blocked at the non-permissive end, but unexpectedly the isolated polymerase was not, unless C(6) was unpaired. Instead, the polymerase was blocked at the permissive end. This indicates that the Tus–Ter mechanism is sensitive to the translocation polarity of the DNA motor. The polymerase tracking along the template strand traps the C(6) to prevent lock formation; the helicase tracking along the other strand traps the complementary G(6) to aid lock formation. Our results are consistent with the model where strand separation by the helicase unpairs the GC(6) base pair and triggers lock formation immediately before the polymerase can sequester the C(6) base.

  15. Peptide ligands specific to the oxidized form of escherichia coli thioredoxin.

    Energy Technology Data Exchange (ETDEWEB)

    Scholle, M. D.; Banach, B. S.; Hamdan, S. M.; Richardson, C. C.; Kay, B. K.; Biosciences Division; Amunix, Inc.; Univ. of Illinois at Chicago; Harvard Medical School

    2008-11-01

    Thioredoxin (Trx) is a highly conserved redox protein involved in several essential cellular processes. In this study, our goal was to isolate peptide ligands to Escherichia coli Trx that mimic protein-protein interactions, specifically the T7 polymerase-Trx interaction. To do this, we subjected Trx to affinity selection against a panel of linear and cysteine-constrained peptides using M13 phage display. A novel cyclized conserved peptide sequence, with a motif of C(D/N/S/T/G)D(S/T)-hydrophobic-C-X-hydrophobic-P, was isolated to Trx. These peptides bound specifically to the E. coli Trx when compared to the human and spirulina homologs. An alanine substitution of the active site cysteines (CGPC) resulted in a significant loss of peptide binding affinity to the Cys-32 mutant. The peptides were also characterized in the context of Trx's role as a processivity factor of the T7 DNA polymerase (gp5). As the interaction between gp5 and Trx normally takes place under reducing conditions, which might interfere with the conformation of the disulfide-bridged peptides, we made use of a 22 residue deletion mutant of gp5 in the thioredoxin binding domain (gp5{Delta}22) that bypassed the requirements of reducing conditions to interact with Trx. A competition study revealed that the peptide selectively inhibits the interaction of gp5{Delta}22 with Trx, under oxidizing conditions, with an IC50 of {approx} 10 {micro}M.

  16. PATHOGENIC POTENTIALS OF ESCHERICHIA COLI ISOLATED ...

    African Journals Online (AJOL)

    Electrolyte and haematological parameters in rabbits infected with pathogenic isolates of Escherichia coli from rural water supplies in Rivers State, Nigeria, where monitored. Rabbits were orally infected with suspension containing 3x107 cfu /ml of Escherichia coli to induce diarrhoea, and the electrolyte (sodium, potassium ...

  17. Multiplex polymerase chain reaction for identification of Escherichia coli, Escherichia albertii and Escherichia fergusonii.

    Science.gov (United States)

    Lindsey, Rebecca L; Garcia-Toledo, L; Fasulo, D; Gladney, L M; Strockbine, N

    2017-09-01

    Escherichia coli, Escherichia albertii, and Escherichia fergusonii are closely related bacteria that can cause illness in humans, such as bacteremia, urinary tract infections and diarrhea. Current identification strategies for these three species vary in complexity and typically rely on the use of multiple phenotypic and genetic tests. To facilitate their rapid identification, we developed a multiplex PCR assay targeting conserved, species-specific genes. We used the Daydreamer™ (Pattern Genomics, USA) software platform to concurrently analyze whole genome sequence assemblies (WGS) from 150 Enterobacteriaceae genomes (107 E. coli, 5 Shigella spp., 21 E. albertii, 12 E. fergusonii and 5 other species) and design primers for the following species-specific regions: a 212bp region of the cyclic di-GMP regulator gene (cdgR, AW869_22935 from genome K-12 MG1655, CP014225) for E. coli/Shigella; a 393bp region of the DNA-binding transcriptional activator of cysteine biosynthesis gene (EAKF1_ch4033 from genome KF1, CP007025) for E. albertii; and a 575bp region of the palmitoleoyl-acyl carrier protein (ACP)-dependent acyltransferase (EFER_0790 from genome ATCC 35469, CU928158) for E. fergusonii. We incorporated the species-specific primers into a conventional multiplex PCR assay and assessed its performance with a collection of 97 Enterobacteriaceae strains. The assay was 100% sensitive and specific for detecting the expected species and offers a quick and accurate strategy for identifying E. coli, E. albertii, and E. fergusonii in either a single reaction or by in silico PCR with sequence assemblies. Published by Elsevier B.V.

  18. Transiting exoplanets from the CoRoT space mission. VIII. CoRoT-7b: the first super-Earth with measured radius

    Science.gov (United States)

    Léger, A.; Rouan, D.; Schneider, J.; Barge, P.; Fridlund, M.; Samuel, B.; Ollivier, M.; Guenther, E.; Deleuil, M.; Deeg, H. J.; Auvergne, M.; Alonso, R.; Aigrain, S.; Alapini, A.; Almenara, J. M.; Baglin, A.; Barbieri, M.; Bruntt, H.; Bordé, P.; Bouchy, F.; Cabrera, J.; Catala, C.; Carone, L.; Carpano, S.; Csizmadia, Sz.; Dvorak, R.; Erikson, A.; Ferraz-Mello, S.; Foing, B.; Fressin, F.; Gandolfi, D.; Gillon, M.; Gondoin, Ph.; Grasset, O.; Guillot, T.; Hatzes, A.; Hébrard, G.; Jorda, L.; Lammer, H.; Llebaria, A.; Loeillet, B.; Mayor, M.; Mazeh, T.; Moutou, C.; Pätzold, M.; Pont, F.; Queloz, D.; Rauer, H.; Renner, S.; Samadi, R.; Shporer, A.; Sotin, Ch.; Tingley, B.; Wuchterl, G.; Adda, M.; Agogu, P.; Appourchaux, T.; Ballans, H.; Baron, P.; Beaufort, T.; Bellenger, R.; Berlin, R.; Bernardi, P.; Blouin, D.; Baudin, F.; Bodin, P.; Boisnard, L.; Boit, L.; Bonneau, F.; Borzeix, S.; Briet, R.; Buey, J.-T.; Butler, B.; Cailleau, D.; Cautain, R.; Chabaud, P.-Y.; Chaintreuil, S.; Chiavassa, F.; Costes, V.; Cuna Parrho, V.; de Oliveira Fialho, F.; Decaudin, M.; Defise, J.-M.; Djalal, S.; Epstein, G.; Exil, G.-E.; Fauré, C.; Fenouillet, T.; Gaboriaud, A.; Gallic, A.; Gamet, P.; Gavalda, P.; Grolleau, E.; Gruneisen, R.; Gueguen, L.; Guis, V.; Guivarc'h, V.; Guterman, P.; Hallouard, D.; Hasiba, J.; Heuripeau, F.; Huntzinger, G.; Hustaix, H.; Imad, C.; Imbert, C.; Johlander, B.; Jouret, M.; Journoud, P.; Karioty, F.; Kerjean, L.; Lafaille, V.; Lafond, L.; Lam-Trong, T.; Landiech, P.; Lapeyrere, V.; Larqué, T.; Laudet, P.; Lautier, N.; Lecann, H.; Lefevre, L.; Leruyet, B.; Levacher, P.; Magnan, A.; Mazy, E.; Mertens, F.; Mesnager, J.-M.; Meunier, J.-C.; Michel, J.-P.; Monjoin, W.; Naudet, D.; Nguyen-Kim, K.; Orcesi, J.-L.; Ottacher, H.; Perez, R.; Peter, G.; Plasson, P.; Plesseria, J.-Y.; Pontet, B.; Pradines, A.; Quentin, C.; Reynaud, J.-L.; Rolland, G.; Rollenhagen, F.; Romagnan, R.; Russ, N.; Schmidt, R.; Schwartz, N.; Sebbag, I.; Sedes, G.; Smit, H.; Steller, M. B.; Sunter, W.; Surace, C.; Tello, M.; Tiphène, D.; Toulouse, P.; Ulmer, B.; Vandermarcq, O.; Vergnault, E.; Vuillemin, A.; Zanatta, P.

    2009-10-01

    Aims: We report the discovery of very shallow (Δ F/F ≈ 3.4× 10-4), periodic dips in the light curve of an active V = 11.7 G9V star observed by the CoRoT satellite, which we interpret as caused by a transiting companion. We describe the 3-colour CoRoT data and complementary ground-based observations that support the planetary nature of the companion. Methods: We used CoRoT colours information, good angular resolution ground-based photometric observations in- and out- of transit, adaptive optics imaging, near-infrared spectroscopy, and preliminary results from radial velocity measurements, to test the diluted eclipsing binary scenarios. The parameters of the host star were derived from optical spectra, which were then combined with the CoRoT light curve to derive parameters of the companion. Results: We examined all conceivable cases of false positives carefully, and all the tests support the planetary hypothesis. Blends with separation >0.40´´or triple systems are almost excluded with a 8 × 10-4 risk left. We conclude that, inasmuch we have been exhaustive, we have discovered a planetary companion, named CoRoT-7b, for which we derive a period of 0.853 59 ± 3 × 10-5 day and a radius of Rp = 1.68 ± 0.09 R_Earth. Analysis of preliminary radial velocity data yields an upper limit of 21 M_Earth for the companion mass, supporting the finding. Conclusions: CoRoT-7b is very likely the first Super-Earth with a measured radius. This object illustrates what will probably become a common situation with missions such as Kepler, namely the need to establish the planetary origin of transits in the absence of a firm radial velocity detection and mass measurement. The composition of CoRoT-7b remains loosely constrained without a precise mass. A very high surface temperature on its irradiated face, ≈1800-2600 K at the substellar point, and a very low one, ≈50 K, on its dark face assuming no atmosphere, have been derived. The CoRoT space mission, launched on 27

  19. Photoreactivating enzyme from Escherichia coli

    International Nuclear Information System (INIS)

    Snapka, R.M.; Fuselier, C.O.

    1977-01-01

    Escherichia coli photoreactivating enzyme (PRE) has been purified in large amounts from an E.coli strain lysogenic for a defective lambda bacteriophage carrying the phr gene. The resulting enzyme had a pH optimum of 7.2 and an ionic strength optimum of 0.18. It consisted of an apoprotein and cofactor, both of which were necessary for catalytic activity. The apoprotein had a monomer molecular weight of 35,200 and showed stable aggregates under denaturing conditions. The amino acid analysis of the E.coli enzyme was very similar to that of the photoreactivating enzyme from orchid seedlings (Cattelya aurantiaca). Both had arginine at the amino terminus. The cofactor, like the holoenzyme, showed absorption, magnetic circular dichroism, and emission properties indicative of an adenine moiety. Although the isolated enzyme had an action spectrum which peaked at about 360 nm, neither the cofactor, apoenzyme nor holoenzyme showed any detectable absorption between 300 and 400 nm. (author)

  20. Genomics of Escherichia and Shigella

    Science.gov (United States)

    Perna, Nicole T.

    The laboratory workhorse Escherichia coli K-12 is among the most intensively studied living organisms on earth, and this single strain serves as the model system behind much of our understanding of prokaryotic molecular biology. Dense genome sequencing and recent insightful comparative analyses are making the species E. coli, as a whole, an emerging system for studying prokaryotic population genetics and the relationship between system-scale, or genome-scale, molecular evolution and complex traits like host range and pathogenic potential. Genomic perspective has revealed a coherent but dynamic species united by intraspecific gene flow via homologous lateral or horizontal transfer and differentiated by content flux mediated by acquisition of DNA segments from interspecies transfers.

  1. Photoreactivating enzyme from Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Snapka, R M; Fuselier, C O [California Univ., Irvine (USA)

    1977-05-01

    Escherichia coli photoreactivating enzyme (PRE) has been purified in large amounts from an E.coli strain lysogenic for a defective lambda bacteriophage carrying the phr gene. The resulting enzyme had a pH optimum of 7.2 and an ionic strength optimum of 0.18. It consisted of an apoprotein and cofactor, both of which were necessary for catalytic activity. The apoprotein had a monomer molecular weight of 35,200 and showed stable aggregates under denaturing conditions. The amino acid analysis of the E.coli enzyme was very similar to that of the photoreactivating enzyme from orchid seedlings (Cattelya aurantiaca). Both had arginine at the amino terminus. The cofactor, like the holoenzyme, showed absorption, magnetic circular dichroism, and emission properties indicative of an adenine moiety. Although the isolated enzyme had an action spectrum which peaked at about 360 nm, neither the cofactor, apoenzyme nor holoenzyme showed any detectable absorption between 300 and 400 nm.

  2. Hydrolysis of R7T7 nuclear waste glass in dilute media: mechanisms and rate as a function of pH

    International Nuclear Information System (INIS)

    Advocat, T.; Vernaz, E.; Charpentier, H.; Crovisier, J.L.; Ehret, G.

    1990-01-01

    R7T7 nuclear waste glass dissolution in highly dilute aqueous media under static conditions at 90 0 C occurs according to two different mechanisms depending on the solution acidity. In acid media (pH 4.8 and 5.5), preferential extraction of glass network modifiers results in the formation of an alkali metal-depleted surface region on which amorphous and crystallized (phosphate) compounds rich in transition elements precipitate. Steady-state dissolution conditions are not reached, as attested by variable normalized Si, B and Na mass losses. Glass dissolution is stoichiometric in basic media (pH 7 to 10): the strong bonds of the silicated network are broken at a rate that increases with the pH: the glass dissolution rate increases by a factor of 15 between pH 7 and 10. Under these conditions, alteration products at the glass/solution interface do not constitute a short-term kinetic barrier against the release of the major glass components

  3. Experimental alteration of R7T7 nuclear model glass in solutions with different salinities (90/sup 0/C, 1 bar): implications for the selection of geological repositories

    Energy Technology Data Exchange (ETDEWEB)

    Godon, N.; Thomassin, J.H.; Touray, J.C.; Vernaz, E.

    1988-01-01

    In order to simulate the leaching of nuclear wastes in repositories percolated by solutions of variable salinity, leaching tests of R7T7 glass in solutions with different NaCl contents have been performed at 90/sup 0/C and 1 bar using a static procedure. A comparison of the efficiency of the different leachants indicated that the alteration was maximum in pure water and in 23.7 g (NaCl) kg/sup -1/ solution. In deionized water, uranium- and rare-earth elements simulating the actinides were found quite immobile: they have not been detected in solution but are present in the alteration layer. On the other hand, in the 23.7 g (NaCl) kg/sup -1/ solution, high amounts of uranium, cerium and neodymium have been detected in solution and did not accumulate in the solid phases. In the highest salinity brines, the bulk reactivity of the glass decreased. In all leachants, the alteration layer was structured in two parts: hydrated glass and flakes. The flakes were mainly nickel-and zinc-bearing aluminosilicate phases. When crystallized, the flakes were identified as berthierine.

  4. Experimental investigation of aqueous corrosion of R7T7 nuclear glass at 90 degrees C in the presence of humic acids: A kinetic approach

    International Nuclear Information System (INIS)

    Gin, S.; Godon, N.; Mestre, J.P.; Vernaz, E.Y.; Beaufort, D.

    1994-01-01

    The dissolution kinetics of the French open-quotes R7T7close quotes nonradioactive LWR reference glass in solutions containing dissolved humic acids were investigated at 9O degrees C during static tests with imposed or free pH. Experiments conducted in highly dilute media, with a glass-surface-area-to-solution-volume (SA/V) ratio of 5 m -1 , showed that the glass dissolution surface reaction is catalyzed by humic acids. With higher degrees of reaction progress (SA/V = 100 m -1 and free pH) the humic acids impose pH modifications on the system compared with inorganic media; moreover, they directly or indirectly enhance the dissolution of certain alkali metals and transition elements, forming aqueous complexes with the latter. During experiments with an imposed pH of 8.5 (SA/V = 1300 and 5300 m -1 ), the humic acids appear to cause increased silica solubility that cannot be accounted for by the formation of silica complexes. A residual corrosion rate in the humic acid media exceeding the rate measured in inorganic media suggests that, in addition to silica, one or more element complexes formed by humic acids may be a kinetically limiting factor. This hypothesis must be confirmed, however, as the quantity of humic acids per unit glass surface area was too small in this experiment to allow unambiguous characterization of the phenomenon

  5. Construction of pRMES and pTMES plasmids to test their expression ability of Nanobodies via the efficient T7 promoter

    International Nuclear Information System (INIS)

    Masoud, H.; Quider, M.; Abbady, A.

    2014-01-01

    Nanobody technology is considered as a promising molecular biology technique performed by means of the genetic engineering of special type of antibodies, existing exclusively in Camelidea. It enables the obtaining of small proteins, referred to as Nanobodies, which are characterized by high stability and solubility, are able to link to their specific antigens. After production, the Nanobody genes are cloned within plasmids of protein expression in bacteria, allowing their stable and continuous production for research and applied purposes. This work aimed to design new plasmids for Nanobody genes cloning in order to ensure a strong expression via the efficient T7 promoter, thus enhancing the quantity of the produced Nanobodies. These plasmids were called pRMES and pTMES and their ability to express Nanobodies, NbBruc02 and Nb16M, was tested. The plasmid pTMES showed an enhanced production condition of this Nanobody. These new plasmids, by their variable characteristics, could represent efficient tools for general production of recombinant proteins, including Nanobodies (author).

  6. Lepton mixing and cancellation of the Dirac mass hierarchy in SO(10) GUTs with flavor symmetries T7 and Σ(81)

    International Nuclear Information System (INIS)

    Hagedorn, Claudia; Schmidt, Michael A.; Smirnov, Alexei Yu.

    2009-01-01

    In SO(10) grand unified theories the hierarchy which is present in the Dirac mass term of the neutrinos is generically as strong as the one in the up-type quark mass term. We propose a mechanism to partially or completely cancel this hierarchy in the light neutrino mass matrix in the seesaw context. The two main ingredients of the cancellation mechanism are the existence of three fermionic gauge singlets and of a discrete flavor symmetry G f which is broken at a higher scale than SO(10). Two realizations of the cancellation mechanism are presented. The realization based on the Frobenius group T 7 ≅Z 7 xZ 3 leads to a partial cancellation of the hierarchy and relates maximal 2-3 lepton mixing with the geometric hierarchy of the up-quark masses. In the realization with the group Σ(81) the cancellation is complete and tribimaximal lepton mixing is reproduced at the lowest order. In both cases, to fully accommodate the leptonic data we take into account additional effects such as effects of higher-dimensional operators involving more than one flavon. The heavy neutral fermion mass spectra are considered. For both realizations we analyze the flavon potential at the renormalizable level as well as ways to generate the Cabibbo angle.

  7. R7T7 glass alteration mechanism in an aqueous closed system: understanding and modelling the long term alteration kinetic; Etude des mecanismes d'alteration par l'eau du verre R7T7 en milieu confine: comprehension et modelisation de la cinetique residuelle

    Energy Technology Data Exchange (ETDEWEB)

    Chave, T

    2007-10-15

    The long term alteration rate of the French R7T7 nuclear glass has been investigated since many years because it will define the overall resistance of the radionuclide containment matrix. Recent studies have shown that the final rate remains constant or is slightly decreasing with time. It never reaches zero. Though this residual rate is very low, only 5 nm per year at 50 C, it would be the dominant alteration phenomenon in a geological repository. Two mechanisms are suggested for explaining such behaviour: diffusion in solution of elements from glass through an amorphous altered layer and precipitation of neo-formed phases. The diffusion processes are in agreement with a solid state diffusion mechanism and can lead to secondary phase precipitation due to solution concentration increases. Observed phases are mainly phyllosilicates and zeolites, in specific conditions. Phyllosilicates are expected to maintain the residual kinetic rate whereas alteration resumption could be observed in presence of zeolites at very high pH or temperature (10.5 at 90 C or temperature above 150 C). Both diffusion and neo-formed phase precipitation have been investigated in order to better understand their impact on the residual alteration rate and have then been modelled by a calculation code, coupling chemistry and transport, in order to be able to better anticipate the long term behaviour of the glass R7T7 in an aqueous closed system. (author)

  8. Method for determining transcriptional linkage by means of inhibition of deoxyribonucleic acid transcription by ultraviolet irradiation: evaluation in application to the investigation of in vivo transcription in bacteriophage T7

    International Nuclear Information System (INIS)

    Brautigam, A.R.

    1975-01-01

    A technique is presented for mapping promotor sites that utilizes the introduction of transcription-terminating lesions in DNA through uv irradiation which prevents transcription of genes in proportion to their distance from the promotor. This technique was applied to and evaluated in investigations of the transcriptional linkage of bacteriophage T7. All results substantiate the hypothesis that transcription in vivo does not proceed beyond the first uv lesion encountered in the template DNA and that such premature termination of transcription is the principal effect of the uv irradiation on the transcriptional template function of DNA. UV-induced inhibition of the initiation of transcription is insignificant by comparison. Uv inactivation of expression of individual T7 genes was found to follow pseudo first-order kinetics, allowing a gene-specific uv inactivation cross section to be evaluated for each gene. Promotor locations were inferred from the discontinuity in the numerical values of inactivation cross sections arising at the start of each new unit. By such analysis the bacteriophage T7 genome was found to consist of seven transcription units. In vivo E. coli RNA polymerase transcribes the T7 early region as a single unit from a pomotor region located at the left end of the genome. The T7 late region was found to consist of six transcription units, with promotors located just ahead of genes 1.7, 7, 9, 11, 13 and 17

  9. (ESBL) producing Escherichia coli and Klebsiella pneumoniae

    African Journals Online (AJOL)

    use

    2011-11-21

    Nov 21, 2011 ... the most common serious bacterial infections in infants ... UTI is a common cause of morbidity .... of ESBL and non-ESBL producing Escherichia coli and Klebsiella pneumonia. ... in hospital and community acquired infections.

  10. Characterization of Escherichia coli Phylogenetic Groups ...

    African Journals Online (AJOL)

    tract infection (UTI), bacteremia, pneumonia, soft-tissue infection, and ... Keywords: Drug resistance, Escherichia coli, Extraintestinal infections, Polymerase chain reaction, .... gynecology, 12 from orthopedics and 5 from pediatrics units.

  11. escherichia coli serotypes confirmed in experimental mammary ...

    African Journals Online (AJOL)

    DJFLEX

    VARIATIONS IN VIRULENCE OF THREE (3) ESCHERICHIA COLI. SEROTYPES CONFIRMED IN ... ows are susceptible to E. coli infection because. E. coli exist in the .... Coli infections in mice: A laboratory animal model for research in.

  12. Frameshift mutations in infectious cDNA clones of Citrus tristeza virus: a strategy to minimize the toxicity of viral sequences to Escherichia coli

    International Nuclear Information System (INIS)

    Satyanarayana, Tatineni; Gowda, Siddarame; Ayllon, Maria A.; Dawson, William O.

    2003-01-01

    The advent of reverse genetics revolutionized the study of positive-stranded RNA viruses that were amenable for cloning as cDNAs into high-copy-number plasmids of Escherichia coli. However, some viruses are inherently refractory to cloning in high-copy-number plasmids due to toxicity of viral sequences to E. coli. We report a strategy that is a compromise between infectivity of the RNA transcripts and toxicity to E. coli effected by introducing frameshift mutations into 'slippery sequences' near the viral 'toxicity sequences' in the viral cDNA. Citrus tristeza virus (CTV) has cDNA sequences that are toxic to E. coli. The original full-length infectious cDNA of CTV and a derivative replicon, CTV-ΔCla, cloned into pUC119, resulted in unusually limited E. coli growth. However, upon sequencing of these cDNAs, an additional uridinylate (U) was found in a stretch of U's between nts 3726 and 3731 that resulted in a change to a reading frame with a stop codon at nt 3734. Yet, in vitro produced RNA transcripts from these clones infected protoplasts, and the resulting progeny virus was repaired. Correction of the frameshift mutation in the CTV cDNA constructs resulted in increased infectivity of in vitro produced RNA transcripts, but also caused a substantial increase of toxicity to E. coli, now requiring 3 days to develop visible colonies. Frameshift mutations created in sequences not suspected to facilitate reading frame shifting and silent mutations introduced into oligo(U) regions resulted in complete loss of infectivity, suggesting that the oligo(U) region facilitated the repair of the frameshift mutation. Additional frameshift mutations introduced into other oligo(U) regions also resulted in transcripts with reduced infectivity similarly to the original clones with the +1 insertion. However, only the frameshift mutations introduced into oligo(U) regions that were near and before the toxicity region improved growth and stability in E. coli. These data demonstrate that

  13. Purification and characterization of the d-xylose isomerase gene from Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Ho, N W.Y.; Rosenfeld, S; Stevis, P; Tsao, G T

    1983-11-01

    A DNA fragment containing both the Escherichia coli D-xylose isomerase (D-xylose ketol-isomerase, EC 5.3.1.5) gene and the D-xylulokinase (ATP: D-xylulose 5-phosphotransferase, EC 2.7.1.17) gene has been cloned on an E. coli plasmid. The D-xylose isomerase gene was separated from the D-xylulokinase gene by the construction of a new deletion plasmid, pLX7. The D-xylose isomerase gene cloned on pLX7 was found still to be an intact gene. The precise location of the D-xylose isomerase gene on the plasmid pLX7 was further determined by the construction of two more plasmids, pLX8 and pLX9. This is believed to be the first D-xylose isomerase gene that has been isolated and extensively purified from any organism. D-Xylose isomerase, the enzyme product of the D-xylose isomerase gene, is responsible for the conversion of D-xylose to D-xylulose, as well as D-glucose to D-fructose. It is widely believed that yeast cannot ferment D-xylose to ethanol primarily because of the lack of D-xylose isomerase in yeast. D-Xylose isomerase (also known as D-glucose isomerase) is also used for the commercial production of high-fructose syrups. The purification of the D-xylose isomerase gene may lead to the following industrial applications: (1) cloning and expression of the gene in yeast to make the latter organism capable of directly fermenting D-xylose to ethanol, and (2) cloning of the gene on a high-copy-number plasmid in a proper host to overproduce the enzyme, which should have a profound impact on the high-fructose syrup technology. 14 references.

  14. Peptidoglycan Hydrolases of Escherichia coli

    Science.gov (United States)

    van Heijenoort, Jean

    2011-01-01

    Summary: The review summarizes the abundant information on the 35 identified peptidoglycan (PG) hydrolases of Escherichia coli classified into 12 distinct families, including mainly glycosidases, peptidases, and amidases. An attempt is also made to critically assess their functions in PG maturation, turnover, elongation, septation, and recycling as well as in cell autolysis. There is at least one hydrolytic activity for each bond linking PG components, and most hydrolase genes were identified. Few hydrolases appear to be individually essential. The crystal structures and reaction mechanisms of certain hydrolases having defined functions were investigated. However, our knowledge of the biochemical properties of most hydrolases still remains fragmentary, and that of their cellular functions remains elusive. Owing to redundancy, PG hydrolases far outnumber the enzymes of PG biosynthesis. The presence of the two sets of enzymes acting on the PG bonds raises the question of their functional correlations. It is difficult to understand why E. coli keeps such a large set of PG hydrolases. The subtle differences in substrate specificities between the isoenzymes of each family certainly reflect a variety of as-yet-unidentified physiological functions. Their study will be a far more difficult challenge than that of the steps of the PG biosynthesis pathway. PMID:22126997

  15. Partitioning of the rare earths and actinides between R7T7 nuclear glass alteration products and solution according to disposal conditions; Partage des terres rares et des actinides entre solution et produits d`alteration du verre nucleaire type R7T7 en fonction des conditions de stockage

    Energy Technology Data Exchange (ETDEWEB)

    Menard, O

    1995-10-25

    The alteration of nuclear glass by water is liable to release radionuclides into the environment. Determining the release kinetics of these elements and their aqueous chemical forms are therefore essential steps in establishing the safety of a geological repository site. Leach tests were conducted with a nonradioactive specimen of the French ``R7T7`` light water containment glass spiked with U and Th, and with two R7T7 specimens spiked with {sup 237}Np and {sup 239}Pu, respectively. The alteration solution compositions were representative of deep groundwater and contained carbonate, sulfate, phosphate, fluorine and chlorine ions. The release of U, Th, Np and Pu, as well as of the rare earths La, Ce and Nd were monitored by ICP mass spectrometry and by {alpha} spectrometry. Scanning and transmission electron microscopic examination of the nonradioactive altered glass surfaces was also performed to assess the partitioning balance for the rare earths, U and Th between the glass alteration products and solution. The mobility of these elements depends on two competing mechanisms. The rare earths and thorium are incorporated in the alteration products (gel); the retention process is assumed to involve chemisorption or coprecipitation, enhanced in the gel layer by the presence of phosphate ions in particular. Conversely, the aqueous species in the alteration solutions (mainly anions) form complexes with the actinides and rare earths; this phenomenon is particularly evident with U and Np. The presence of carbonate ions favors this mobility. Plutonium differs from U and Np in that it is adsorbed mainly on colloids formed by glass dissolution, the principal factors governing its chemical evolution in solution. (author). refs., 122 figs., 185 tabs.

  16. Analysis of whole genome sequencing for the Escherichia coli O157:H7 typing phages.

    Science.gov (United States)

    Cowley, Lauren A; Beckett, Stephen J; Chase-Topping, Margo; Perry, Neil; Dallman, Tim J; Gally, David L; Jenkins, Claire

    2015-04-08

    Shiga toxin producing Escherichia coli O157 can cause severe bloody diarrhea and haemolytic uraemic syndrome. Phage typing of E. coli O157 facilitates public health surveillance and outbreak investigations, certain phage types are more likely to occupy specific niches and are associated with specific age groups and disease severity. The aim of this study was to analyse the genome sequences of 16 (fourteen T4 and two T7) E. coli O157 typing phages and to determine the genes responsible for the subtle differences in phage type profiles. The typing phages were sequenced using paired-end Illumina sequencing at The Genome Analysis Centre and the Animal Health and Veterinary Laboratories Agency and bioinformatics programs including Velvet, Brig and Easyfig were used to analyse them. A two-way Euclidian cluster analysis highlighted the associations between groups of phage types and typing phages. The analysis showed that the T7 typing phages (9 and 10) differed by only three genes and that the T4 typing phages formed three distinct groups of similar genomic sequences: Group 1 (1, 8, 11, 12 and 15, 16), Group 2 (3, 6, 7 and 13) and Group 3 (2, 4, 5 and 14). The E. coli O157 phage typing scheme exhibited a significantly modular network linked to the genetic similarity of each group showing that these groups are specialised to infect a subset of phage types. Sequencing the typing phage has enabled us to identify the variable genes within each group and to determine how this corresponds to changes in phage type.

  17. 77 FR 9888 - Shiga Toxin-Producing Escherichia coli

    Science.gov (United States)

    2012-02-21

    ... Toxin-Producing Escherichia coli in Certain Raw Beef Products AGENCY: Food Safety and Inspection Service... toxin-producing Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, and O145). This new date..., that are contaminated with Shiga toxin-producing Escherichia coli (STEC) O26, O45, O103, O111, O121...

  18. 21 CFR 866.3255 - Escherichia coli serological reagents.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Escherichia coli serological reagents. 866.3255... coli serological reagents. (a) Identification. Escherichia coli serological reagents are devices that consist of antigens and antisera used in serological tests to identify Escherichia coli from cultured...

  19. Alteration of 'R7T7' type nuclear glasses: statistical approach, experimental validation, local evolution model; Alteration des verres nucleaires de type 'R7T7': demarche statistique, validation experimentale, modele local d'evolution

    Energy Technology Data Exchange (ETDEWEB)

    Thierry, F

    2003-02-01

    The aim of this work is to propose an evolution of nuclear (R7T7-type) glass alteration modeling. The first part of this thesis is about development and validation of the 'r(t)' model. This model which predicts the decrease of alteration rates in confined conditions is based upon a coupling between a first-order dissolution law and a diffusion barrier effect of the alteration gel layer. The values and the uncertainties regarding the main adjustable parameters of the model ({alpha}, Dg and C*) have been determined from a systematic study of the available experimental data. A program called INVERSION has been written for this purpose. This work lead to characterize the validity domain of the 'r(t)' model and to parametrize it. Validation experiments have been undertaken, confirming the validity of the parametrization over 200 days. A new model is proposed in the second part of this thesis. It is based on an inhibition of glass dissolution reaction by silicon coupled with a local description of silicon retention in the alteration gel layer. This model predicts the evolutions of boron and silicon concentrations in solution as well as the concentrations and retention profiles in the gel layer. These predictions have been compared to measurements of retention profiles by the secondary ion mass spectrometry (SIMS) method. The model has been validated on fractions of gel layer which reactivity present low or moderate disparities. (author)

  20. Quality control of inclusion bodies in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Schweder Thomas

    2010-05-01

    Full Text Available Abstract Background Bacterial inclusion bodies (IBs are key intermediates for protein production. Their quality affects the refolding yield and further purification. Recent functional and structural studies have revealed that IBs are not dead-end aggregates but undergo dynamic changes, including aggregation, refunctionalization of the protein and proteolysis. Both, aggregation of the folding intermediates and turnover of IBs are influenced by the cellular situation and a number of well-studied chaperones and proteases are included. IBs mostly contain only minor impurities and are relatively homogenous. Results IBs of α-glucosidase of Saccharomyces cerevisiae after overproduction in Escherichia coli contain a large amount of (at least 12 different major product fragments, as revealed by two-dimensional polyacrylamide gel electrophoresis (2D PAGE. Matrix-Assisted-Laser-Desorption/Ionization-Time-Of-Flight Mass-Spectrometry (MALDI-ToF MS identification showed that these fragments contain either the N- or the C-terminus of the protein, therefore indicate that these IBs are at least partially created by proteolytic action. Expression of α-glucosidase in single knockout mutants for the major proteases ClpP, Lon, OmpT and FtsH which are known to be involved in the heat shock like response to production of recombinant proteins or to the degradation of IB proteins, clpP, lon, ompT, and ftsH did not influence the fragment pattern or the composition of the IBs. The quality of the IBs was also not influenced by the sampling time, cultivation medium (complex and mineral salt medium, production strategy (shake flask, fed-batch fermentation process, production strength (T5-lac or T7 promoter, strain background (K-12 or BL21, or addition of different protease inhibitors during IB preparation. Conclusions α-glucosidase is fragmented before aggregation, but neither by proteolytic action on the IBs by the common major proteases, nor during downstream IB

  1. Altered membrane permeability in multidrug resistant Escherichia ...

    African Journals Online (AJOL)

    PRECIOUS

    2009-11-02

    Nov 2, 2009 ... involvement during the transport of β - lactams in multidrug resistant Escherichia coli isolated from extra-intestinal infections. Also, the ... lactam resistance in multidrug resistant E. coli in ESBL and non-ESBL isolates. .... and decreased susceptibility to carbapenems, particularly ertapenem (Perez et al.,.

  2. Altered membrane permeability in multidrug resistant Escherichia ...

    African Journals Online (AJOL)

    The study was conducted with the objective of examining the outer membrane proteins and their involvement during the transport of β - lactams in multidrug resistant Escherichia coli isolated from extra-intestinal infections. Also, the response of gram negative bacterial biomembrane alteration was studied using extended ...

  3. Optimization of plasmid electrotransformation into Escherichia coli ...

    African Journals Online (AJOL)

    In order to improve electroporation, optical density of bacteria, recovery time and electrical parameter (field strength and capacitance) were optimized using the Taguchi statistical method. ANOVA of obtained data indicated that the optimal conditions of electrotransformation of pET-28a (+) plasmid into Escherichia coli ...

  4. Inhibition of Escherichia Coli, Salmonella and Staphylococcus ...

    African Journals Online (AJOL)

    Escherichia coli O157:H7, Salmonella typhimurium and Staphylococcus. aureus are of great concern to the food industry, especially in foods stored under refrigerated conditions where, unlike most food-borne pathogens are able to multiply. This investigation was conducted to study the inhibitory effect of some spice ...

  5. (ESBL) producing Escherichia coli and Klebsiella pneumoniae

    African Journals Online (AJOL)

    Emerging antibiotic resistance due to extended spectrum β-lactamase (ESBL) production limited the use of β-lactam antibiotics against Escherichia coli and Klebsiella pneumoniae. This observational study was conducted at the Microbiology department of the Children's Hospital, Lahore Pakistan, from June, 2009 to ...

  6. Antimicrobial resistance among commensal Escherichia coli from ...

    African Journals Online (AJOL)

    Commensal bacteria contribute to the distribution and persistence of antimicrobial resistance in the environment. This study monitored antimicrobial resistance in commensal Escherichia coli from the faeces of on-farm and slaughter cattle and beef. A total of 342 (89.5%) E. coli isolates were obtained from 382 samples.

  7. Characterization of Escherichia coli Phylogenetic Groups ...

    African Journals Online (AJOL)

    Background: Escherichia coli strains mainly fall into four phylogenetic groups (A, B1, B2, and D) and that virulent extra‑intestinal strains mainly belong to groups B2 and D. Aim: The aim was to determine the association between phylogenetic groups of E. coli causing extraintestinal infections (ExPEC) regarding the site of ...

  8. Mutagenic DNA repair in Escherichia coli

    International Nuclear Information System (INIS)

    Bridges, B.A.; Sharif, Firdaus

    1986-01-01

    The authors report a study of the misincorporation step in excision proficient umuC Escherichia coli as revealed by delayed photoreversal and show that it parallels the loss of photoreversibility of mutations induced in isogenic umu + bacteria; in both cases the end-point was mutation to streptomycin resistance. (author)

  9. Fosfomycin Resistance in Escherichia coli, Pennsylvania, USA.

    Science.gov (United States)

    Alrowais, Hind; McElheny, Christi L; Spychala, Caressa N; Sastry, Sangeeta; Guo, Qinglan; Butt, Adeel A; Doi, Yohei

    2015-11-01

    Fosfomycin resistance in Escherichia coli is rare in the United States. An extended-spectrum β-lactamase-producing E. coli clinical strain identified in Pennsylvania, USA, showed high-level fosfomycin resistance caused by the fosA3 gene. The IncFII plasmid carrying this gene had a structure similar to those found in China, where fosfomycin resistance is commonly described.

  10. Strategies for Protein Overproduction in Escherichia coli.

    Science.gov (United States)

    Mott, John E.

    1984-01-01

    Examines heterologous expression in Escherichia coli and the role of regulatory sequences which control gene expression at transcription resulting in abundant production of messenger RNA and regulatory sequences in mRNA which promote efficient translation. Also examines the role of E. coli cells in stabilizing mRNA and protein that is…

  11. Antibiotic resistance properties of uropathogenic Escherichia coli ...

    African Journals Online (AJOL)

    Purpose: To investigate the antibiotic resistance pattern of uropathogenic Escherichia coli (UPEC) strains isolated from pregnant women with history of recurrent urinary tract infections (RUTIs) and healthy pregnant women. Methods: A total of 485 high vaginal swab specimens were collected from pregnant women with ...

  12. Escherichia coli survival in waters: Temperature dependence

    Science.gov (United States)

    Knowing the survival rates of water-borne Escherichia coli is important in evaluating microbial contamination and making appropriate management decisions. E. coli survival rates are dependent on temperature, a dependency that is routinely expressed using an analogue of the Q10 mo...

  13. Comparison of 61 Sequenced Escherichia coli Genomes

    DEFF Research Database (Denmark)

    Lukjancenko, Oksana; Wassenaar, T. M.; Ussery, David

    2010-01-01

    Escherichia coli is an important component of the biosphere and is an ideal model for studies of processes involved in bacterial genome evolution. Sixty-one publically available E. coli and Shigella spp. sequenced genomes are compared, using basic methods to produce phylogenetic and proteomics...

  14. Antimicrobial resistance among commensal Escherichia coli from ...

    African Journals Online (AJOL)

    user1

    2012-07-19

    Jul 19, 2012 ... Commensal bacteria contribute to the distribution and persistence of antimicrobial resistance in the environment. This study monitored antimicrobial resistance in commensal Escherichia coli from the faeces of on-farm and slaughter cattle and beef. A total of 342 (89.5%) E. coli isolates were obtained.

  15. Prevalence of Arcobacter, Escherichia coli, Staphylococcus aureus ...

    African Journals Online (AJOL)

    In this study, varying level of resistance of Escherichia coli 66(84.6%), Salmonella 6(100%) and Arcobacter 57(100%) to amoxicillin was observed. The susceptibility pattern indicates that the bacterial isolates exhibited a varying level of resistance to two or more antimicrobial agents with maximum resistance to amoxicillin.

  16. Fimbrial adhesins from extraintestinal Escherichia coli

    DEFF Research Database (Denmark)

    Klemm, Per; Hancock, Viktoria; Schembri, Mark A.

    2010-01-01

    Extraintestinal pathogenic Escherichia coli (ExPEC) represent an important subclass of E. coli that cause a wide spectrum of diseases in human and animal hosts. Fimbriae are key virulence factors of ExPEC strains. These long surface located rod-shaped organelles mediate receptor-specific attachment...

  17. Leaner and meaner genomes in Escherichia coli

    DEFF Research Database (Denmark)

    Ussery, David

    2006-01-01

    A 'better' Escherichia coli K-12 genome has recently been engineered in which about 15% of the genome has been removed by planned deletions. Comparison with related bacterial genomes that have undergone a natural reduction in size suggests that there is plenty of scope for yet more deletions....

  18. lactamase in clinical isolates of Escherichia coli

    African Journals Online (AJOL)

    Jane

    2011-08-22

    Aug 22, 2011 ... The beta lactamase enzyme producing E. coli, resistant to β-lactam antibiotics, created many problems ... Key words: Escherichia coli, β-lactamase enzymes, TEM-type extended spectrum ... difficulties in treatment using antibiotics that are currently ... and chloramphenicol (30 µg) (Mast Diagnostics Ltd., UK).

  19. ANTIMICIROBIAL SUSCEPTIBILITY PATTERNS OF Escherichia coli ...

    African Journals Online (AJOL)

    DR. AMINU

    A total of 56 and 24 strains of E. coli and Shigella sp. isolated from children less than five years with diarrhoea attending 3 ... parasitic infections, as well as food intolerance, reaction to ..... Escherichia coil 0157:H7 as a model of entry of a new.

  20. The formation of an aberrant PAX5 transcript in a patient with mixed phenotype acute leukemia harboring der(9t(7;9(q11.2;p13

    Directory of Open Access Journals (Sweden)

    Jun Amaki

    2016-01-01

    Full Text Available We experienced the case of a 56-year-old male with B-lymphoid/myeloid lineage mixed phenotype acute leukemia (MPAL. A cytogenetic analysis of the patient's bone marrow revealed a complex karyotype, including der(9t(7;9(q11.2;p13. We identified an aberrant PAX5 transcript, including the exons 1A to 5 and the contiguous intron 5/6 sequence using the 3′ rapid amplification of cDNA ends-polymerase chain reaction method, and confirmed their expression in the leukemic cells. Our case suggests that der(9t(7;9(q11.2;p13 can cause the truncation of the PAX5 transcript, which is supposed to contribute to the generation of MPAL, in addition to three previously reported types of PAX5 fusion.

  1. Generation of an Infectious Clone of a New Korean Isolate of Apple chlorotic leaf spot virus Driven by Dual 35S and T7 Promoters in a Versatile Binary Vector

    Directory of Open Access Journals (Sweden)

    Ik-Hyun Kim

    2017-12-01

    Full Text Available The full-length sequence of a new isolate of Apple chlorotic leaf spot virus (ACLSV from Korea was divergent, but most closely related to the Japanese isolate A4, at 84% nucleotide identity. The full-length cDNA of the Korean isolate of ACLSV was cloned into a binary vector downstream of the bacteriophage T7 RNA promoter and the Cauliflower mosaic virus 35S promoter. Chenopodium quinoa was successfully infected using in vitro transcripts synthesized using the T7 promoter, detected at 20 days post inoculation (dpi, but did not produce obvious symptoms. Nicotiana occidentalis and C. quinoa were inoculated through agroinfiltration. At 32 dpi the infection rate was evaluated; no C. quinoa plants were infected by agroinfiltration, but infection of N. occidentalis was obtained.

  2. Study of rhyolitic glasses alteration in contact with natural brines (Bolivia). Application to the study of the long-term behaviour of the R7T7 nuclear glass

    International Nuclear Information System (INIS)

    Abdelouas, A.

    1996-01-01

    The purpose of this work is to complement an experimental program on the R7T7 nuclear waste glass alteration in brines at 190 deg C in Germany by the analysis of the structure and the chemical composition of the alteration layers, and to study the alteration of rhyolitic glasses in natural brines from Bolivia as analogue for nuclear waste glasses disposed in salt formations. Alteration experiments with the R7T7 and basaltic glasses and obsidian in MgCl 2 -CaCl 2 -saturated brine at 190 deg. C were also conducted in order to study the influence of the glass composition on the nature of the secondary phases. The experiments with the R7T7 glass in three salt brines, saturated respectively in MgCl 2 , MgCl 2 -CaCl 2 and NaCl, showed that the solubilities of most radionuclides are controlled by the secondary phases. Nd, La, and Pr are trapped in powellite, Ce in cerianite, U in coffinite, and Sr is partially immobilized in barite. These phases are stable for more than one year. There is a good similarity between the secondary phases formed experimentally on volcanic glasses and the R7T7 glass altered in MgCl 2 -CaCl 2 -saturated brine. The abundance of Mg in solution permits the formation of similar magnesian clays on the glass samples independently of the nature of the initial glasses. These results support the use of volcanic glasses alteration patterns in Mg-rich solutions to understand the long-term behavior of nuclear waste glasses and to evaluate the stability of the secondary phases. The study of the sediments of Uyuni (Bolivia) showed that the corrosion rate of the rhyolitic glass in brines at 10 deg. C is 12 to 30 time lower than those of rhyolitic glasses altered in high dilute conditions. The low alteration rate of rhyolitic glasses in brines and the formation of secondary phases such as smectite, barite and cerianite (also formed during the experimental alteration of the R7T7 glass), permit us to expect the low alteration of nuclear waste glasses at long

  3. Purification and properties of poliovirus RNA polymerase expressed in Escherichia coli

    International Nuclear Information System (INIS)

    Plotch, S.J.; Palant, O.; Gluzman, Y.

    1989-01-01

    A cDNA clone encoding the RNA polymerase of poliovirus has been expressed in Escherichia coli under the transcriptional control of a T7 bacteriophage promoter. This poliovirus enzyme was designed to contain only a single additional amino acid, the N-terminal methionine. The recombinant enzyme has been purified to near homogeneity, and polyclonal antibodies have been prepared against it. The enzyme exhibits poly(A)-dependent oligo(U)-primed ply(U) polymerase activity as well as RNA polymerase activity. In the presence of an oligo(U) primer, the enzyme catalyzes the synthesis of a full-length copy of either poliovirus or globin RNA templates. In the absence of added primer, RNA products up to twice the length of the template are synthesized. When incubated in the presence of a single nucleoside triphosphate, [α- 32 P]UTP, the enzyme catalyzes the incorporation of radioactive label into template RNA. These results are discussed in light of previously proposed models of poliovirus RNA synthesis in vitro

  4. Over-expression in Escherichia coli and characterization of two recombinant isoforms of human FAD synthetase

    International Nuclear Information System (INIS)

    Brizio, Carmen; Galluccio, Michele; Wait, Robin; Torchetti, Enza Maria; Bafunno, Valeria; Accardi, Rosita; Gianazza, Elisabetta; Indiveri, Cesare; Barile, Maria

    2006-01-01

    FAD synthetase (FADS) (EC 2.7.7.2) is a key enzyme in the metabolic pathway that converts riboflavin into the redox cofactor FAD. Two hypothetical human FADSs, which are the products of FLAD1 gene, were over-expressed in Escherichia coli and identified by ESI-MS/MS. Isoform 1 was over-expressed as a T7-tagged protein which had a molecular mass of 63 kDa on SDS-PAGE. Isoform 2 was over-expressed as a 6-His-tagged fusion protein, carrying an extra 84 amino acids at the N-terminal with an apparent molecular mass of 60 kDa on SDS-PAGE. It was purified near to homogeneity from the soluble cell fraction by one-step affinity chromatography. Both isoforms possessed FADS activity and had a strict requirement for MgCl 2 , as demonstrated using both spectrophotometric and chromatographic methods. The purified recombinant isoform 2 showed a specific activity of 6.8 ± 1.3 nmol of FAD synthesized/min/mg protein and exhibited a K M value for FMN of 1.5 ± 0.3 μM. This is First report on characterization of human FADS, and First cloning and over-expression of FADS from an organism higher than yeast

  5. The Protein Interaction Network of Bacteriophage Lambda with Its Host, Escherichia coli

    Science.gov (United States)

    Blasche, Sonja; Wuchty, Stefan; Rajagopala, Seesandra V.

    2013-01-01

    Although most of the 73 open reading frames (ORFs) in bacteriophage λ have been investigated intensively, the function of many genes in host-phage interactions remains poorly understood. Using yeast two-hybrid screens of all lambda ORFs for interactions with its host Escherichia coli, we determined a raw data set of 631 host-phage interactions resulting in a set of 62 high-confidence interactions after multiple rounds of retesting. These links suggest novel regulatory interactions between the E. coli transcriptional network and lambda proteins. Targeted host proteins and genes required for lambda infection are enriched among highly connected proteins, suggesting that bacteriophages resemble interaction patterns of human viruses. Lambda tail proteins interact with both bacterial fimbrial proteins and E. coli proteins homologous to other phage proteins. Lambda appears to dramatically differ from other phages, such as T7, because of its unusually large number of modified and processed proteins, which reduces the number of host-virus interactions detectable by yeast two-hybrid screens. PMID:24049175

  6. Energetics of sodium efflux from Escherichia coli

    International Nuclear Information System (INIS)

    Borbolla, M.G.; Rosen, B.P.

    1984-01-01

    When energy-starved cells of Escherichia coli were passively loaded with 22 Na+, efflux of sodium could be initiated by addition of a source of metabolic energy. Conditions were established where the source of energy was phosphate bond energy, an electrochemical proton gradient, or both. Only an electrochemical proton gradient was required for efflux from intact cells. These results are consistent with secondary exchange of Na+ for H+ catalyzed by a sodium/proton antiporter

  7. Comparison of the oxygen enhancement ratio for γ-ray-induced double-strand breaks in the DNA of bacteriophage T7 as determined by two different methods of analysis

    International Nuclear Information System (INIS)

    Schans, G.P. van der; Drift, A.C.M. van der.

    1975-01-01

    Bacteriophage T7 was irradiated in a protecting medium under nitrogen and oxygen with 60 Co gamma rays. Double-strand breaks were measured by sucrose gradient sedimentation and by boundary sedimentation analysis. Both methods showed that the presence of oxygen during irradiation enhances the production of double-strand breaks. This is in contrast to a recent report which suggests that boundary sedimentation analysis does not show the effect of oxygen. The discrepancy must be ascribed to differences in the interpretation of the sedimentation data

  8. Targeted capture and heterologous expression of the Pseudoalteromonas alterochromide gene cluster in Escherichia coli represents a promising natural product exploratory platform.

    Science.gov (United States)

    Ross, Avena C; Gulland, Lauren E S; Dorrestein, Pieter C; Moore, Bradley S

    2015-04-17

    Marine pseudoalteromonads represent a very promising source of biologically important natural product molecules. To access and exploit the full chemical capacity of these cosmopolitan Gram-(-) bacteria, we sought to apply universal synthetic biology tools to capture, refactor, and express biosynthetic gene clusters for the production of complex organic compounds in reliable host organisms. Here, we report a platform for the capture of proteobacterial gene clusters using a transformation-associated recombination (TAR) strategy coupled with direct pathway manipulation and expression in Escherichia coli. The ~34 kb pathway for production of alterochromide lipopeptides by Pseudoalteromonas piscicida JCM 20779 was captured and heterologously expressed in E. coli utilizing native and E. coli-based T7 promoter sequences. Our approach enabled both facile production of the alterochromides and in vivo interrogation of gene function associated with alterochromide's unusual brominated lipid side chain. This platform represents a simple but effective strategy for the discovery and biosynthetic characterization of natural products from marine proteobacteria.

  9. Translational coupling in Escherichia coli of a heterologous Bacillus subtilis-Escherichia coli gene fusion.

    OpenAIRE

    Zaghloul, T I; Doi, R H

    1986-01-01

    The efficient expression in Escherichia coli of the Tn9-derived chloramphenicol acetyltransferase (EC 2.3.1.28) gene fused distal to the promoter and N terminus of the Bacillus subtilis aprA gene was dependent on the initiation of translation from the ribosome-binding site in the aprA gene.

  10. ELECTROPHORETIC MOBILITIES OF ESCHERICHIA COLI 0157:H7 AND WILD-TYPE ESCHERICHIA COLI STRAINS

    Science.gov (United States)

    The electrophoretic mobility (EPM) of a number of human-virulent and "wild-type" Escherichia coli strains in phosphate buffered water was measured. The impact of pH, ionic strength, cation type (valence) and concentration, and bacterial strain on the EPM was investigated. Resul...

  11. Enteropathogenic Escherichia coli: foe or innocent bystander?

    Science.gov (United States)

    Hu, Jia; Torres, Alfredo G.

    2015-01-01

    Enteropathogenic Escherichia coli (EPEC) remain one the most important pathogens infecting children and they are one of the main causes of persistent diarrhea worldwide. Historically, typical EPEC (tEPEC), defined as those isolates with the attaching and effacement (A/E) genotype (eae+), which possess bfpA+ and lack the stx- genes are found strongly associated with diarrheal cases. However, occurrence of atypical EPEC (aEPEC; eae+ bfpA- stx-) in diarrheal and asymptomatic hosts has made investigators question the role of these pathogens in human disease. Current epidemiological data is helping answering the question whether EPEC is mainly a foe or an innocent bystander during infection. PMID:25726041

  12. Dynamics of chromosome segregation in Escherichia coli

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck

    2007-01-01

    Since the 1960’es the conformation and segregation of the chromosome in Escherichia coli has been a subject of interest for many scientists. However, after 40 years of research, we still know incredibly little about how the chromosome is organized inside the cell, how it manages to duplicate...... this incredibly big molecule and separate the two daughter chromosomes and how it makes sure that the daughter cells receives one copy each. The fully extended chromosome is two orders of magnitude larger than the cell in which it is contained. Hence the chromosome is heavily compacted in the cell...

  13. Hydrogen production by recombinant Escherichia coli strains

    Science.gov (United States)

    Maeda, Toshinari; Sanchez‐Torres, Viviana; Wood, Thomas K.

    2012-01-01

    Summary The production of hydrogen via microbial biotechnology is an active field of research. Given its ease of manipulation, the best‐studied bacterium Escherichia coli has become a workhorse for enhanced hydrogen production through metabolic engineering, heterologous gene expression, adaptive evolution, and protein engineering. Herein, the utility of E. coli strains to produce hydrogen, via native hydrogenases or heterologous ones, is reviewed. In addition, potential strategies for increasing hydrogen production are outlined and whole‐cell systems and cell‐free systems are compared. PMID:21895995

  14. Vaginal Lactobacillus isolates inhibit uropathogenic Escherichia coli.

    OpenAIRE

    Atassi , Fabrice; Brassart , Dominique; Grob , Philipp; Graf , Federico; Servin , Alain ,

    2006-01-01

    The purpose of this study was to investigate the antibacterial activities of Lactobacillus jensenii KS119.1 and KS121.1, and Lactobacillus gasserii KS120.1 and KS124.3 strains isolated from the vaginal microflora of healthy women, against uropathogenic, diffusely adhering Afa/Dr Escherichia coli (Afa/Dr DAEC) strains IH11128 and 7372 involved in recurrent cystitis. We observed that some of the Lactobacillus isolates inhibited the growth and decreased the viability of E. coli IH11128 and 7372....

  15. 99mTechnetium labelled Escherichia coli

    International Nuclear Information System (INIS)

    Diniz, S.O.F.; Cardoso, V.N.; Resende, B.M.; Nunan, E.A.; Simal, C.J.R.

    1999-01-01

    Samples of a culture of unlabeled Escherichia coli were incubated with different concentrations of stannous chloride for various time periods. 99m Tc (26.0 MBq) was added to each preparation and the results showed a labelling yield of 98% for E. coli. Since the bacterial viability of 99m Tc-E. coli and E. coli did not show any statistical differences, these results demonstrate that labelling of E. coli with 99m Tc does not modify the bacterial viability, and the radiolabelled bacteria may be a good model to study bacterial translocation

  16. Escherichia coli photoreactivating enzyme: purification and properties

    International Nuclear Information System (INIS)

    Snapka, R.M.; Sutherland, B.M.

    1980-01-01

    Researchers have purified large quantities of Escherichia coli photoreactivating enzyme to apparent homogeneity and have studied its physical and chemical properties. The enzyme has a molecular weight of 36,800 and a S/sub 20,w/ 0 of 3.72 S. Amino acid analysis revealed an apparent absence of tryptophan, a low content of aromatic residues, and the presence of no unusual amino acids. The N terminus is arginine. The purified enzyme contained up to 13% carbohydrate by weight. The carbohydrate was composed of mannose, galactose, glucose, and N-acetylglucosamine. The enzyme is also associated with RNA containing uracil, adenine, guanine, and cytosine with no unusual bases detected

  17. Multiplex Genome Editing in Escherichia coli

    DEFF Research Database (Denmark)

    Ingemann Jensen, Sheila; Nielsen, Alex Toftgaard

    2018-01-01

    Lambda Red recombineering is an easy and efficient method for generating genetic modifications in Escherichia coli. For gene deletions, lambda Red recombineering is combined with the use of selectable markers, which are removed through the action of, e.g., flippase (Flp) recombinase. This PCR......-based engineering method has also been applied to a number of other bacteria. In this chapter, we describe a recently developed one plasmid-based method as well as the use of a strain with genomically integrated recombineering genes, which significantly speeds up the engineering of strains with multiple genomic...

  18. Infectious endocarditis caused by Escherichia coli

    DEFF Research Database (Denmark)

    Lauridsen, Trine Kiilerich; Arpi, Magnus; Fritz-Hansen, Thomas

    2011-01-01

    Although Escherichia coli is among the most common causes of Gram-negative bacteraemia, infectious endocarditis (IE) due to this pathogen is rare. A 67-y-old male without a previous medical history presented with a new mitral regurgitation murmur and persisting E. coli bacteraemia in spite of broad......-spectrum intravenous antibiotics. Transthoracic and transoesophageal echocardiography revealed a severe mitral endocarditis. E. coli DNA was identified from the mitral valve and the vegetation, and no other pathogen was found. The case was further complicated by spondylodiscitis and bilateral endophthalmitis. Extra...

  19. Resistant plasmid profile analysis of multidrug resistant Escherichia ...

    African Journals Online (AJOL)

    Background: Multi-drug resistant Escherichia coli has become a major threat and cause of many urinary tract infections (UTIs) in Abeokuta, Nigeria. Objectives: This study was carried out to determine the resistant plasmids of multidrug resistant Escherichia coli isolated from (Urinary tract infections)UTIs in Abeokuta.

  20. Antibiotic resistant Salmonella and Escherichia coli isolated from ...

    African Journals Online (AJOL)

    Results: A hundred and four indigenous chicken rectal swabs were analysed, of which 67.3% were contaminated with Escherichia coli and 12.5% with Salmonella typhimurium. Seventy Escherichia coli isolates showed resistance phenotypes to one, two or more antibiotics. The most common antimicrobial resistance pattern ...

  1. Gene encoding virulence markers among Escherichia coli isolates ...

    African Journals Online (AJOL)

    River water sources and diarrhoeic stools of residents in the Venda Region, Limpopo Province of South Africa were analysed for the prevalence of Escherichia coli (E. coli) and the presence of virulence genes among the isolates. A control group of 100 nondiarrhoeic stool samples was included. Escherichia coli was ...

  2. Chromatin architecture and gene expression in Escherichia coli

    DEFF Research Database (Denmark)

    Willenbrock, Hanni; Ussery, David

    2004-01-01

    Two recent genome-scale analyses underscore the importance of DNA topology and chromatin structure in regulating transcription in Escherichia coli.......Two recent genome-scale analyses underscore the importance of DNA topology and chromatin structure in regulating transcription in Escherichia coli....

  3. Escherichia coli clearance after splenic autotransplants

    International Nuclear Information System (INIS)

    Marques, R.G.; Petroianu, A.; Oliveira, M.B.N.; Bernardo-Filho, M.; Portela, M.C.

    2002-01-01

    Background: Splenic autotransplantation seems to be the only alternative for preservation of splenic tissue, after total splenectomy. The present study was carried out to analyze Escherichia coli depuration by mononuclear phagocyte system organs after total splenectomy and splenic autotransplantation. Methods: We utilized an experimental model including young and adult Wistar rats, of both sexes, submitted to total splenectomy and splenic autotransplantation. The evaluation method was intravenous inoculation of a suspension of Escherichia coli labeled with technetium-99m. We analyzed bacteria uptake by mononuclear phagocyte system organs and bacteria remnant in the bloodstream. Results: There was no difference between young and adult animals in bacteria uptake by mononuclear phagocyte system organs. In the comparison of groups, it was found out that the mean percent uptake by spleen and liver of animals in the control group was higher than that observed for animals with splenic implants. However, bacteria uptake in the lung was higher in the splenic implant group than in the control group. Although spleen bacteria uptake in the control group animals has been higher than that of animals in the splenic implant group, the remnant bacteria in the bloodstream was similar. Animals submitted to isolated total splenectomy showed higher bacteria remnant in the bloodstream than animals of the control group or the group submitted to total splenectomy combined with splenic autotransplantation. Conclusion: Our results indicate that autogenous splenic implant is efficacious in bacteria depuration in rats, by means of their macrophages phagocytosis. In addition, it does not modify bacteria removal function of liver and lung

  4. High-level expression of Bacillus naganoensis pullulanase from recombinant Escherichia coli with auto-induction: effect of lac operator.

    Directory of Open Access Journals (Sweden)

    Yao Nie

    Full Text Available Pullulanase plays an important role in specific hydrolysis of branch points in amylopectin and is generally employed as an important enzyme in starch-processing industry. So far, however, the production level of pullulanase is still somewhat low from wide-type strains and even heterologous expression systems. Here the gene encoding Bacillus naganoensis pullulanase was amplified and cloned. For expression of the protein, two recombinant systems, Escherichia coli BL21(DE3/pET-20b(+-pul and E. coli BL21(DE3/pET-22b(+-pul, were constructed, both bearing T7 promoter and signal peptide sequence, but different in the existance of lac operator and lacI gene encoding lac repressor. Recombinant pullulanase was initially expressed with the activity of up to 14 U/mL by E. coli BL21(DE3/pET-20b(+-pul with IPTG induction in LB medium, but its expression level reduced continually with the extension of cryopreservation time and basal expression was observed. However, E. coli BL21(DE3/pET-22b(+-pul , involving lac operator downstream of T7 promoter to regulate foreign gene transcription, exhibited pullulanase activity consistently without detected basal expression. By investigating the effect of lac operator, basal expression of foreign protein was found to cause expression instability and negative effect on production of target protein. Thus double-repression strategy was proposed that lac operators in both chromosome and plasmid were bound with lac repressor to repress T7 RNA polymerase synthesis and target protein expression before induction. Consequently, the total activity of pullulanase was remarkably increased to 580 U/mL with auto-induction by lac operator-involved E. coli BL21(DE3/pET-22b(+-pul. When adding 0.6% glycine in culture, the extracellular production of pullulanase was significantly improved with the extracellular activity of 502 U/mL, which is a relatively higher level achieved to date for extracellular production of pullulanase. The

  5. Escherichia coli in broiler chickens with airsacculitis

    Directory of Open Access Journals (Sweden)

    Leandro S. Machado

    2014-09-01

    Full Text Available ABSTRACT. Machado L.S., do Nascimento E.R., Pereira V.L.A., Abreu D.L.C., Gouvea R. & Santos L.M.M. 2014. [Escherichia coli in broiler chickens with airsacculitis.] Escherichia coli em frangos de corte com aerossaculite. Revista Brasileira de Medicina Veterinária, 36(3:261-265, 2014. Departamento de Medicina Veterinária Preventiva e Saúde Pública, Faculdade de Veterinária, Universidade Federal Fluminense, Rua Dr. Vital Brazil Filho 64, Vital Brazil, Niterói, RJ 24230-340, Brazil. E-mail: leandromachadovet@yahoo.com.br The Brazilian poultry industry grows each year and becomes increasingly representative in the production and export of products. The health care with poultry have accompanied and favored this evolution, however, respiratory agents that affect the weight and carcass quality, continue to cause great damage to the poultry industry. Airsacculitis is considered the main cause of total and partial condemnation of carcasses of broilers, and has been attributed to Mycoplasmosis mostly caused by Mycoplasma gallisepticum (MG and Mycoplasma synoviae (MS and Escherichia coli. The aim of this study was to relate the positivity of MG / MS and E. coli detected by PCR as a risk factor for airsacculitis in condemnation of broilers in Health Inspection Service. We studied 30 broiler poultry slaughtered in a slaughterhouse under Federal Sanitary Inspection, located in the State of Rio de Janeiro. 30 chickens were randomly collected from different lots and tracheas obtained in each PCR. DNA was extracted by phenol-chloroform method and amplified using pairs of “primer”specific for MG, MS and E. coli. Of the 30 chickens analyzed by PCR, 30% (9/30 had lesions in air sacs. None of the birds showed infection with MG and/or MS PCR, however 33.3% (3/9 birds were positive for airsacculitis iss gene from E.coli. E.coli found in broiler chickens that were negative for mycoplasma airsacculitis, implying the presence of such bacteria may be sufficient

  6. Identification and Prevalence of Escherichia coli and Escherichia coli O157: H7 in Foods

    Directory of Open Access Journals (Sweden)

    Ancuta Mihaela Rotar

    2013-11-01

    Full Text Available The objective of this study is to investigate the incidence of Escherichia coli in animal and non-animal foods, and mainly the incidence of the serotype O157: H7 producing verotoxin. The presence of common Escherichia coli and Escherichia coli O157: H7 in various foods (of animal and non animal origin was performed in Transylvania area. We analyzed a total of one hundred forty-one samples of minced meat, one hundred twenty-six samples of meat , twenty six samples of meat products, five samples of alcoholic beverages, three samples of seafood, one hundred samples of cheese from pasteurized milk, seventeen samples of butter, four samples of vegetables and one sample of milk powder, using the standard cultural method and Vidas Eco method for E. coli O157: H7 strains. E. coli was identified in 50 samples of minced meat, 55 samples of meat prepared, 4 samples of meat products, 2 samples of alcoholic beverages, 25 samples of cheese from pasteurized milk, 6 samples of butter and 1 sample of vegetables. In this study were not been identified any foods contaminated with the E. coli O157: H7 serotype. The results of this reasearch have demostrated that E. coli wich represents a hygienic indicator of recent food contamination, can be destroyed with heat treatment and hygienic handling of foods. Our country over the years has been among the few countries where the incidence of the E. coli O157: H7 serotype has been minimal.

  7. Action of sodium deoxycholate on Escherichia coli

    International Nuclear Information System (INIS)

    D'Mello, A.; Yotis, W.W.

    1987-01-01

    Sodium deoxycholate is used in a number of bacteriological media for the isolation and classification of gram-negative bacteria from food and the environment. Initial experiments to study the effect of deoxycholate on the growth parameters of Escherichia coli showed an increase in the lag time constant and generation time and a decrease in the growth rate constant total cell yield of this microorganisms. Cell fractionation studies indicated that sodium deoxycholate at levels used in bacteriological media interferes with the incorporation of [U- 14 C]glucose into the cold-trichloroacetic acid-soluble, ethanol-soluble, and trypsin-soluble cellular fractions of E. coli. Finally, sodium deoxycholate interfered with the flagellation and motility of Proteus mirabilis and E. coli. It would appear then that further improvement of the deoxycholate medium may be in order

  8. Prodigiosin - A Multifaceted Escherichia coli Antimicrobial Agent.

    Directory of Open Access Journals (Sweden)

    Tjaša Danevčič

    Full Text Available Despite a considerable interest in prodigiosin, the mechanism of its antibacterial activity is still poorly understood. In this work, Escherichia coli cells were treated with prodigiosin to determine its antimicrobial effect on bacterial physiology. The effect of prodigiosin was concentration dependent. In prodigiosin treated cells above MIC value no significant DNA damage or cytoplasmic membrane disintegration was observed. The outer membrane, however, becomes leaky. Cells had severely decreased respiration activity. In prodigiosin treated cells protein and RNA synthesis were inhibited, cells were elongated but could not divide. Pre-treatment with prodigiosin improved E. coli survival rate in media containing ampicillin, kanamycin and erythromycin but not phleomycin. The results suggest that prodigiosin acts as a bacteriostatic agent in E. coli cells. If prodigiosin was diluted, cells resumed growth. The results indicate that prodigiosin has distinct mode of antibacterial action in different bacteria.

  9. Deuterium incorporation into Escherichia-coli proteins

    DEFF Research Database (Denmark)

    Lederer, H.; May, R. P.; Kjems, Jørgen

    1986-01-01

    Neutron small-angle scattering studies of single protein subunits in a protein-DNA complex require the adjustment of the neutron scattering-length densities of protein and DNA, which is attainable by specific deuteration of the protein. The neutron scattering densities of unlabelled DNA and DNA......-dependent RNA polymerase of Escherichia coli match when RNA polymerase is isolated from cells grown in a medium containing 46% D2O and unlabelled glucose as carbon source. Their contrasts vanish simultaneously in a dialysis buffer containing 65% D2O. An expression was evaluated which allows the calculation...... of the degree of deuteration and match point of any E. coli protein from the D2O content of the growth medium, taking the 2H incorporation into RNA polymerase amino acids to be representative for all amino acids in E. coli proteins. The small-angle scattering results, on which the calculation of the degree...

  10. FTIR nanobiosensors for Escherichia coli detection

    Directory of Open Access Journals (Sweden)

    Stefania Mura

    2012-07-01

    Full Text Available Infections due to enterohaemorrhagic E. coli (Escherichia coli have a low incidence but can have severe and sometimes fatal health consequences, and thus represent some of the most serious diseases due to the contamination of water and food. New, fast and simple devices that monitor these pathogens are necessary to improve the safety of our food supply chain. In this work we report on mesoporous titania thin-film substrates as sensors to detect E. coli O157:H7. Titania films treated with APTES ((3-aminopropyltriethoxysilane and GA (glutaraldehyde were functionalized with specific antibodies and the absorption properties monitored. The film-based biosensors showed a detection limit for E. coli of 1 × 102 CFU/mL, constituting a simple and selective method for the effective screening of water samples.

  11. Initiation of Replication in Escherichia coli

    DEFF Research Database (Denmark)

    Frimodt-Møller, Jakob

    The circular chromosome of Escherichia coli is replicated by two replisomes assembled at the unique origin and moving in the opposite direction until they meet in the less well defined terminus. The key protein in initiation of replication, DnaA, facilitates the unwinding of double-stranded DNA...... to single-stranded DNA in oriC. Although DnaA is able to bind both ADP and ATP, DnaA is only active in initiation when bound to ATP. Although initiation of replication, and the regulation of this, is thoroughly investigated it is still not fully understood. The overall aim of the thesis was to investigate...... the regulation of initiation, the effect on the cell when regulation fails, and if regulation was interlinked to chromosomal organization. This thesis uncovers that there exists a subtle balance between chromosome replication and reactive oxygen species (ROS) inflicted DNA damage. Thus, failure in regulation...

  12. Escherichia coli pyomyositis in an immunocompromised host.

    Science.gov (United States)

    Sharma, Umesh; Schwan, William R; Agger, William A

    2011-08-01

    Pyomyositis due to Escherichia coli (E. coil) is rarely reported in immunocompromised patients with hematological malignancy. We present a case report of a 34-year-old man who developed E. coli pyomyositis as a complication of acute myelogenous leukemia (AML). Magnetic resonance imaging (MRI) of the right hip suggested myofascial infection of the gluteal muscles, and a needle muscle aspiration grew E. coli phylogenetic group B2. The patient responded to intravenous piperacillin/tazobactam followed by prolonged oral levofloxacin. Pyomyositis should be suspected in all immunocompromised patients complaining of muscle pain and may exhibit signs of localized muscle infection. Appropriate antibiotic therapy targeting fluoroquinolone-resistant E. coli should be considered for initial empiric therapy of pyomyositis in immunocompromised patients.

  13. Control of Ribosome Synthesis in Escherichia coli

    DEFF Research Database (Denmark)

    Molin, Søren; Meyenburg, K. von; Måløe, O.

    1977-01-01

    The rate of ribosome synthesis and accumulation in Escherichia coli during the transition after an energy source shift-down was analyzed. The shift was imposed on cultures of stringent and relaxed strains growing in glucose minimal medium by the addition of the glucose analogue {alpha...... and to estimate the transcription time for the rRNA operon under different conditions. In steady states of growth with growth rates ranging from 0.75 to 2.3 doublings/h, as well as during the transition after a shift-down, the transcription time of the rRNA operon was constant. The rate of synthesis of r......RNA correlated during this transition – in contrast to the rate of accumulation (M. T. Hansen et al., J. Bacteriol. 122: 585-591, 1975) – with the ppGpp pool in the same way as has been observed during partial amino acid starvation....

  14. Repair replication in permeabilized Escherichia coli

    International Nuclear Information System (INIS)

    Masker, W.E.; Simon, T.J.; Hanawalt, P.C.

    1975-01-01

    We have examined the modes of DNA synthesis in Escherichia coli strains made permeable to nucleoside triphosphates by treatment with toluene. In this quasi in vitro system, polymerase-I-deficient mutants exhibit a nonconservative mode of synthesis with properties expected for the resynthesis step of excision-repair. This uv-stimulated DNA synthesis can be performed by either DNA polymerase II or III and it also requires the uvrA gene product. It requires the four deoxynucleoside triphosphates; but, in contrast to the semiconservative mode, the ATP requirement can be partially satisfied by other nucleoside triphosphates. The ATP-dependent recBC nuclease is not involved. The observed uv-stimulated mode of DNA synthesis may be part of an alternate excision-repair mechanism which supplements or complements DNA-polymerase-I-dependent repair in vivo

  15. Expression of maize prolamins in Escherichia Coli

    International Nuclear Information System (INIS)

    Wang, Szu-zhen; Esen, Asim

    1985-01-01

    We have constructed a cDNA expression library of developing corn (Zea manys L.) endosperm using plasmid pUC8 as vector and Escherichia coli strain DH1 as host. The expression library was screened with non-radioactive immunological probes to detect the expression of gamma-zein and alpha-zein. When anti-gamma-zein antibody was used as the probe, 23 colonies gave positive reactions. The lengths of cDNA inserts of the 23 colonies were found to be 250-900 base pairs. When anti-alpha zein antibody was used, however, fewer colonies gave positive reactions. The library was also screened by colony-hybridization with 32 P-labeled DNA probes. Based on immunological and hybridization screening of the library and other evidence, we conclude that alpha-zein was either toxic to E. coli cells or rapidly degraded whereas gamma-zein and its fragments were readily expressed. (author)

  16. Progressive segregation of the Escherichia coli chromosome

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck; Youngren, Brenda; Hansen, Flemming G.

    2006-01-01

    We have followed the fate of 14 different loci around the Escherichia coli chromosome in living cells at slow growth rate using a highly efficient labelling system and automated measurements. Loci are segregated as they are replicated, but with a marked delay. Most markers segregate in a smooth...... temporal progression from origin to terminus. Thus, the overall pattern is one of continuous segregation during replication and is not consistent with recently published models invoking extensive sister chromosome cohesion followed by simultaneous segregation of the bulk of the chromosome. The terminus......, and a region immediately clockwise from the origin, are exceptions to the overall pattern and are subjected to a more extensive delay prior to segregation. The origin region and nearby loci are replicated and segregated from the cell centre, later markers from the various positions where they lie...

  17. Infectious endocarditis caused by Escherichia coli

    DEFF Research Database (Denmark)

    Lauridsen, Trine Kiilerich; Arpi, Magnus; Fritz-Hansen, Thomas

    2011-01-01

    Although Escherichia coli is among the most common causes of Gram-negative bacteraemia, infectious endocarditis (IE) due to this pathogen is rare. A 67-y-old male without a previous medical history presented with a new mitral regurgitation murmur and persisting E. coli bacteraemia in spite of broad......-spectrum intravenous antibiotics. Transthoracic and transoesophageal echocardiography revealed a severe mitral endocarditis. E. coli DNA was identified from the mitral valve and the vegetation, and no other pathogen was found. The case was further complicated by spondylodiscitis and bilateral endophthalmitis. Extra......-intestinal pathogenic E. coli (ExPEC) are able to colonize tissue outside the gastrointestinal tract and contain a variety of virulence factors that may enable the pathogens to invade and induce infections in the cardiac endothelia. In these cases echocardiography as the imaging technology is of paramount importance...

  18. Genes under positive selection in Escherichia coli

    DEFF Research Database (Denmark)

    Petersen, Lise; Bollback, Jonathan P; Dimmic, Matt

    2007-01-01

    We used a comparative genomics approach to identify genes that are under positive selection in six strains of Escherichia coli and Shigella flexneri, including five strains that are human pathogens. We find that positive selection targets a wide range of different functions in the E. coli genome......, including cell surface proteins such as beta barrel porins, presumably because of the involvement of these genes in evolutionary arms races with other bacteria, phages, and/or the host immune system. Structural mapping of positively selected sites on trans-membrane beta barrel porins reveals...... that the residues under positive selection occur almost exclusively in the extracellular region of the proteins that are enriched with sites known to be targets of phages, colicins, or the host immune system. More surprisingly, we also find a number of other categories of genes that show very strong evidence...

  19. Synergistic effects in mixed Escherichia coli biofilms

    DEFF Research Database (Denmark)

    Reisner, A.; Holler, B.M.; Molin, Søren

    2006-01-01

    Bacterial biofilms, often composed of multiple species and genetically distinct strains, develop under complex influences of cell-cell interactions. Although detailed knowledge about the mechanisms underlying formation of single-species laboratory biofilms has emerged, little is known about...... the pathways governing development of more complex heterogeneous communities. In this study, we established a laboratory model where biofilm-stimulating effects due to interactions between genetically diverse strains of Escherichia coli were monitored. Synergistic induction of biofilm formation resulting from...... the cocultivation of 403 undomesticated E. coli strains with a characterized E. coli K-12 strain was detected at a significant frequency. The survey suggests that different mechanisms underlie the observed stimulation, yet synergistic development of biofilm within the subset of E. coli isolates (n = 56) exhibiting...

  20. Action of sodium deoxycholate on Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    D' Mello, A.; Yotis, W.W.

    1987-08-01

    Sodium deoxycholate is used in a number of bacteriological media for the isolation and classification of gram-negative bacteria from food and the environment. Initial experiments to study the effect of deoxycholate on the growth parameters of Escherichia coli showed an increase in the lag time constant and generation time and a decrease in the growth rate constant total cell yield of this microorganisms. Cell fractionation studies indicated that sodium deoxycholate at levels used in bacteriological media interferes with the incorporation of (U-/sup 14/C)glucose into the cold-trichloroacetic acid-soluble, ethanol-soluble, and trypsin-soluble cellular fractions of E. coli. Finally, sodium deoxycholate interfered with the flagellation and motility of Proteus mirabilis and E. coli. It would appear then that further improvement of the deoxycholate medium may be in order.

  1. In vivo Assembly in Escherichia coli of Transformation Vectors for Plastid Genome Engineering

    Directory of Open Access Journals (Sweden)

    Yuyong Wu

    2017-08-01

    Full Text Available Plastid transformation for the expression of recombinant proteins and entire metabolic pathways has become a promising tool for plant biotechnology. However, large-scale application of this technology has been hindered by some technical bottlenecks, including lack of routine transformation protocols for agronomically important crop plants like rice or maize. Currently, there are no standard or commercial plastid transformation vectors available for the scientific community. Construction of a plastid transformation vector usually requires tedious and time-consuming cloning steps. In this study, we describe the adoption of an in vivo Escherichia coli cloning (iVEC technology to quickly assemble a plastid transformation vector. The method enables simple and seamless build-up of a complete plastid transformation vector from five DNA fragments in a single step. The vector assembled for demonstration purposes contains an enhanced green fluorescent protein (GFP expression cassette, in which the gfp transgene is driven by the tobacco plastid ribosomal RNA operon promoter fused to the 5′ untranslated region (UTR from gene10 of bacteriophage T7 and the transcript-stabilizing 3′UTR from the E. coli ribosomal RNA operon rrnB. Successful transformation of the tobacco plastid genome was verified by Southern blot analysis and seed assays. High-level expression of the GFP reporter in the transplastomic plants was visualized by confocal microscopy and Coomassie staining, and GFP accumulation was ~9% of the total soluble protein. The iVEC method represents a simple and efficient approach for construction of plastid transformation vector, and offers great potential for the assembly of increasingly complex vectors for synthetic biology applications in plastids.

  2. Development of expression vectors for Escherichia coli based on the pCR2 replicon

    Directory of Open Access Journals (Sweden)

    Deb J K

    2007-05-01

    Full Text Available Abstract Background Recent developments in metabolic engineering and the need for expanded compatibility required for co-expression studies, underscore the importance of developing new plasmid vectors with properties such as stability and compatibility. Results We utilized the pCR2 replicon of Corynebacterium renale, which harbours multiple plasmids, for constructing a range of expression vectors. Different antibiotic-resistance markers were introduced and the vectors were found to be 100% stable over a large number of generations in the absence of selection pressure. Compatibility of this plasmid was studied with different Escherichia coli plasmid replicons viz. pMB1 and p15A. It was observed that pCR2 was able to coexist with these E.coli plasmids for 60 generations in the absence of selection pressure. Soluble intracellular production was checked by expressing GFP under the lac promoter in an expression plasmid pCR2GFP. Also high level production of human IFNγ was obtained by cloning the h-IFNγ under a T7 promoter in the expression plasmid pCR2-IFNγ and using a dual plasmid heat shock system for expression. Repeated sub-culturing in the absence of selection pressure for six days did not lead to any fall in the production levels post induction, for both GFP and h-IFNγ, demonstrating that pCR2 is a useful plasmid in terms of stability and compatibility. Conclusion We have constructed a series of expression vectors based on the pCR2 replicon and demonstrated its high stability and sustained expression capacity, in the absence of selection pressure which will make it an efficient tool for metabolic engineering and co-expression studies, as well as for scale up of expression.

  3. Epidemiology and clinical manifestations of enteroaggregative Escherichia coli

    DEFF Research Database (Denmark)

    Hebbelstrup Jensen, Betina; Olsen, Katharina E P; Struve, Carsten

    2014-01-01

    Enteroaggregative Escherichia coli (EAEC) represents a heterogeneous group of E. coli strains. The pathogenicity and clinical relevance of these bacteria are still controversial. In this review, we describe the clinical significance of EAEC regarding patterns of infection in humans, transmission...

  4. TRIMETHOPRIM-SULFAMETHOXAZOLE RESISTANCE IN SEWAGE ISOLATES OF ESCHERICHIA COLI

    Science.gov (United States)

    Sewage samples from seven locations in the United States were analyzed for Escherichia coli isolates which were resistant to trimethoprim-sulfamethoxazole (SXT). The prevalence rate of SXT resistant organisms varied between the different geographical locales. The majority of th...

  5. lactamases genes among0 Escherichia coli from patients with ...

    African Journals Online (AJOL)

    -lactamases (ESBLs) that mediate resistance to b-lactam drugs among Escherichia coli and other uropathogens have been reported worldwide. However, there is little information on the detection of ESBLs genes in E. coli from patients with ...

  6. The Prevalence of Enterhaemorrhagic Escherichia Coli in children ...

    African Journals Online (AJOL)

    EHEC), the pathogenicity of other strains of Escherichia coli and other organisms in children presenting with and without diarrhoea in the hospital. Subjects and Methods: A total of 247 stool samples collected from children aged 1 month to 7 ...

  7. Plasmid-Mediated Quinolone Resistance Genes in Escherichia coli ...

    African Journals Online (AJOL)

    Erah

    PMQR) genes and the prevalence of extended spectrum β-lactamase (ESBL) types in Escherichia coli clinical isolates. Methods: Sixty-one ESBL-producing urinary E. coli isolates were studied. An antibiotic susceptibility test was performed ...

  8. GLYCOSYLATED YGHJ POLYPEPTIDES FROM ENTEROTOXIGENIC ESCHERICHIA COLI (ETEC)

    DEFF Research Database (Denmark)

    2017-01-01

    The present invention relates to glycosylated YghJ polypeptides from or derived from enterotoxigenic Escherichia coli (ETEC) that are immunogenic. In particular, the present invention relates to compositions or vaccines comprising the polypeptides and their application in immunization, vaccination...

  9. Efficient production of mutant phytase (phyA-7) derived from Selenomonas ruminantium using recombinant Escherichia coli in pilot scale.

    Science.gov (United States)

    Chi-Wei Lan, John; Chang, Chih-Kai; Wu, Ho-Shing

    2014-09-01

    A mutant gene of rumen phytase (phyA-7) was cloned into pET23b(+) vector and expressed in the Escherichia coli BL21 under the control of the T7 promoter. The study of fermentation conditions includes the temperature impacts of mutant phytase expression, the effect of carbon supplements over induction stage, the inferences of acetic acid accumulation upon enzyme expression and the comparison of one-stage and two-stage operations in batch mode. The maximum value of phytase activity was reached 107.0 U mL(-1) at induction temperature of 30°C. Yeast extract supplement demonstrated a significant increase on both protein concentration and phytase activity. The acetic acid (2 g L(-1)) presented in the modified synthetic medium demonstrated a significant decrease on expressed phytase activity. A two-stage batch operation enhanced the level of phytase activity from 306 to 1204 U mL(-1) in the 20 L of fermentation scale. An overall 3.7-fold improvement in phytase yield (35,375.72-1,31,617.50 U g(-1) DCW) was achieved in the two-stage operation. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  10. Differential causes of mutation and killing in Escherichia coli after psoralen plus light treatment: monoadducts and cross-links

    Energy Technology Data Exchange (ETDEWEB)

    Seki, T; Nozu, K [Nara Medical Univ., Kashihara (Japan); Kondo, S

    1978-01-01

    On treatment with 8-methoxypsoralen plus near uv light, an excision (uvrB/sup -/) strain of Escherichia coli showed about 3- and 10 times higher sensitivities to killing and mutation, respectively, than its parental strain. On re-irradiation with near uv in the absence of unbound psoralen, the uvrB/sup -/ strain pretreated with psoralen plus near uv showed a decrease in both survival and mutation. After treatment with psoralen plus near uv, re-irradiation of T7DNA in the absence of unbound psoralen caused an increase in the cross-linked fraction with an equivalent decrease in the non-cross-linked fraction. From these and previous results, it is concluded that monoadducts produced by treatment with psoralen plus near uv are converted to cross-links by further irradiation and that, in E.coli, monoadducts are responsible for the mutation induced by psoralen-plus-light whereas cross-links are the major cause of its lethal action.

  11. Identifying New Small Proteins in Escherichia coli.

    Science.gov (United States)

    VanOrsdel, Caitlin E; Kelly, John P; Burke, Brittany N; Lein, Christina D; Oufiero, Christopher E; Sanchez, Joseph F; Wimmers, Larry E; Hearn, David J; Abuikhdair, Fatimeh J; Barnhart, Kathryn R; Duley, Michelle L; Ernst, Sarah E G; Kenerson, Briana A; Serafin, Aubrey J; Hemm, Matthew R

    2018-04-12

    The number of small proteins (SPs) encoded in the Escherichia coli genome is unknown, as current bioinformatics and biochemical techniques make short gene and small protein identification challenging. One method of small protein identification involves adding an epitope tag to the 3' end of a short open reading frame (sORF) on the chromosome, with synthesis confirmed by immunoblot assays. In this study, this strategy was used to identify new E. coli small proteins, tagging 80 sORFs in the E. coli genome, and assayed for protein synthesis. The selected sORFs represent diverse sequence characteristics, including degrees of sORF conservation, predicted transmembrane domains, sORF direction with respect to flanking genes, ribosome binding site (RBS) prediction, and ribosome profiling results. Of 80 sORFs, 36 resulted in encoded synthesized proteins-a 45% success rate. Modeling of detected versus non-detected small proteins analysis showed predictions based on RBS prediction, transcription data, and ribosome profiling had statistically-significant correlation with protein synthesis; however, there was no correlation between current sORF annotation and protein synthesis. These results suggest substantial numbers of small proteins remain undiscovered in E. coli, and existing bioinformatics techniques must continue to improve to facilitate identification. © 2018 The Authors. Proteomics Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim, Towson University.

  12. Engineering Escherichia coli for methanol conversion.

    Science.gov (United States)

    Müller, Jonas E N; Meyer, Fabian; Litsanov, Boris; Kiefer, Patrick; Potthoff, Eva; Heux, Stéphanie; Quax, Wim J; Wendisch, Volker F; Brautaset, Trygve; Portais, Jean-Charles; Vorholt, Julia A

    2015-03-01

    Methylotrophic bacteria utilize methanol and other reduced one-carbon compounds as their sole source of carbon and energy. For this purpose, these bacteria evolved a number of specialized enzymes and pathways. Here, we used a synthetic biology approach to select and introduce a set of "methylotrophy genes" into Escherichia coli based on in silico considerations and flux balance analysis to enable methanol dissimilation and assimilation. We determined that the most promising approach allowing the utilization of methanol was the implementation of NAD-dependent methanol dehydrogenase and the establishment of the ribulose monophosphate cycle by expressing the genes for hexulose-6-phosphate synthase (Hps) and 6-phospho-3-hexuloisomerase (Phi). To test for the best-performing enzymes in the heterologous host, a number of enzyme candidates from different donor organisms were selected and systematically analyzed for their in vitro and in vivo activities in E. coli. Among these, Mdh2, Hps and Phi originating from Bacillus methanolicus were found to be the most effective. Labeling experiments using (13)C methanol with E. coli producing these enzymes showed up to 40% incorporation of methanol into central metabolites. The presence of the endogenous glutathione-dependent formaldehyde oxidation pathway of E. coli did not adversely affect the methanol conversion rate. Taken together, the results of this study represent a major advancement towards establishing synthetic methylotrophs by gene transfer. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  13. Multiple loci affecting photoreactivation in Escherichia coli

    International Nuclear Information System (INIS)

    Sutherland, B.M.; Hausrath, S.G.

    1979-01-01

    Sutherland et al. mapped a phr gene in Escherichia coli at 17 min and found that induction of an E. coli stain lysogenic for a lambda phage carrying this gene increased photoreactivating enzyme levels 2,000-fold. Recently, Smith and Youngs and Sancar and Rupert located a phr gene at 15.9 min. We have therefore investigated the properties of photoreactivating enzyme and cellular photoreactivation in cells containing deletions of the gene at 17 min. Cells with this deletion photoreactivated ultraviolet-induced killing at a rate 20% of normal; they also contained approximately 20% of the normal photoreactivating enzyme level. The residual enzyme in these cells was characterized to determine whether the reduced cellular photoreactivation rate and photoreactivating enzyme levels resulted from reduced numbers of normal enzymes or from an altered enzyme. Photoreactivating enzymes from strains carrying a deletion of the region at 17 min has an apparent K/sub m/ about two- to threefold higher than normal enzyme and showed markedly increased heat lability. The gene at 17 min thus contains information determining the function of the E. coli photoreactivating enzyme rather than the quantity of the enzyme. It is proposed that the gene at 17 min be termed phrA and that located at 15.9 min be termed phrB

  14. Transport proteins promoting Escherichia coli pathogenesis

    Science.gov (United States)

    Tang, Fengyi; Saier, Milton H.

    2014-01-01

    Escherichia coli is a genetically diverse species infecting hundreds of millions of people worldwide annually. We examined seven well-characterized E. coli pathogens causing urinary tract infections, gastroenteritis, pyelonephritis and haemorrhagic colitis. Their transport proteins were identified and compared with each other and a non-pathogenic E. coli K12 strain to identify transport proteins related to pathogenesis. Each pathogen possesses a unique set of protein secretion systems for export to the cell surface or for injecting effector proteins into host cells. Pathogens have increased numbers of iron siderophore receptors and ABC iron uptake transporters, but the numbers and types of low-affinity secondary iron carriers were uniform in all strains. The presence of outer membrane iron complex receptors and high-affinity ABC iron uptake systems correlated, suggesting co-evolution. Each pathovar encodes a different set of pore-forming toxins and virulence-related outer membrane proteins lacking in K12. Intracellular pathogens proved to have a characteristically distinctive set of nutrient uptake porters, different from those of extracellular pathogens. The results presented in this report provide information about transport systems relevant to various types of E. coli pathogenesis that can be exploited in future basic and applied studies. PMID:24747185

  15. Transport proteins promoting Escherichia coli pathogenesis.

    Science.gov (United States)

    Tang, Fengyi; Saier, Milton H

    2014-01-01

    Escherichia coli is a genetically diverse species infecting hundreds of millions of people worldwide annually. We examined seven well-characterized E. coli pathogens causing urinary tract infections, gastroenteritis, pyelonephritis and haemorrhagic colitis. Their transport proteins were identified and compared with each other and a non-pathogenic E. coli K12 strain to identify transport proteins related to pathogenesis. Each pathogen possesses a unique set of protein secretion systems for export to the cell surface or for injecting effector proteins into host cells. Pathogens have increased numbers of iron siderophore receptors and ABC iron uptake transporters, but the numbers and types of low-affinity secondary iron carriers were uniform in all strains. The presence of outer membrane iron complex receptors and high-affinity ABC iron uptake systems correlated, suggesting co-evolution. Each pathovar encodes a different set of pore-forming toxins and virulence-related outer membrane proteins lacking in K12. Intracellular pathogens proved to have a characteristically distinctive set of nutrient uptake porters, different from those of extracellular pathogens. The results presented in this report provide information about transport systems relevant to various types of E. coli pathogenesis that can be exploited in future basic and applied studies. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Tiamulin resistance mutations in Escherichia coli.

    Science.gov (United States)

    Böck, A; Turnowsky, F; Högenauer, G

    1982-01-01

    Forty "two-step" and 13 "three-step" tiamulin-resistant mutants of Escherichia coli PR11 were isolated and tested for alteration of ribosomal proteins. Mutants with altered ribosomal proteins S10, S19, L3, and L4 were detected. The S19, L3, and L4 mutants were studied in detail. The L3 and L4 mutations did not segregate from the resistance character in transductional crosses and therefore seem to be responsible for the resistance. Extracts of these mutants also exhibited an increased in vitro resistance to tiamulin in the polyuridylic acid and phage R17 RNA-dependent polypeptide synthesis systems, and it was demonstrated that this was a property of the 50S subunit. In the case of the S19 mutant, genetic analysis showed segregation between resistance and the S19 alteration and therefore indicated that mutation of a protein other than S19 was responsible for the resistance phenotype. The isolated ribosomes of the S19, L3, and L4 mutants bound radioactive tiamulin with a considerably reduced strength when compared with those of wild-type cells. The association constants were lower by factors ranging from approximately 20 to 200. When heated in the presence of ammonium chloride, these ribosomes partially regained their avidity for tiamulin. Images PMID:7050084

  17. Photoinactivation of mcr-1 positive Escherichia coli

    Science.gov (United States)

    Caires, C. S. A.; Leal, C. R. B.; Rodrigues, A. C. S.; Lima, A. R.; Silva, C. M.; Ramos, C. A. N.; Chang, M. R.; Arruda, E. J.; Oliveira, S. L.; Nascimento, V. A.; Caires, A. R. L.

    2018-01-01

    The emergence of plasmid-mediated colistin resistance in Enterobacteriaceae, mostly in Escherichia coli due to the mcr-1 gene, has revealed the need to develop alternative approaches in treating mcr-1 positive bacterial infections. This is because colistin is a broad-spectrum antibiotic and one of the ‘last-resort’ antibiotics for multidrug resistant bacteria. The present study evaluated for the first time, to the best of our knowledge, the efficacy of photoinactivation processes to kill a known mcr-1 positive E. coli strain. Eosin methylene-blue (EMB) was investigated as a photoantimicrobial agent for inhibiting the growth of a mcr-1 positive E. coli strain obtained from a patient with a diabetic foot infection. The photoantimicrobial activity of EMB was also tested in a non-multidrug resistant E. coli strain. The photoinactivation process was tested using light doses in the 30-45 J cm-2 range provided by a LED device emitting at 625 nm. Our findings demonstrate that a mcr-1 positive E. coli strain is susceptible to photoinactivation. The results show that the EMB was successfully photoactivated, regardless of the bacterial multidrug resistance; inactivating the bacterial growth by oxidizing the cells in accordance with the generation of the oxygen reactive species. Our results suggest that bacterial photoinactivation is an alternative and effective approach to kill mcr-1 positive bacteria.

  18. Profiling of Escherichia coli Chromosome database.

    Science.gov (United States)

    Yamazaki, Yukiko; Niki, Hironori; Kato, Jun-ichi

    2008-01-01

    The Profiling of Escherichia coli Chromosome (PEC) database (http://www.shigen.nig.ac.jp/ecoli/pec/) is designed to allow E. coli researchers to efficiently access information from functional genomics studies. The database contains two principal types of data: gene essentiality and a large collection of E. coli genetic research resources. The essentiality data are based on data compilation from published single-gene essentiality studies and on cell growth studies of large-deletion mutants. Using the circular and linear viewers for both whole genomes and the minimal genome, users can not only gain an overview of the genome structure but also retrieve information on contigs, gene products, mutants, deletions, and so forth. In particular, genome-wide exhaustive mutants are an essential resource for studying E. coli gene functions. Although the genomic database was constructed independently from the genetic resources database, users may seamlessly access both types of data. In addition to these data, the PEC database also provides a summary of homologous genes of other bacterial genomes and of protein structure information, with a comprehensive interface. The PEC is thus a convenient and useful platform for contemporary E. coli researchers.

  19. Revisiting the Acanthamoeba species that form star-shaped cysts (genotypes T7, T8, T9, and T17): characterization of seven new Brazilian environmental isolates and phylogenetic inferences.

    Science.gov (United States)

    Magliano, Ana C M; Teixeira, Marta M G; Alfieri, Silvia C

    2012-01-01

    Free-living amoebae of the genus Acanthamoeba are the agents of both opportunistic and non-opportunistic infections and are frequently isolated from the environment. Of the 17 genotypes (T1-T17) identified thus far, 4 (T7, T8, T9, and T17) accommodate the rarely investigated species of morphological group I, those that form large, star-shaped cysts. We report the isolation and characterization of 7 new Brazilian environmental Acanthamoeba isolates, all assigned to group I. Phylogenetic analyses based on partial (~1200 bp) SSU rRNA gene sequences placed the new isolates in the robustly supported clade composed of the species of morphological group I. One of the Brazilian isolates is closely related to A. comandoni (genotype T9), while the other 6, together with 2 isolates recently assigned to genotype T17, form a homogeneous, well-supported group (2·0% sequence divergence) that likely represents a new Acanthamoeba species. Thermotolerance, osmotolerance, and cytophatic effects, features often associated with pathogenic potential, were also examined. The results indicated that all 7 Brazilian isolates grow at temperatures up to 40°C, and resist under hyperosmotic conditions. Additionally, media conditioned by each of the new Acanthamoeba isolates induced the disruption of SIRC and HeLa cell monolayers.

  20. Role of wild birds as carriers of multi-drug resistant Escherichia coli and Escherichia vulneris

    Directory of Open Access Journals (Sweden)

    Mohammed Y. Shobrak

    2014-12-01

    Full Text Available Emergence and distribution of multi-drug resistant (MDR bacteria in environments pose a risk to human and animal health. A total of 82 isolates of Escherichia spp. were recovered from cloacal swabs of migrating and non-migrating wild birds. All bacterial isolates were identified and characterized morphologically and biochemically. 72% and 50% of isolates recovered from non-migrating and migrating birds, respectively, showed positive congo red dye binding (a virulence factor. Also, hemolysin production (a virulence factor was showed in 8% of isolates recovered from non-migrating birds and 75% of isolates recovered from migrating birds. All isolates recovered from non-migrating birds were found resistant to Oxacillin while all isolates recovered from migrating birds demonstrated resistance to Oxacillin, Chloramphenicol, Oxytetracycline and Lincomycin. Some bacterial isolates recovered from non-migrating birds and migrating birds exhibited MDR phenotype. The MDR isolates were further characterized by API 20E and 16S rRNA as E. coli and E. vulneris. MDR Escherichia isolates contain ~1-5 plasmids of high-molecular weights. Accordingly, wild birds could create a potential threat to human and animal health by transmitting MDR bacteria to water streams and other environmental sources through their faecal residues, and to remote regions by migration.

  1. Role of wild birds as carriers of multi-drug resistant Escherichia coli and Escherichia vulneris

    Science.gov (United States)

    Shobrak, Mohammed Y.; Abo-Amer, Aly E.

    2014-01-01

    Emergence and distribution of multi-drug resistant (MDR) bacteria in environments pose a risk to human and animal health. A total of 82 isolates of Escherichia spp. were recovered from cloacal swabs of migrating and non-migrating wild birds. All bacterial isolates were identified and characterized morphologically and biochemically. 72% and 50% of isolates recovered from non-migrating and migrating birds, respectively, showed positive congo red dye binding (a virulence factor). Also, hemolysin production (a virulence factor) was showed in 8% of isolates recovered from non-migrating birds and 75% of isolates recovered from migrating birds. All isolates recovered from non-migrating birds were found resistant to Oxacillin while all isolates recovered from migrating birds demonstrated resistance to Oxacillin, Chloramphenicol, Oxytetracycline and Lincomycin. Some bacterial isolates recovered from non-migrating birds and migrating birds exhibited MDR phenotype. The MDR isolates were further characterized by API 20E and 16S rRNA as E. coli and E. vulneris. MDR Escherichia isolates contain ~1–5 plasmids of high-molecular weights. Accordingly, wild birds could create a potential threat to human and animal health by transmitting MDR bacteria to water streams and other environmental sources through their faecal residues, and to remote regions by migration. PMID:25763023

  2. Nanotextile membranes for bacteria Escherichia coli capturing

    Directory of Open Access Journals (Sweden)

    Jaroslav Lev

    2010-01-01

    Full Text Available The article describes an experimental study dealing with the possibility of nanotextile materials usa­ge for microbiologically contaminated water filtration. The aim of the study is to verify filtration ability of different nanotextile materials and evaluate the possibilities of practical usage. Good detention ability of these materials in the air filtration is the presumption for nanotextile to be used for bacteria filtration from a liquid. High nanotextile porosity with the nanotextile pores dimensions smaller than a bacteria size predicates the possibility of a successful usage of these materials. For the experiment were used materials made from electrospinning nanofibres under the label PA612, PUR1, PUR2 s PUR3 on the supporting unwoven textiles (viscose and PP. As a model simulation of the microbial contamination, bacteria Escherichia coli was chosen. Contaminated water was filtered during the overpressure activity of 105Pa on the input side of the filter from the mentioned material. After three-day incubation on the nutrient medium, cultures found in the samples before and after filtration were compared. In the filtrated water, bacteria E. coli were indicated, which did not verify the theoretical presumptions about an absolut bacteria detention. However, used materials caught at least 94% of bacteria in case of material PUR1 and up to 99,996% in case of material PUR2. These results predict the possibility of producing effective nanotextile filters for microbiologically contaminated water filtration.Recommendation: For the production of materials with better filtrating qualities, experiments need to be done, enabling better understanding of the bacteria detention mechanisms on the nanotextile material, and parameters of the used materials that influence the filtrating abilities need to be verified.

  3. Duplication of C7orf58, WNT16 and FAM3C in an obese female with a t(7;22)(q32.1;q11.2) chromosomal translocation and clinical features resembling Coffin-Siris Syndrome.

    Science.gov (United States)

    Zhu, Jun; Qiu, Jun; Magrane, Gregg; Abedalthagafi, Malak; Zanko, Andrea; Golabi, Mahin; Chehab, Farid F

    2012-01-01

    We characterized the t(7;22)(q32;q11.2) chromosomal translocation in an obese female with coarse features, short stature, developmental delay and a hypoplastic fifth digit. While these clinical features suggest Coffin-Siris Syndrome (CSS), we excluded a CSS diagnosis by exome sequencing based on the absence of deleterious mutations in six chromatin-remodeling genes recently shown to cause CSS. Thus, molecular characterization of her translocation could delineate genes that underlie other syndromes resembling CSS. Comparative genomic hybridization microarrays revealed on chromosome 7 the duplication of a 434,682 bp region that included the tail end of an uncharacterized gene termed C7orf58 (also called CPED1) and spanned the entire WNT16 and FAM3C genes. Because the translocation breakpoint on chromosome 22 did not disrupt any apparent gene, her disorder was deemed to result from the rearrangement on chromosome 7. Mapping of yeast and bacterial artificial chromosome clones by fluorescent in situ hybridization on chromosome spreads from this patient showed that the duplicated region and all three genes within it were located on both derivative chromosomes 7 and 22. Furthermore, DNA sequencing of exons and splice junctional regions from C7orf58, WNT16 and FAM3C revealed the presence of potential splice site and promoter mutations, thereby augmenting the detrimental effect of the duplicated genes. Hence, dysregulation and/or disruptions of C7orf58, WNT16 and FAM3C underlie the phenotype of this patient, serve as candidate genes for other individuals with similar clinical features and could provide insights into the physiological role of the novel gene C7orf58.

  4. Duplication of C7orf58, WNT16 and FAM3C in an obese female with a t(7;22(q32.1;q11.2 chromosomal translocation and clinical features resembling Coffin-Siris Syndrome.

    Directory of Open Access Journals (Sweden)

    Jun Zhu

    Full Text Available We characterized the t(7;22(q32;q11.2 chromosomal translocation in an obese female with coarse features, short stature, developmental delay and a hypoplastic fifth digit. While these clinical features suggest Coffin-Siris Syndrome (CSS, we excluded a CSS diagnosis by exome sequencing based on the absence of deleterious mutations in six chromatin-remodeling genes recently shown to cause CSS. Thus, molecular characterization of her translocation could delineate genes that underlie other syndromes resembling CSS. Comparative genomic hybridization microarrays revealed on chromosome 7 the duplication of a 434,682 bp region that included the tail end of an uncharacterized gene termed C7orf58 (also called CPED1 and spanned the entire WNT16 and FAM3C genes. Because the translocation breakpoint on chromosome 22 did not disrupt any apparent gene, her disorder was deemed to result from the rearrangement on chromosome 7. Mapping of yeast and bacterial artificial chromosome clones by fluorescent in situ hybridization on chromosome spreads from this patient showed that the duplicated region and all three genes within it were located on both derivative chromosomes 7 and 22. Furthermore, DNA sequencing of exons and splice junctional regions from C7orf58, WNT16 and FAM3C revealed the presence of potential splice site and promoter mutations, thereby augmenting the detrimental effect of the duplicated genes. Hence, dysregulation and/or disruptions of C7orf58, WNT16 and FAM3C underlie the phenotype of this patient, serve as candidate genes for other individuals with similar clinical features and could provide insights into the physiological role of the novel gene C7orf58.

  5. Synthesis of avenanthramides using engineered Escherichia coli.

    Science.gov (United States)

    Lee, Su Jin; Sim, Geun Young; Kang, Hyunook; Yeo, Won Seok; Kim, Bong-Gyu; Ahn, Joong-Hoon

    2018-03-22

    Hydroxycinnamoyl anthranilates, also known as avenanthramides (avns), are a group of phenolic alkaloids with anti-inflammatory, antioxidant, anti-itch, anti-irritant, and antiatherogenic activities. Some avenanthramides (avn A-H and avn K) are conjugates of hydroxycinnamic acids (HC), including p-coumaric acid, caffeic acid, and ferulic acid, and anthranilate derivatives, including anthranilate, 4-hydroxyanthranilate, and 5-hydroxyanthranilate. Avns are primarily found in oat grain, in which they were originally designated as phytoalexins. Knowledge of the avns biosynthesis pathway has now made it possible to synthesize avns through a genetic engineering strategy, which would help to further elucidate their properties and exploit their beneficial biological activities. The aim of the present study was to synthesize natural avns in Escherichia coli to serve as a valuable resource. We synthesized nine avns in E. coli. We first synthesized avn D from glucose in E. coli harboring tyrosine ammonia lyase (TAL), 4-coumarate:coenzyme A ligase (4CL), anthranilate N-hydroxycinnamoyl/benzoyltransferase (HCBT), and anthranilate synthase (trpEG). A trpD deletion mutant was used to increase the amount of anthranilate in E. coli. After optimizing the incubation temperature and cell density, approximately 317.2 mg/L of avn D was synthesized. Avn E and avn F were then synthesized from avn D, using either E. coli harboring HpaBC and SOMT9 or E. coli harboring HapBC alone, respectively. Avn A and avn G were synthesized by feeding 5-hydroxyanthranilate or 4-hydroxyanthranilate to E. coli harboring TAL, 4CL, and HCBT. Avn B, avn C, avn H, and avn K were synthesized from avn A or avn G, using the same approach employed for the synthesis of avn E and avn F from avn D. Using different HCs, nine avns were synthesized, three of which (avn D, avn E, and avn F) were synthesized from glucose in E. coli. These diverse avns provide a strategy to synthesize both natural and unnatural avns

  6. Complete Genome Sequence of Escherichia coli Strain WG5

    DEFF Research Database (Denmark)

    Imamovic, Lejla; Misiakou, Maria-Anna; van der Helm, Eric

    2018-01-01

    Escherichia coli strain WG5 is a widely used host for phage detection, including somatic coliphages employed as standard ISO method 10705-1 (2000). Here, we present the complete genome sequence of a commercial E. coli WG5 strain.......Escherichia coli strain WG5 is a widely used host for phage detection, including somatic coliphages employed as standard ISO method 10705-1 (2000). Here, we present the complete genome sequence of a commercial E. coli WG5 strain....

  7. Findings of Escherichia coli and Enterococcus spp. in homemade cheese

    Directory of Open Access Journals (Sweden)

    Tambur Zoran

    2007-01-01

    Full Text Available During the period from February until March 2004, 108 samples of soft cheese originating from markets of Pancevo, Subotica and Belgrade were examined. Microbiological analyses of the cheese samples to the presence of Escherichia coli was performed using methods described in the Regulations on methods for performing microbiological analyses and super analyses of consumer articles, while the presence of bacteria Enteroccocus spp. was performed on the dexter agar. From 108 samples of soft cheese from the territories of Pancevo, Belgrade and Subotica were isolated: Enterococcus spp. from 96% and Escherichia coli from 69%, cheese samples. Verocytotoxic E.coli was not isolated from any of the taken cheese samples.

  8. Transverse water relaxation in whole blood and erythrocytes at 3T, 7T, 9.4T, 11.7T and 16.4T; determination of intracellular hemoglobin and extracellular albumin relaxivities.

    Science.gov (United States)

    Grgac, Ksenija; Li, Wenbo; Huang, Alan; Qin, Qin; van Zijl, Peter C M

    2017-05-01

    Blood is a physiological substance with multiple water compartments, which contain water-binding proteins such as hemoglobin in erythrocytes and albumin in plasma. Knowing the water transverse (R 2 ) relaxation rates from these different blood compartments is a prerequisite for quantifying the blood oxygenation level-dependent (BOLD) effect. Here, we report the Carr-Purcell-Meiboom-Gill (CPMG) based transverse (R 2CPMG ) relaxation rates of water in bovine blood samples circulated in a perfusion system at physiological temperature in order to mimic blood perfusion in humans. R 2CPMG values of blood plasma, lysed packed erythrocytes, lysed plasma/erythrocyte mixtures, and whole blood at 3 T, 7 T, 9.4 T, 11.7 T and 16.4 T were measured as a function of hematocrit or hemoglobin concentration, oxygenation, and CPMG inter-echo spacing (τ cp ). R 2CPMG in lysed cells showed a small τ cp dependence, attributed to the water exchange rate between free and hemoglobin-bound water to be much faster than τ cp . This was contrary to the tangential dependence in whole blood, where a much slower exchange between cells and blood plasma applies. Whole blood data were fitted as a function of τ cp using a general tangential correlation time model applicable for exchange as well as diffusion contributions to R 2CPMG , and the intercept R 20blood at infinitely short τ cp was determined. The R 20blood values at different hematocrit and the R 2CPMG values of lysed erythrocyte/plasma mixtures at different hemoglobin concentration were used to determine the relaxivity of hemoglobin inside the erythrocyte (r 2Hb ) and albumin (r 2Alb ) in plasma. The r 2Hb values obtained from lysed erythrocytes and whole blood were comparable at full oxygenation. However, while r 2Hb determined from lysed cells showed a linear dependence on oxygenation, this dependence became quadratic in whole blood. This possibly suggests an additional relaxation effect inside intact cells, perhaps due to hemoglobin

  9. WGS accurately predicts antimicrobial resistance in Escherichia coli

    Science.gov (United States)

    Objectives: To determine the effectiveness of whole-genome sequencing (WGS) in identifying resistance genotypes of multidrug-resistant Escherichia coli (E. coli) and whether these correlate with observed phenotypes. Methods: Seventy-six E. coli strains were isolated from farm cattle and measured f...

  10. Differential expression of the Escherichia coli autoaggregation factor antigen 43

    DEFF Research Database (Denmark)

    Schembri, Mark; Hjerrild, Louise; Gjermansen, Morten

    2003-01-01

    Antigen 43 (Ag43) is a self-recognizing surface adhesin found in most Escherichia coli strains. Due to its excellent cell-to-cell aggregation characteristics, Ag43 expression confers clumping and fluffing of cells and promotes biofilm formation. Ag43 expression is repressed by the cellular redox...

  11. Increased multi-drug resistant Escherichia coli from hospitals in ...

    African Journals Online (AJOL)

    Background: Multidrug-resistant Escherichia coli (MDR E. coli) has become a major public health concern in Sudan and many countries, causing failure in treatment with consequent huge health burden. Objectives: To determine the prevalence and susceptibility of MDR E. coli isolated from patients in hospitals at Khartoum ...

  12. Escherichia coli bacteraemia in patients with and without haematological malignancies

    DEFF Research Database (Denmark)

    Olesen, B; Kolmos, H J; Orskov, F

    1998-01-01

    We compared serotypes, virulence factors and susceptibility to antibiotics of Escherichia coli strains isolated from 282 patients with bacteraemia. Thirty-five of these were neutropenic patients with haematological malignancy and 247 were patients with a normal or raised total white blood cell co...

  13. Changes in Escherichia coli resistance to co-trimoxazole in ...

    African Journals Online (AJOL)

    In Thyolo district, Malawi, an operational research study is being conducted on the efficacy and feasibility of co-trimoxazole prophylaxis in preventing deaths in HIV-positive patients with tuberculosis (TB). A series of cross-sectional studies were carried out to determine i) whether faecal Escherichia coli (E.coli) resistance to ...

  14. Effects of recombinant human collagen VI from Escherichia coli on ...

    African Journals Online (AJOL)

    Jane

    2011-07-20

    Jul 20, 2011 ... In this study, we reported the cloning and over expression of a gene coding for human collagen peptide. (CP6) in Escherichia coli and investigated the protective effects of CP6 on UVA-irradiated human skin fibroblasts cells. The collagen peptide (CP6) was highly soluble and the expression level was.

  15. Escherichia coli growth modeling using neural network | Shamsudin ...

    African Journals Online (AJOL)

    technique that has the ability to predict with efficient and good performance. Using NARX, a highly accurate model was developed to predict the growth of Escherichia coli (E. coli) based on pH water parameter. The multiparameter portable sensor and spectrophotometer data were used to build and train the neural network.

  16. DNA supercoiling depends on the phosphorylation potential in Escherichia coli

    DEFF Research Database (Denmark)

    Van Workum, M.; van Dooren, S.J.M; Oldenburg, N

    1996-01-01

    ATP/ADP ratios were varied in different ways and the degree of negative supercoiling was determined in Escherichia coli. Independent of whether the ATP/ADP ratio was reduced by a shift to anaerobic conditions, by addition of protonophore (dinitrophenol) or by potassium cyanide addition, DNA super...

  17. Effect of visible range electromagnetic radiations on Escherichia coli ...

    African Journals Online (AJOL)

    Background: Escherichia coli is the agent responsible for a range of clinical diseases. With emerging antimicrobial resistance, other treatment options including solar/photo-therapy are becoming increasingly common. Visible Range Radiation Therapy/Colour Therapy is an emerging technique in the field of ...

  18. Modulation of allele leakiness and adaptive mutability in Escherichia ...

    Indian Academy of Sciences (India)

    It is shown that partial phenotypic suppression of two ochre mutations (argE3 and lacZU118) and an amber mutation (in argE) by sublethal concentrations of streptomycin in an rpsL+ (streptomycin-sensitive) derivative of the Escherichia coli strain AB1157 greatly enhances their adaptive mutability under selection.

  19. Properties of in situ Escherichia coli -D-glucuronidase (GUS ...

    African Journals Online (AJOL)

    A study of the activity of Escherichia coli -D-glucuronidase (GUS) in polluted stagnant and running water samples was performed with an objective of assessing the viability of a direct marker enzyme assay as a suitable alternative to membrane filtration for the indication of faecal pollution in water intended for drinking ...

  20. QSAR study of benzimidazole derivatives inhibition on escherichia ...

    African Journals Online (AJOL)

    The paper describes a quantitative structure-activity relationship (QSAR) study of IC50 values of benzimidazole derivatives on escherichia coli methionine aminopeptidase. The activity of the 32 inhibitors has been estimated by means of multiple linear regression (MLR) and artificial neural network (ANN) techniques.

  1. Multiple-Resistant Commensal Escherichia Coli from Nigerian ...

    African Journals Online (AJOL)

    Purpose: The antimicrobial susceptibility and virulence traits of 150 strains of Escherichia coli ... and ethical approval was obtained from the Health .... persist in the guts by virtue of the ability of such ... cases of diarrhoea in Ile-Ife and environs.

  2. Cytokine response to Escherichia coli in gnotobiotic pigs

    Czech Academy of Sciences Publication Activity Database

    Šplíchal, Igor; Šplíchalová, Alla; Trebichavský, Ilja

    2008-01-01

    Roč. 53, č. 2 (2008), s. 161-164 ISSN 0015-5632 R&D Projects: GA ČR GA523/05/0249 Institutional research plan: CEZ:AV0Z50200510 Keywords : germ-free pigs * escherichia coli * cytokine response Subject RIV: EE - Microbiology, Virology Impact factor: 1.172, year: 2008

  3. Escherichia coli as other Enterobacteriaceae: food poisoning and health effects

    Science.gov (United States)

    Many Escherichia coli strains are harmless, and they are an important commensal in the intestinal microflora; however, pathogenic strains also exist. The pathogenic strains can be divided into diarrhea-inducing strains and strains that reside in the intestines but only cause disease in bodily sites...

  4. Escherichia coli and other Enterobacteriaceae: Food poisoning and health effects

    Science.gov (United States)

    The family Enterobactericeae consists of rod-shaped, Gram-negative, facultatively anaerobic, non-spore forming bacteria and also includes the food-borne pathogens, Cronobacter spp., Escherichia coli, Salmonella enterica, Shigella spp., and Yersinia spp. Illness caused by these pathogens is acquired...

  5. Binding of Divalent Magnesium by Escherichia coli Phosphoribosyl Diphosphate Synthetase

    DEFF Research Database (Denmark)

    Willemoës, Martin; Hove-Jensen, Bjarne

    1997-01-01

    The mechanism of binding of the substrates MgATP and ribose 5-phosphate as well as Mg2+ to the enzyme 5-phospho-d-ribosyl a-1-diphosphate synthetase from Escherichia coli has been analyzed. By use of the competive inhibitors of ATP and ribose 5-phosphate binding, a,ß-methylene ATP and (+)-1-a,2-a...

  6. Production of 3-O-xylosyl quercetin in Escherichia coli

    DEFF Research Database (Denmark)

    Pandey, Ramesh Prasad; Malla, Sailesh; Simkhada, Dinesh

    2013-01-01

    Quercetin, a flavonol aglycone, is one of the most abundant flavonoids with high medicinal value. The bioavailability and pharmacokinetic properties of quercetin are influenced by the type of sugars attached to the molecule. To efficiently diversify the therapeutic uses of quercetin, Escherichia ...

  7. Kwantitatief gevoeligheidsonderzoek met intra- en extramurale isolaten van Escherichia coli

    NARCIS (Netherlands)

    de Neeling AJ; de Jong J; Overbeek BP; de Bruin RW; Dessens-Kroon M; van Klingeren B

    1990-01-01

    Three Dutch laboratories for medical microbiology collected a total number of 1432 strains of Escherichia coli. Of these 995 were obtained from routine samples taken in clinic and policlinic, 290 had been sent spontaneously by general practitioners for microbiological examination and 147 had been

  8. Growth modeling of uropathogenic Escherichia coli in ground chicken meat

    Science.gov (United States)

    Extraintestinal Pathogenic Escherichia coli (ExPEC), including Uropathogenic E. coli (UPEC), are common contaminants in poultry meat, and are a major pathogen associated with inflammatory bowel disease, ulcerative colitis, sepsis, and urinary tract infections. The purpose of this study was to determ...

  9. in Escherichia coli with native cholesterol oxidase expressed

    African Journals Online (AJOL)

    The structure and bio-activity of an endogenous cholesterol oxidase from Brevibacterium sp. was compared to the same enzyme exogenously expressed in Escherichia coli BL21 (DE3) with and without N- or C-terminal his-tags. The different proteins were purified with affinity and subtractive protocols. The specific activity of ...

  10. Sequencing of Escherichia coli that cause persistent and transient Mastitis

    Science.gov (United States)

    The genomes of two strains of Escherichia coli that cause bovine mastitis were sequenced. These strains are known to be associated with persistent and transient mastitis: strain ECA-B causes a transient infection, and ECC-M leads to a persistent infection....

  11. Escherichia coli. A sanitary methodology for faecal water pollution tests

    International Nuclear Information System (INIS)

    Bonadonna, L.

    2001-01-01

    Among the traditional indictors of faecal water pollution, Escherichia coli has shown to fit better with the definition of indicator organism. Till now its recovery has been time-consuming and needs confirmation tests. In this report more rapid and direct methods, based on enzymatic reactions, are presented [it

  12. Antimicrobial susceptibilities of avian Escherichia coli isolates in ...

    African Journals Online (AJOL)

    Colibacillosis is a poultry disease of economic importance in Iran and all around the world. The aim of this study is to test the antibiotic sensitivity of Escherichia coli strains which were isolated in Tabriz. A total of 100 E. coli strains isolated from avian colibacillosis of 50 farms from 2008 to 2009 in Tabriz, were investigated for ...

  13. Antibiotic resistance profile of Escherichia coli isolated from five ...

    African Journals Online (AJOL)

    Information on the resistance profiles of clinical and non clinical human bacteria isolates in the developing countries can serve as important means of understanding the human pathogens drug resistance interactions in the zone. Escherichia coli isolated from five geopolitical zones of Nigeria were screened for anti-microbial ...

  14. Effect of high pressurized carbon dioxide on Escherichia coli ...

    African Journals Online (AJOL)

    Carbon dioxide at high pressure can retard microbial growth and sometimes kill microorganisms depending on values of applied pressure, temperature and exposure time. In this study the effect of high pressurised carbon dioxide (HPCD) on Escherichia coli was investigated. Culture of E. coli was subjected to high ...

  15. Modeling base excision repair in Escherichia coli bacterial cells

    International Nuclear Information System (INIS)

    Belov, O.V.

    2011-01-01

    A model describing the key processes in Escherichia coli bacterial cells during base excision repair is developed. The mechanism is modeled of damaged base elimination involving formamidopyrimidine DNA glycosylase (the Fpg protein), which possesses several types of activities. The modeling of the transitions between DNA states is based on a stochastic approach to the chemical reaction description

  16. Prevalence of Aeromonas species and Escherichia coli in stool ...

    African Journals Online (AJOL)

    Background: Diarrhoea is one of the main causes of mortality and morbidity in childhood. Bacterial diarrhoea is a common disorder. Aeromonas species and Escherichia coli (E. coli) are some of the aetiological agents associated with diarrhoea in children. Objective: To determine the prevalence of Aeromonas species and ...

  17. Isolation and genomic characterization of Escherichia coli O157:NM ...

    African Journals Online (AJOL)

    Human diseases caused by Escherichia coli O157:NM and E. coli O157:H7 strains have been reported throughout the world. In developed countries, serotype O157:H7 represents the major cause of human diseases; however, there have been increasing reports of non-O157 Shiga toxin (Stx)-producing E. coli strains ...

  18. Adsorption of Escherichia coli Using Bone Char | Rezaee | Journal ...

    African Journals Online (AJOL)

    The aim of study was providing a novel adsorbent for the removal of Escherichia coli (E.coli) as a microbial model from contaminated air especially in hospital units using bone char (BC). The BC was prepared from cattle animal bone by pyrolysis in a furnace at 450°C for 2 h. The characteristics of BC have been determined ...

  19. Occurrence of Escherichia coli in Brassica rapa L. chinensis ...

    African Journals Online (AJOL)

    Low quality water has become valuable resource with restricted or unrestricted use in food production depending on its quality. This study has quantified the occurrence of Escherichia coli in Brassica rapa L. chinensis (Chinese cabbage) vegetables and low quality irrigation water. A total of 106 samples including Chinese ...

  20. Biofilm production and antibiotic susceptibility profile of Escherichia ...

    African Journals Online (AJOL)

    Samie.Amidou

    2012-04-26

    Apr 26, 2012 ... Full Length Research Paper. Biofilm production and antibiotic susceptibility profile of Escherichia coli isolates from HIV and AIDS patients in the Limpopo Province. Samie, A. and Nkgau, T. F.. Department of Microbiology, University of Venda, Private Bag X5050, Thohoyandou 0950, Limpopo, South Africa.

  1. Draft Genome Sequence of Escherichia coli K-12 (ATCC 10798)

    OpenAIRE

    Dimitrova, Daniela; Engelbrecht, Kathleen C.; Putonti, Catherine; Koenig, David W.; Wolfe, Alan J.

    2017-01-01

    ABSTRACT Here, we present the draft genome sequence of Escherichia coli ATCC 10798. E.?coli ATCC 10798 is a K-12 strain, one of the most well-studied model microorganisms. The size of the genome was 4,685,496?bp, with a G+C content of 50.70%. This assembly consists of 62 contigs and the F plasmid.

  2. Physiological responses of Escherichia coli to far-ultraviolet radiation

    International Nuclear Information System (INIS)

    Swenson, P.A.

    1976-01-01

    The following topics are reviewed: photochemical damage to DNA; measurement of cell survival; DNA repair processes and genetics of radiation sensitivity; degradation of DNA and RNA; biochemical and physiological consequences; reactivation of bacteriophage in Escherichia coli cells; filament formation; influence of growth phase on survival after uv irradiation; and post-uv-irradiation treatment

  3. Production of jet fuel precursor monoterpenoids from engineered Escherichia coli

    DEFF Research Database (Denmark)

    Mendez-Perez, Daniel; Alonso-Gutierrez, Jorge; Hu, Qijun

    2017-01-01

    ). FPP biosynthesis diverts the carbon flux from monoterpene production to C15 products and quinone biosynthesis. In this study, we tested a chromosomal mutation of Escherichia coli's native FPP synthase (IspA) to improve GPP availability for the production of monoterpenes using a heterologous mevalonate...

  4. The incidence and antibiotics susceptibility of Escherichia coli O157 ...

    African Journals Online (AJOL)

    STORAGESEVER

    2010-02-22

    Feb 22, 2010 ... The incidence of Escherichia coli 0157: H7 was assessed in meat samples from slaughtered cattle in. Ibadan metropolis by culturing ... high quality farm to fork wholesome and safe meat for public consumption in Nigeria. Key words: EHEC .... Prevalence and in vitro antimicrobial susceptibility. Trop. Vet. 26.

  5. Prevalence of Escherichia coli virulence genes in patients with ...

    African Journals Online (AJOL)

    In this study, we investigated the prevalence of the virulence genes specific for five major pathogroups of diarrheagenic Escherichia coli (DEC) in primary cultures from diarrhoeagenic patients in Burkina Faso. Methodology: From September 2016 to Mars 2017, a total of 211 faecal samples from diarrhoeagenic patients from ...

  6. Comparative Genomics of Escherichia coli Strains Causing Urinary Tract Infections

    DEFF Research Database (Denmark)

    Vejborg, Rebecca Munk; Hancock, Viktoria; Schembri, Mark A.

    2011-01-01

    The virulence determinants of uropathogenic Escherichia coli have been studied extensively over the years, but relatively little is known about what differentiates isolates causing various types of urinary tract infections. In this study, we compared the genomic profiles of 45 strains from a range...

  7. Effect of phytoplankton on Escherichia coli survival in laboratory microcosms

    Science.gov (United States)

    Fecal contamination of water sources is an important water quality issue for agricultural irrigation ponds. Escherichia coli is a common microbial indicator used to evaluate recreational and irrigation water quality. Nuisance algae commonly grow in low- or no-flow irrigation water source The objecti...

  8. Neonatal infections caused by Escherichia coli at the National ...

    African Journals Online (AJOL)

    Background: Escherichia coli (E.coli) has been implicated as a common cause of both early and late onset neonatal infections. The emergence of different strains of E.coli that are multiply resistant to commonly used antibiotics has made continuous antibiotics surveillance relevant. Knowledge about common infections ...

  9. neonatal infections caused by escherichia coli at the national

    African Journals Online (AJOL)

    boaz

    Background: Escherichia coli (E.coli) has been implicated as a common cause of both early and late onset neonatal infections. The emergence of different strains of E.coli that are multiply resistant to commonly used antibiotics has made continuous antibiotics surveillance relevant. Knowledge about common infections ...

  10. Expression and purification of recombinant hemoglobin in Escherichia coli

    DEFF Research Database (Denmark)

    Natarajan, Chandrasekhar; Jiang, Xiaoben; Fago, Angela

    2011-01-01

    BACKGROUND: Recombinant DNA technologies have played a pivotal role in the elucidation of structure-function relationships in hemoglobin (Hb) and other globin proteins. Here we describe the development of a plasmid expression system to synthesize recombinant Hbs in Escherichia coli, and we describe...

  11. Search for Enterohaemorrhagic Escherichia coli O157:H7 and ...

    African Journals Online (AJOL)

    Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 and Salmonella enterica are important zoonotic bacteria responsible for enteric infections in humans. The present study investigated the possible role of kittens in the zoonotic transmission of antimicrobial resistant EHEC O157 and Salmonella enterica to human using ...

  12. Antibiotic Sensitivity Profile of Escherichia coli Isolated from Poultry ...

    African Journals Online (AJOL)

    A cross sectional study involving 300 cloaca swabs from apparently healthy birds from 8 small-medium scale poultry farms in Ibadan Oyo State was carried out. A total of 201 (67%) Escherichia coli isolates were recovered from the birds and they were subjected to in-vitro antibiotic sensitivity test by agar gel diffusion method.

  13. Increasing the permeability of Escherichia coli using MAC13243

    DEFF Research Database (Denmark)

    Muheim, Claudio; Götzke, Hansjörg; Eriksson, Anna U.

    2017-01-01

    molecules that make the outer membrane of Escherichia coli more permeable. We identified MAC13243, an inhibitor of the periplasmic chaperone LolA that traffics lipoproteins from the inner to the outer membrane. We observed that cells were (1) more permeable to the fluorescent probe 1-N...

  14. Molecular characterization of the Escherichia coli asymptomatic bacteriuria strain 83972

    DEFF Research Database (Denmark)

    Klemm, Per; Hancock, Viktoria; Ulett, G.C.

    2006-01-01

    Escherichia coli 83972 is a clinical asymptomatia bacteriuric isolate that is able to colonize the human urinary bladder without inducing an immune response. Here we demonstrate that one of the mechanisms by which this strain has become attenuated is through the mutation of its genes encoding type...

  15. Catheter Related Escherichia hermannii Sepsis in a Haemodialysis Patient

    DEFF Research Database (Denmark)

    Utke Rank, Cecilie; Kristensen, Peter Lommer; Hansen, Dennis Schrøder

    2016-01-01

    Escherichia hermannii is an extremely rare etiological agent of invasive infection, and thus, the bacterium was initially considered non-pathogenic. However, in five previously reported case reports E. hermannii has been implicated as the sole pathogen. Our case report describes blood stream infe...

  16. Inactivation of Escherichia coli by titanium dioxide photocatalytic oxidation.

    Science.gov (United States)

    Titanium dioxide in the anatase crystalline form was used as a photocatalyst to generate hydroxyl radicals in a flowthrough water reactor. Experiments were performed on pure cultures of Escherichia coli in dechlorinated tap water and a surface water sample to evaluate the disinfe...

  17. Expression of green fluorescent protein (GFPuv) in Escherichia coli ...

    African Journals Online (AJOL)

    Administrator

    The recombinant green fluorescent protein (GFPuv) was expressed by transformed cells of Escherichia coli DH5-α grown in LB/amp broth at 37oC, for 8 h and 24 h. To evaluate the effectiveness of different parameters to improve the expression of GFPuv by E. coli, four variable culturing conditions were set up for assays by ...

  18. Complete Genome Sequence of Enterotoxigenic Escherichia coli Siphophage Seurat.

    Science.gov (United States)

    Doan, Dung P; Lessor, Lauren E; Hernandez, Adriana C; Kuty Everett, Gabriel F

    2015-02-26

    Enterotoxigenic Escherichia coli (ETEC) is one of the leading causes of diarrhea in developing countries. Bacteriophage therapy has the potential to aid in the prevention and treatment of ETEC-related illness. To that end, we present here the complete genome of ETEC siphophage Seurat and describe its major features. Copyright © 2015 Doan et al.

  19. Protein export in bacillus subtilis and escherichia coli

    NARCIS (Netherlands)

    Dijl, Jan Maarten van

    1990-01-01

    The export of heterologous proteins in Bacillus subtilis and Escherichia coli is often inefficient. Frequently observed problems are: 1) accumulation of the precursor form of the exported protein in the cytoplasm or in the membrane; 2), inefficient or incorrect processing of the precursor; 3),

  20. Escherichia coli and virus isolated from ''sticky kits''

    DEFF Research Database (Denmark)

    Jørgensen, M.; Scheutz, F.; Strandbygaard, Bertel

    1996-01-01

    A total of 121 Escherichia coli strains isolated from 3-week-old mink kits were serotyped and examined for virulence factors. 56 strains were isolated from healthy kits while 65 were from ''sticky kits''. Among these, 34 different serotypes were detected. No difference in serotypes or the presenc...

  1. Colonization of Enteroaggregative Escherichia coli and Shiga toxin-producing Escherichia coli in chickens and humans in southern Vietnam

    NARCIS (Netherlands)

    Trung, Nguyen Vinh; Nhung, Hoang Ngoc; Carrique-Mas, Juan J.; Mai, Ho Huynh; Tuyen, Ha Thanh; Campbell, James; Nhung, Nguyen Thi; van Minh, Pham; Wagenaar, Jaap A.; Mai, Nguyen Thi Nhu; Hieu, Thai Quoc; Schultsz, Constance; Hoa, Ngo Thi

    2016-01-01

    Enteroaggregative (EAEC) and Shiga-toxin producing Escherichia coli (STEC) are a major cause of diarrhea worldwide. E. coli carrying both virulence factors characteristic for EAEC and STEC and producing extended-spectrum beta-lactamase caused severe and protracted disease during an outbreak of E.

  2. Exacerbation of substrate toxicity by IPTG in Escherichia coli BL21(DE3) carrying a synthetic metabolic pathway.

    Science.gov (United States)

    Dvorak, Pavel; Chrast, Lukas; Nikel, Pablo I; Fedr, Radek; Soucek, Karel; Sedlackova, Miroslava; Chaloupkova, Radka; de Lorenzo, Víctor; Prokop, Zbynek; Damborsky, Jiri

    2015-12-21

    Heterologous expression systems based on promoters inducible with isopropyl-β-D-1-thiogalactopyranoside (IPTG), e.g., Escherichia coli BL21(DE3) and cognate LacI(Q)/P(lacUV5)-T7 vectors, are commonly used for production of recombinant proteins and metabolic pathways. The applicability of such cell factories is limited by the complex physiological burden imposed by overexpression of the exogenous genes during a bioprocess. This burden originates from a combination of stresses that may include competition for the expression machinery, side-reactions due to the activity of the recombinant proteins, or the toxicity of their substrates, products and intermediates. However, the physiological impact of IPTG-induced conditional expression on the recombinant host under such harsh conditions is often overlooked. The physiological responses to IPTG of the E. coli BL21(DE3) strain and three different recombinants carrying a synthetic metabolic pathway for biodegradation of the toxic anthropogenic pollutant 1,2,3-trichloropropane (TCP) were investigated using plating, flow cytometry, and electron microscopy. Collected data revealed unexpected negative synergistic effect of inducer of the expression system and toxic substrate resulting in pronounced physiological stress. Replacing IPTG with the natural sugar effector lactose greatly reduced such stress, demonstrating that the effect was due to the original inducer's chemical properties. IPTG is not an innocuous inducer; instead, it exacerbates the toxicity of haloalkane substrate and causes appreciable damage to the E. coli BL21(DE3) host, which is already bearing a metabolic burden due to its content of plasmids carrying the genes of the synthetic metabolic pathway. The concentration of IPTG can be effectively tuned to mitigate this negative effect. Importantly, we show that induction with lactose, the natural inducer of P lac , dramatically lightens the burden without reducing the efficiency of the synthetic TCP degradation

  3. Genetic Transfer of Salmonella typhimurium and Escherichia coli Lipopolysaccharide Antigens to Escherichia coli K-12

    Science.gov (United States)

    Jones, Randall T.; Koeltzow, Donald E.; Stocker, B. A. D.

    1972-01-01

    Escherichia coli K-12 ϰ971 was crossed with a smooth Salmonella typhimurium donor, HfrK6, which transfers early the ilv-linked rfa region determining lipopolysaccharide (LPS) core structure. Two ilv+ hybrids differing in their response to the LPS-specific phages FO and C21 were then crossed with S. typhimurium HfrK9, which transfers early the rfb gene cluster determining O repeat unit structure. Most recombinants selected for his+ (near rfb) were agglutinated by Salmonella factor 4 antiserum. Transfer of an F′ factor (FS400) carrying the rfb–his region of S. typhimurium to the same two ilv+ hybrids gave similar results. LPS extracted from two ilv+,his+, factor 4-positive hybrids contained abequose, the immunodominant sugar for factor 4 specificity. By contrast, his+ hybrids obtained from ϰ971 itself by similar HfrK9 and F′FS400 crosses were not agglutinated by factor 4 antiserum, indicating that the parental E. coli ϰ971 does not have the capacity to attach Salmonella O repeat units to its LPS core. It is concluded that the Salmonella rfb genes are expressed only in E. coli ϰ971 hybrids which have also acquired ilv-linked genes (presumably rfa genes affecting core structure or O-translocase ability, or both) from a S. typhimurium donor. When E. coli ϰ971 was crossed with a smooth E. coli donor, Hfr59, of serotype O8, which transfers his early, most his+ recombinants were agglutinated by E. coli O8 antiserum and lysed by the O8-specific phage, Ω8. This suggests that, although the parental E. coli K-12 strain ϰ971 cannot attach Salmonella-specific repeat units to its LPS core, it does have the capacity to attach E. coli O8-specific repeat units. PMID:4559827

  4. Tunable Control of an Escherichia coli Expression System for the Overproduction of Membrane Proteins by Titrated Expression of a Mutant lac Repressor.

    Science.gov (United States)

    Kim, Seong Keun; Lee, Dae-Hee; Kim, Oh Cheol; Kim, Jihyun F; Yoon, Sung Ho

    2017-09-15

    Most inducible expression systems suffer from growth defects, leaky basal induction, and inhomogeneous expression levels within a host cell population. These difficulties are most prominent with the overproduction of membrane proteins that are toxic to host cells. Here, we developed an Escherichia coli inducible expression system for membrane protein production based on titrated expression of a mutant lac repressor (mLacI). Performance of the mLacI inducible system was evaluated in conjunction with commonly used lac operator-based expression vectors using a T7 or tac promoter. Remarkably, expression of a target gene can be titrated by the dose-dependent addition of l-rhamnose, and the expression levels were homogeneous in the cell population. The developed system was successfully applied to overexpress three membrane proteins that were otherwise difficult to produce in E. coli. This gene expression control system can be easily applied to a broad range of existing protein expression systems and should be useful in constructing genetic circuits that require precise output signals.

  5. Improved antifungal activity of barley derived chitinase I gene that overexpress a 32 kDa recombinant chitinase in Escherichia coli host

    Directory of Open Access Journals (Sweden)

    Nida Toufiq

    Full Text Available Abstract Agricultural crops suffer many diseases, including fungal and bacterial infections, causing significant yield losses. The identification and characterisation of pathogenesis-related protein genes, such as chitinases, can lead to reduction in pathogen growth, thereby increasing tolerance against fungal pathogens. In the present study, the chitinase I gene was isolated from the genomic DNA of Barley (Hordeum vulgare L. cultivar, Haider-93. The isolated DNA was used as template for the amplification of the ∼935 bp full-length chitinase I gene. Based on the sequence of the amplified gene fragment, class I barley chitinase shares 93% amino acid sequence homology with class II wheat chitinase. Interestingly, barley class I chitinase and class II chitinase do not share sequence homology. Furthermore, the amplified fragment was expressed in Escherichia coli Rosetta strain under the control of T7 promoter in pET 30a vector. Recombinant chitinase protein of 35 kDa exhibited highest expression at 0.5 mM concentration of IPTG. Expressed recombinant protein of 35 kDa was purified to homogeneity with affinity chromatography. Following purification, a Western blot assay for recombinant chitinase protein measuring 35 kDa was developed with His-tag specific antibodies. The purified recombinant chitinase protein was demonstrated to inhibit significantly the important phytopathogenic fungi Alternaria solani, Fusarium spp, Rhizoctonia solani and Verticillium dahliae compared to the control at concentrations of 80 µg and 200 µg.

  6. Pathogenic Escherichia coli and food handlers in luxury hotels in Nairobi, Kenya.

    Science.gov (United States)

    Onyango, Abel O; Kenya, Eucharia U; Mbithi, John J N; Ng'ayo, Musa O

    2009-11-01

    The epidemiology and virulence properties of pathogenic Escherichia coli among food handlers in tourist destination hotels in Kenya are largely uncharacterized. This cross-sectional study among consenting 885 food handlers working in nine luxurious tourist hotels in Nairobi, Kenya determined the epidemiology, virulence properties, antibiotics susceptibility profiles and conjugation abilities of pathogenic Escherichia coli. Pathogenic Escherichia coli was detected among 39 (4.4%) subjects, including 1.8% enteroaggregative Escherichia coli (EAEC) harboring aggR genes, 1.2% enterotoxigenic Escherichia coli (ETEC) expressing both LT and STp toxins, 1.1% enteropathogenic Escherichia coli (EPEC) and 0.2% Shiga-like Escherichia coli (EHEC) both harboring eaeA and stx2 genes respectively. All the pathotypes had increased surface hydrophobicity. Using multivariate analyses, food handlers with loose stools were more likely to be infected with pathogenic Escherichia coli. Majority 53.8% of the pathotypes were resistant to tetracycline with 40.2% being multi-drug resistant. About 85.7% pathotypes trans-conjugated with Escherichia coli K12 F(-) NA(r) LA. The carriage of multi-drug resistant, toxin expressing pathogenic Escherichia coli by this population is of public health concern because exposure to low doses can result in infection. Screening food handlers and implementing public awareness programs is recommended as an intervention to control transmission of enteric pathogens.

  7. The Escherichia coli transcriptome linked to growth fitness

    Directory of Open Access Journals (Sweden)

    Bei-Wen Ying

    2016-03-01

    Full Text Available A series of Escherichia coli strains with varied genomic sequences were subjected to high-density microarray analyses to elucidate the fitness-correlated transcriptomes. Fitness, which is commonly evaluated by the growth rate during the exponential phase, is not only determined by the genome but is also linked to growth conditions, e.g., temperature. We previously reported genetic and environmental contributions to E. coli transcriptomes and evolutionary transcriptome changes in thermal adaptation. Here, we describe experimental details on how to prepare microarray samples that truly represent the growth fitness of the E. coli cells. A step-by-step record of sample preparation procedures that correspond to growing cells and transcriptome data sets that are deposited at the GEO database (GSE33212, GSE52770, GSE61739 are also provided for reference. Keywords: Transcriptome, Growth fitness, Escherichia coli, Microarray

  8. Escherichia fergusonii Associated with Pneumonia in a Beef Cow

    Directory of Open Access Journals (Sweden)

    Guillermo M. Rimoldi

    2013-01-01

    Full Text Available An adult Angus cow developed hyperthermia, prostration, and respiratory distress, dying 36 hours after the onset of clinical signs. The main finding during postmortem examination was a severe focally extensive pneumonia. Icterus and a chronic mastitis were also noticed. Histologic examination of the lungs detected fibrinonecrotic pneumonia, with large number of oat cells and intralesional Gram-negative bacterial colonies. Samples from lung lesions were collected, and a pure growth of Escherichia fergusonii was obtained. E. fergusonii is a member of Enterobacteriaceae, related to Escherichia coli and Salmonella sp. In veterinary medicine, E. fergusonii has been reported in calves and sheep with clinical cases suggestive of salmonellosis; in a horse and a goat with enteritis and septicemia; and in ostriches with fibrinonecrotic typhlitis. To our knowledge, this report represents the first description of E. fergusonii associated with an acute pneumonia in cattle.

  9. Tranformasi Fragmen Dna Kromosom Xanthomonas Campestris ke dalam Escherichia Coli

    Directory of Open Access Journals (Sweden)

    Wibowo Mangunwardoyo

    2002-04-01

    Full Text Available Research on DNA transformation of Xanthomonas campestris into Escherichia coli DH5αα using plasmid vector Escherichia coli (pUC19. was carried out. DNA chromosome was isolated using CTAB method, alkali lysis method was used to isolate DNA plasmid. Both of DNA plasmid and chromosome were digested using restriction enzyme EcoRI. Competent cell was prepared with CaCl2 and heat shock method for transformation procedure. The result revealed transformation obtain 5 white colonies, with transformation frequency was 1,22 x 10-8 colony/competent cell. Electrophoresis analysis showed the DNA fragment (insert in range 0.5 – 7,5 kb. Further research should be carried out to prepare the genomic library to obtain better result of transformant.

  10. Respiration shutoff in Escherichia coli after far-uv irradiation

    International Nuclear Information System (INIS)

    Swenson, P.A.; Norton, I.L.

    1984-01-01

    Damage to DNA of Escherichia coli by uv, ionizing radiation and chemicals causes a number of responses that require the recA + and lexA + gene products. The responses include error prone repair (as indicated by mutagenesis), filamentation and induction of prophage lambda. Another important rec/lex response, shutoff of respiration, which occurs 60 min after exposure to uv, is studied. Objective is to understand the genetic and biochemical bases of the shutoff process and its control

  11. flu, a metastable gene controlling surface properties of Escherichia coli.

    OpenAIRE

    Diderichsen, B

    1980-01-01

    flu, a gene of Escherichia coli K-12, was discovered and mapped between his and shiA. It is shown that flu is a metastable gene that changes frequently between the flu+ and flu states. flu+ variants give stable homogeneous suspensions, are piliated, and form glossy colonies. flu variants aggregate, fluff and sediment from suspensions, are nonpiliated, and form frizzy colonies. flu+ and flu variants can be isolated from most strains. Implications of these observations are discussed, and it is ...

  12. Multiplex PCR Assay for Identification of Human Diarrheagenic Escherichia coli

    OpenAIRE

    Toma, Claudia; Lu, Yan; Higa, Naomi; Nakasone, Noboru; Chinen, Isabel; Baschkier, Ariela; Rivas, Marta; Iwanaga, Masaaki

    2003-01-01

    A multiplex PCR assay for the identification of human diarrheagenic Escherichia coli was developed. The targets selected for each category were eae for enteropathogenic E. coli, stx for Shiga toxin-producing E. coli, elt and est for enterotoxigenic E. coli, ipaH for enteroinvasive E. coli, and aggR for enteroaggregative E. coli. This assay allowed the categorization of a diarrheagenic E. coli strain in a single reaction tube.

  13. Two Tales of Prokaryotic Genomic Diversity: Escherichia coli and Halophiles

    Directory of Open Access Journals (Sweden)

    Lejla Pašić

    2014-01-01

    Full Text Available Prokaryotes are generally characterized by vast genomic diversity that has been shaped by mutations, horizontal gene transfer, bacteriocins and phage predation. Enormous genetic diversity has developed as a result of stresses imposed in harsh environments and the ability of microorganisms to adapt. Two examples of prokaryotic diversity are presented: on intraspecies level, exemplified by Escherichia coli, and the diversity of the hypersaline environment, with the discussion of food-related health issues and biotechnological potential.

  14. SENSITIVITY TEST OF Escherichia coli AGAINST EXTRACT Tinospora crispa

    OpenAIRE

    Lucia Ratna Winata Muslimin; abdul wahid jamaluddin

    2017-01-01

    In general, a bacterium such as Escherichia coli produces a kind of toxic protein which can disrupt intestinal wall. Livestock reacts to these toxins by pumping lots of water into the intestine in order to rinse or flush these toxins. As a result, the livestocks have diarrhea as a body response to remove the toxin in the digestive system. In the presence of these problems, breeders take a measure such as using antibiotics freely. Among breeders, antibiotics are often used freely ...

  15. Multidrug-Resistant Escherichia fergusonii: a Case of Acute Cystitis▿

    Science.gov (United States)

    Savini, Vincenzo; Catavitello, Chiara; Talia, Marzia; Manna, Assunta; Pompetti, Franca; Favaro, Marco; Fontana, Carla; Febbo, Fabio; Balbinot, Andrea; Di Berardino, Fabio; Di Bonaventura, Giovanni; Di Zacomo, Silvia; Esattore, Francesca; D'Antonio, Domenico

    2008-01-01

    We report a case in which Escherichia fergusonii, an emerging pathogen in various types of infections, was associated with cystitis in a 52-year-old woman. The offending strain was found to be multidrug resistant. Despite in vitro activity, beta-lactam treatment failed because of a lack of patient compliance with therapy. The work confirms the pathogenic potential of E. fergusonii. PMID:18256229

  16. Nosocomial acquisition of Escherichia coli by infants delivered in hospitals.

    Science.gov (United States)

    Fujita, K; Murono, K

    1996-04-01

    The delivery of infants in hospitals is desirable for obstetric reasons, but exposes the neonates to the microbiological hazards of a maternity unit. When neonates are born and cared for in hospital, the Escherichia coli strains that colonize the intestine tend to be acquired from the environment or from other babies, and are potentially pathogenic. The colonization of the infant with maternal flora should be promoted by strict rooming-in of mother and baby, or by delivery at home.

  17. Removal of Escherichia coli from biological effluents using natural ...

    African Journals Online (AJOL)

    Ability for disinfecting sterile biological effluents inoculated with Escherichia coli ATCC 25922 at concentrations of 105 CFU/m., using a natural mineral aggregate (NMA) and artificial mineral aggregates (AMAfs) consisting of individual oxides as Fe2O3, Cu2O y Ag2O and combined oxides as Fe2O3-Cu2O, Fe2O3-Ag2O, ...

  18. Optimizing the feeding operation of recombinant Escherichia coli ...

    African Journals Online (AJOL)

    Recombinant Escherichia coli BL21 was used to produce human-like collagen in fed-batch culture. After building and analyzing the kinetic models of fed-batch cultures, the maximum specific growth rate, Yx/s and Yp/s were 0.411 h-1 , 0.428 g·g-1 and 0.0716 g/g, respectively. The square error of cell growth models, glucose ...

  19. Draft Genome Sequence of Escherichia coli K-12 (ATCC 10798).

    Science.gov (United States)

    Dimitrova, Daniela; Engelbrecht, Kathleen C; Putonti, Catherine; Koenig, David W; Wolfe, Alan J

    2017-07-06

    Here, we present the draft genome sequence of Escherichia coli ATCC 10798. E. coli ATCC 10798 is a K-12 strain, one of the most well-studied model microorganisms. The size of the genome was 4,685,496 bp, with a G+C content of 50.70%. This assembly consists of 62 contigs and the F plasmid. Copyright © 2017 Dimitrova et al.

  20. Distribution of Diverse Escherichia coli between Cattle and Pasture

    OpenAIRE

    NandaKafle, Gitanjali; Seale, Tarren; Flint, Toby; Nepal, Madhav; Venter, Stephanus N.; Brözel, Volker S.

    2017-01-01

    Escherichia coli is widely considered to not survive for extended periods outside the intestines of warm-blooded animals; however, recent studies demonstrated that E. coli strains maintain populations in soil and water without any known fecal contamination. The objective of this study was to investigate whether the niche partitioning of E. coli occurs between cattle and their pasture. We attempted to clarify whether E. coli from bovine feces differs phenotypically and genotypically from isola...

  1. Prevalence of Antibiotic-Resistant Strains of Escherichia coli in ...

    African Journals Online (AJOL)

    A total of six bacteria species Escherichia coli, Pseudomonas aeruginosa, Bacillus cereus, Klebsiella pneumonia, Staphylococcus aureus, Enterobacter aerogenes were ... Énumération de nombre de plaque standard a été effectuée par la méthode de la plaque de propagation sur des échantillons d'eau dilués en série.

  2. Genes and proteins of Escherichia coli K-12.

    Science.gov (United States)

    Riley, M

    1998-01-01

    GenProtEC is a database of Escherichia coli genes and their gene products, classified by type of function and physiological role and with citations to the literature for each. Also present are data on sequence similarities among E.coli proteins, representing groups of paralogous genes, with PAM values, percent identity of amino acids, length of alignment and percent aligned. GenProtEC can be accessed at the URL http://www.mbl.edu/html/ecoli.html

  3. Predictors Of Non-Escherichia Coli Urinary Tract Infection.

    Science.gov (United States)

    Shaikh, Nader; Wald, Ellen R; Keren, Ron; Gotman, Nathan; Ivanova, Anastasia; Carpenter, Myra A; Moxey-Mims, Marva; Hoberman, Alejandro

    2016-11-01

    We aimed to determine which children are prone to non-Escherichia coli urinary tract infection (UTIs). We included 769 children with UTI. We found that circumcised males, Hispanic children, children without fever and children with grades 3 and 4 vesicoureteral reflux were more likely to have a UTI caused by organisms other than E. coli. This information may guide clinicians in their choice of antimicrobial therapy.

  4. Metabolic and Transcriptional Response to Cofactor Perturbations in Escherichia coli

    DEFF Research Database (Denmark)

    Holm, Anders Koefoed; Blank, L.M.; Oldiges, M.

    2010-01-01

    Metabolic cofactors such as NADH and ATP play important roles in a large number of cellular reactions, and it is of great interest to dissect the role of these cofactors in different aspects of metabolism. Toward this goal, we overexpressed NADH oxidase and the soluble F1-ATPase in Escherichia coli...... of redox and energy metabolism and should help in developing metabolic engineering strategies in E. coli....

  5. Ribosome slowed by mutation to streptomycin resistance. [Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Galas, D J; Branscomb, E W

    1976-08-12

    The effect of mutation to streptomycin resistance on the speed of polypeptide elongation in Escherichia coli was investigated. Translation speed was determined by measuring the time required for the first newly synthesized ..beta..-galactosidase molecules to appear after induction of the lactose operon. The results showed that ribosome speed is not a fixed parameter inherent to the protein synthetic apparatus, but a variable determined by the kinetics of translation and ultimately by the structure of the ribosome. (HLW)

  6. Compilation and analysis of Escherichia coli promoter DNA sequences.

    OpenAIRE

    Hawley, D K; McClure, W R

    1983-01-01

    The DNA sequence of 168 promoter regions (-50 to +10) for Escherichia coli RNA polymerase were compiled. The complete listing was divided into two groups depending upon whether or not the promoter had been defined by genetic (promoter mutations) or biochemical (5' end determination) criteria. A consensus promoter sequence based on homologies among 112 well-defined promoters was determined that was in substantial agreement with previous compilations. In addition, we have tabulated 98 promoter ...

  7. [Virulence markers of Escherichia coli O1 strains].

    Science.gov (United States)

    Makarova, M A; Kaftyreva, L A; Grigor'eva, N S; Kicha, E V; Lipatova, L A

    2011-01-01

    To detect virulence genes in clinical isolates of Escherichia coli O1 using polymerase chain reaction (PCR). One hundred and twenty strains of E.coli O1 strains isolated from faeces of patients with acute diarrhea (n = 45) and healthy persons (n = 75) were studied. PCR with primers for rfb and fliC genes, which control synthesis of O- and H- antigens respectively, was used. Fourteen virulence genes (pap, aaf, sfa, afa, eaeA, bfpA, ial, hly, cnf, stx1, stx2, lt, st, and aer) were detected by PCR primers. K1-antigen was determined by Pastorex Meningo B/E. coli O1 kit (Bio-Rad). rfb gene controlling O-antigen synthesis in serogroup O1 as well as fliC gene controlling synthesis of H7 and K1 antigens were detected in all strains. Thus all E. coli strains had antigenic structure O1:K1 :H-:F7. Virulence genes aafl, sfa, afa, eaeA, bfpA, ial, hly, cnf, stx1, stx2, lt, and st were not detected. All strains owned pap and aer genes regardless of the presence of acute diarrhea symptoms. It was shown that E. coli O1:KI:H-:F7 strains do not have virulence genes which are characteristic for diarrhea-causing Escherichia. In accordance with the presence of pap and aer genes they could be attributed to uropathogenic Escherichia (UPEC) or avian-pathogenic Escherichia (APEC). It is necessary to detect virulence factors in order to determine E. coli as a cause of intestinal infection.

  8. Biomolecular Mechanisms of Pseudomonas aeruginosa and Escherichia coli Biofilm Formation

    Science.gov (United States)

    Laverty, Garry; Gorman, Sean P.; Gilmore, Brendan F.

    2014-01-01

    Pseudomonas aeruginosa and Escherichia coli are the most prevalent Gram-negative biofilm forming medical device associated pathogens, particularly with respect to catheter associated urinary tract infections. In a similar manner to Gram-positive bacteria, Gram-negative biofilm formation is fundamentally determined by a series of steps outlined more fully in this review, namely adhesion, cellular aggregation, and the production of an extracellular polymeric matrix. More specifically this review will explore the biosynthesis and role of pili and flagella in Gram-negative adhesion and accumulation on surfaces in Pseudomonas aeruginosa and Escherichia coli. The process of biofilm maturation is compared and contrasted in both species, namely the production of the exopolysaccharides via the polysaccharide synthesis locus (Psl), pellicle Formation (Pel) and alginic acid synthesis in Pseudomonas aeruginosa, and UDP-4-amino-4-deoxy-l-arabinose and colonic acid synthesis in Escherichia coli. An emphasis is placed on the importance of the LuxR homologue sdiA; the luxS/autoinducer-II; an autoinducer-III/epinephrine/norepinephrine and indole mediated Quorum sensing systems in enabling Gram-negative bacteria to adapt to their environments. The majority of Gram-negative biofilms consist of polysaccharides of a simple sugar structure (either homo- or heteropolysaccharides) that provide an optimum environment for the survival and maturation of bacteria, allowing them to display increased resistance to antibiotics and predation. PMID:25438014

  9. Antibiotic resistance of Verotoxigenic Escherichia coli isolated from vegetables

    Directory of Open Access Journals (Sweden)

    mojtaba boniadian

    2017-01-01

    Full Text Available Introduction: Human gastrointestinal disease caused by verotoxigenic Escherichia coli has been diagnosed for recent decades. Escherichia coli O157:H7 is the most important serotype of verotoxigenic Escherichia coli that cause hemolytic uremic syndrome and hemorrhagic colitis in humans. This study was conducted to determine the occurrence of verotoxigenic E. coli and antibiotic resistance of the isolates from vegetables. Materials and methods: A total of 500 fresh vegetable samples were collected randomly from retail shops in Shahrekord, Iran. E. coli was isolated and identified using bacteriological and biochemical tests. PCR method was used to identify the rbfE, stx1, stx2 and eae genes. Also, antibiotic resistance of the isolates was determined by disk diffusion method. Results: The results represented that among 25 isolates possess virulence genes, 40, 12 and 4% of the isolates contained eaeA, STx2, and both genes, respectively. But none of them contained H7, STx1, and rfbE genes. The antibiotic resistance pattern demonstrated that the isolates were highly resistant to Gentamycin and cefotoxime. Discussion and conclusion: The results of this study showed that the presence of verotoxigenic E.coli in vegetables; and high resistance of the isolates to antibiotics could be hazardous for public health.

  10. Genomic diversity of Escherichia isolates from diverse habitats.

    Directory of Open Access Journals (Sweden)

    Seungdae Oh

    Full Text Available Our understanding of the Escherichia genus is heavily biased toward pathogenic or commensal isolates from human or animal hosts. Recent studies have recovered Escherichia isolates that persist, and even grow, outside these hosts. Although the environmental isolates are typically phylogenetically distinct, they are highly related to and phenotypically indistinguishable from their human counterparts, including for the coliform test. To gain insights into the genomic diversity of Escherichia isolates from diverse habitats, including freshwater, soil, animal, and human sources, we carried out comparative DNA-DNA hybridizations using a multi-genome E. coli DNA microarray. The microarray was validated based on hybridizations with selected strains whose genome sequences were available and used to assess the frequency of microarray false positive and negative signals. Our results showed that human fecal isolates share two sets of genes (n>90 that are rarely found among environmental isolates, including genes presumably important for evading host immune mechanisms (e.g., a multi-drug transporter for acids and antimicrobials and adhering to epithelial cells (e.g., hemolysin E and fimbrial-like adhesin protein. These results imply that environmental isolates are characterized by decreased ability to colonize host cells relative to human isolates. Our study also provides gene markers that can distinguish human isolates from those of warm-blooded animal and environmental origins, and thus can be used to more reliably assess fecal contamination in natural ecosystems.

  11. Biomolecular Mechanisms of Pseudomonas aeruginosa and Escherichia coli Biofilm Formation

    Directory of Open Access Journals (Sweden)

    Garry Laverty

    2014-07-01

    Full Text Available Pseudomonas aeruginosa and Escherichia coli are the most prevalent Gram-negative biofilm forming medical device associated pathogens, particularly with respect to catheter associated urinary tract infections. In a similar manner to Gram-positive bacteria, Gram-negative biofilm formation is fundamentally determined by a series of steps outlined more fully in this review, namely adhesion, cellular aggregation, and the production of an extracellular polymeric matrix. More specifically this review will explore the biosynthesis and role of pili and flagella in Gram-negative adhesion and accumulation on surfaces in Pseudomonas aeruginosa and Escherichia coli. The process of biofilm maturation is compared and contrasted in both species, namely the production of the exopolysaccharides via the polysaccharide synthesis locus (Psl, pellicle Formation (Pel and alginic acid synthesis in Pseudomonas aeruginosa, and UDP-4-amino-4-deoxy-l-arabinose and colonic acid synthesis in Escherichia coli. An emphasis is placed on the importance of the LuxR homologue sdiA; the luxS/autoinducer-II; an autoinducer-III/epinephrine/norepinephrine and indole mediated Quorum sensing systems in enabling Gram-negative bacteria to adapt to their environments. The majority of Gram-negative biofilms consist of polysaccharides of a simple sugar structure (either homo- or heteropolysaccharides that provide an optimum environment for the survival and maturation of bacteria, allowing them to display increased resistance to antibiotics and predation.

  12. Escherichia coli. A sanitary methodology for faecal water pollution tests; Escherichia coli nelle acque. Significato sanitario e metodologie di analisi

    Energy Technology Data Exchange (ETDEWEB)

    Bonadonna, L. [Istituto Superiore di Sanita' , Rome (Italy)

    2001-02-01

    Among the traditional indictors of faecal water pollution, Escherichia coli has shown to fit better with the definition of indicator organism. Till now its recovery has been time-consuming and needs confirmation tests. In this report more rapid and direct methods, based on enzymatic reactions, are presented. [Italian] Per talune peculiari caratteristiche, Escherichia coli sembra meglio soddisfare i requisiti insiti nella definizione di organismo indicatore, rispetto ai tradizionali indicatori di contaminazione fecale dell'acqua. Finora, i substrati disponibili per il suo rilevamento necessitano tutti di almeno una prova di conferma. Di qui l'esigenza di indicare metodi di rilevamento a riposta piu' rapida, anche in relazione all'inserimento, nelle piu' recenti normative nazionali ed europee, del microrganismo tra i parametri microbiologici da ricercare.

  13. Characterization of a new high copy Stowaway family MITE, BRAMI-1 in Brassica genome

    Science.gov (United States)

    2013-01-01

    Background Miniature inverted-repeat transposable elements (MITEs) are expected to play important roles in evolution of genes and genome in plants, especially in the highly duplicated plant genomes. Various MITE families and their roles in plants have been characterized. However, there have been fewer studies of MITE families and their potential roles in evolution of the recently triplicated Brassica genome. Results We identified a new MITE family, BRAMI-1, belonging to the Stowaway super-family in the Brassica genome. In silico mapping revealed that 697 members are dispersed throughout the euchromatic regions of the B. rapa pseudo-chromosomes. Among them, 548 members (78.6%) are located in gene-rich regions, less than 3 kb from genes. In addition, we identified 516 and 15 members in the 470 Mb and 15 Mb genomic shotgun sequences currently available for B. oleracea and B. napus, respectively. The resulting estimated copy numbers for the entire genomes were 1440, 1464 and 2490 in B. rapa, B. oleracea and B. napus, respectively. Concurrently, only 70 members of the related Arabidopsis ATTIRTA-1 MITE family were identified in the Arabidopsis genome. Phylogenetic analysis revealed that BRAMI-1 elements proliferated in the Brassica genus after divergence from the Arabidopsis lineage. MITE insertion polymorphism (MIP) was inspected for 50 BRAMI-1 members, revealing high levels of insertion polymorphism between and within species of Brassica that clarify BRAMI-1 activation periods up to the present. Comparative analysis of the 71 genes harbouring the BRAMI-1 elements with their non-insertion paralogs (NIPs) showed that the BRAMI-1 insertions mainly reside in non-coding sequences and that the expression levels of genes with the elements differ from those of their NIPs. Conclusion A Stowaway family MITE, named as BRAMI-1, was gradually amplified and remained present in over than 1400 copies in each of three Brassica species. Overall, 78% of the members were identified in gene-rich regions, and it is assumed that they may contribute to the evolution of duplicated genes in the highly duplicated Brassica genome. The resulting MIPs can serve as a good source of DNA markers for Brassica crops because the insertions are highly dispersed in the gene-rich euchromatin region and are polymorphic between or within species. PMID:23547712

  14. High-copy sequences reveal distinct evolution of the rye B chromosome

    Czech Academy of Sciences Publication Activity Database

    Klemme, S.; Banaei-Moghaddam, A.M.; Macas, Jiří; Wicker, T.; Novák, Petr; Houben, A.

    2013-01-01

    Roč. 199, č. 2 (2013), s. 550-558 ISSN 0028-646X R&D Projects: GA ČR GBP501/12/G090 Institutional support: RVO:60077344 Keywords : satellite DNA * nondisjunction control region * B chromosome * Secale cereale (rye) Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.545, year: 2013

  15. Cancerous patients and outbreak of Escherichia coli: an important issue in oncology

    OpenAIRE

    Joob, Beuy; Wiwanitkit, Viroj

    2014-01-01

    The widespread of the Escherichia coli outbreak in Europe becomes an important public concern at global level. The infection can be serious and might result in death. The retrospective literature review on this specific topic is performed. In this specific brief article, the author presented and discussed on the problem of Escherichia coli infection in the cancerous patients. This is an actual important issue in medical oncology for the scenario of Escherichia coli epidemic.

  16. Novel roles for the AIDA adhesin from diarrheagenic Escherichia coli:

    DEFF Research Database (Denmark)

    Sherlock, Orla; Schembri, Mark; Reisner, A.

    2004-01-01

    Diarrhea-causing Escherichia coli strains are responsible for numerous cases of gastrointestinal disease and constitute a serious health problem throughout the world. The ability to recognize and attach to host intestinal surfaces is an essential step in the pathogenesis of such strains. AIDA...... binds to mammalian cells. Here, we show that AIDA possesses self-association characteristics and can mediate autoaggregation of E. coli cells. We demonstrate that intercellular AIDA-AIDA interaction is responsible for bacterial autoaggregation. Interestingly, AIDA-expressing cells can interact...

  17. Cellular localization of the Escherichia coli SpoT protein.

    OpenAIRE

    Gentry, D R; Cashel, M

    1995-01-01

    The SpoT protein of Escherichia coli serves as a source of degradation as well as an apparent source of synthesis of (p)ppGpp. Since the subcellular localization of SpoT might be a clue to its function, we have used SpoT-specific antisera to analyze cell extracts fractionated on sucrose gradients. We find that the SpoT protein is not bound to ribosomes or to either inner or outer membrane fractions. Although the SpoT protein is found in large aggregates, its localization is probably cytosolic.

  18. Escherichia coli O26 IN RAW BUFFALO MILK: PRELIMINARY RESULTS

    Directory of Open Access Journals (Sweden)

    A. Rella

    2013-02-01

    Full Text Available Escherichia coli O26 is considered to be one of the most important food-borne pathogen. In this study, 120 buffalo milk samples collected in Lazio and in Apulia regions were tested for the presence of E. coli O26. One buffalo milk sample (0,8% tested positive for E. coli O26; the isolate was positive at the verocytotoxicity test and it showed resistance properties to different antimicrobial classes. These preliminary results highlight the need to monitor the foods of animal origin used for production and eaten by a wide range of persons, respect VTEC organism.

  19. Protein abundance profiling of the Escherichia coli cytosol

    DEFF Research Database (Denmark)

    Ishihama, Y.; Schmidt, T.; Rappsilber, J.

    2008-01-01

    sample. Using a combination of LC-MS/MS approaches with protein and peptide fractionation steps we identified 1103 proteins from the cytosolic fraction of the Escherichia coli strain MC4100. A measure of abundance is presented for each of the identified proteins, based on the recently developed emPAI...... approach which takes into account the number of sequenced peptides per protein. The values of abundance are within a broad range and accurately reflect independently measured copy numbers per cell. As expected, the most abundant proteins were those involved in protein synthesis, most notably ribosomal...

  20. DNA microarray analysis of fim mutations in Escherichia coli

    DEFF Research Database (Denmark)

    Schembri, Mark; Ussery, David; Workman, Christopher

    2002-01-01

    Bacterial adhesion is often mediated by complex polymeric surface structures referred to as fimbriae. Type I fimbriae of Escherichia coli represent the archetypical and best characterised fimbrial system. These adhesive organelles mediate binding to D-mannose and are directly associated...... we have used DNA microarray analysis to examine the molecular events involved in response to fimbrial gene expression in E. coli K-12. Observed differential expression levels of the fim genes were in good agreement with our current knowledge of the stoichiometry of type I fimbriae. Changes in fim...

  1. 4-thiouridine and photoprotection in Escherichia coli K12

    International Nuclear Information System (INIS)

    Thomas, Gilles; Favre, Alain

    1977-01-01

    A high level of protection is observed in the Escherichia coli K 12 strain AB 1157 rec A 1 nuv + whose transfer RNA contains 4-thiouridine. In contrast, the photoprotection level is low and observed at higher doses in a strain which differs from the former by a single mutation nuv - , (lack of 4-thiouridine). This nucleoside is therefore an important chromophore leading to photoprotection. This conclusion is corroborated by the similarity of the action spectra for 8-13 link formation in tRNA and for photoprotection [fr

  2. UV irradiation alters deoxynucleoside triphosphate pools in Escherichia coli

    International Nuclear Information System (INIS)

    Das, S.K.; Loeb, L.A.

    1984-01-01

    UV irradiation of exponentially growing Escherichia coli increased intracellular concentration of dATP and dTTP without significantly changing the concentrations of dGTP and dCTP. These selective increases in dATP and dTTP pools are seen in wild-type E. coli K12 and AB1157, as well as in recA and umuC strains, and are proportional to UV dose. The possible significance of these findings with respect to induction of the SOS response and nontargeted mutagenesis are discussed. (orig.)

  3. A stochastic killing system for biological containment of Escherichia coli

    DEFF Research Database (Denmark)

    Klemm, P.; Jensen, Lars Bogø; Molin, Søren

    1995-01-01

    Bacteria with a stochastic conditional lethal containment system have been constructed. The invertible switch promoter located upstream of the fimA gene from Escherichia coli was inserted as expression cassette in front of the Lethal gef gene deleted of its own natural promoter. The resulting...... fusion was placed on a plasmid and transformed to E. coli. The phenotype connected with the presence of such a plasmid was to reduce the population growth rate with increasing significance as the cell growth rate was reduced. In very fast growing cells, there was no measurable effect on growth rate. When...

  4. Chromosomal replication incompatibility in Dam methyltransferase deficient Escherichia coli cells

    DEFF Research Database (Denmark)

    Freiesleben, Ulrik Von

    1996-01-01

    Dam methyltransferase deficient Escherichia coli cells containing minichromosomes were constructed. Free plasmid DNA could not be detected in these cells and the minichromosomes were found to be integrated in multiple copies in the origin of replication (oriC) region of the host chromosome....... The absence of the initiation cascade in Dam- cells is proposed to account for this observation of apparent incompatibility between plasmid and chromosomal copies of oriC. Studies using oriC-pBR322 chimeric plasmids and their deletion derivatives indicated that the incompatibility determinant is an intact...

  5. Dysfunctional MreB inhibits chromosome segregation in Escherichia coli

    DEFF Research Database (Denmark)

    Kruse, Thomas; Møller-Jensen, Jakob; Løbner-Olesen, Anders

    2003-01-01

    The mechanism of prokaryotic chromosome segregation is not known. MreB, an actin homolog, is a shape-determining factor in rod-shaped prokaryotic cells. Using immunofluorescence microscopy we found that MreB of Escherichia coli formed helical filaments located beneath the cell surface. Flow...... cytometric and cytological analyses indicated that MreB-depleted cells segregated their chromosomes in pairs, consistent with chromosome cohesion. Overexpression of wild-type MreB inhibited cell division but did not perturb chromosome segregation. Overexpression of mutant forms of MreB inhibited cell...... that MreB filaments participate in directional chromosome movement and segregation....

  6. Suppressors of DnaAATP imposed overinitiation in Escherichia coli

    DEFF Research Database (Denmark)

    Charbon, Godefroid; Riber, Leise; Cohen, Malene

    2011-01-01

    Chromosome replication in Escherichia coli is limited by the supply of DnaA associated with ATP. Cells deficient in RIDA (Regulatory Inactivation of DnaA) due to a deletion of the hda gene accumulate suppressor mutations (hsm) to counteract the overinitiation caused by an elevated DnaAATP level....... Eight spontaneous hda suppressor mutations were identified by whole-genome sequencing, and three of these were analysed further. Two mutations (hsm-2 and hsm-4) mapped in the dnaA gene and led to a reduced ability to initiate replication from oriC. One mutation (hsm-1) mapped to the seqA promoter...

  7. Stabilization of Escherichia coli uridine phosphorylase by evolution and immobilization

    CSIR Research Space (South Africa)

    Visser, Daniel F

    2010-08-01

    Full Text Available nucleoside phosphorylase (BHPNP1) from the thermotolerant alkalophile Bacillus halodurans with the Escherichia coli uridine phosphorylase (EcUP) (EC 2.4.2.3) in a one-pot cascade reaction can produce 5-MU in high yield [2, 3]. The optimal operating... reaction temperature of 60?C is within the thermostability range of BHPNP, but the stability of the UP is only 40?C. This requires higher enzyme loading to offset the rate of thermal deactivation. Moreover, due to the low solubility of the reaction...

  8. Spontaneous Escherichia coli Meningitis Associated with Hemophagocytic Lymphohistiocytosis

    Directory of Open Access Journals (Sweden)

    Kuo-Hsuan Chang

    2006-01-01

    Full Text Available Spontaneous Escherichia coli meningitis has not been previously reported in association with hemophago-cytic lymphohistiocytosis (HLH. A previously healthy 72-year-old woman was admitted due to fever, nuchal rigidity, disturbed consciousness and splenomegaly. Anemia, thrombocytopenia and hyperfer-ritinemia developed on the 8th day of hospitalization. Cultures of cerebrospinal fluid and blood grew E. coli. Abundant macrophages overwhelmed erythrocytes in the bone marrow aspirate, confirming the presence of hemophagocytosis. E. coli meningitis was managed with a 40-day course of antibiotic treatment. However, the severity of anemia and thrombocytopenia progressed despite intensive transfusion therapy. The patient died of HLH on the 60th day of hospitalization.

  9. Causes, prevention and treatment of Escherichia coli infections.

    Science.gov (United States)

    Gould, Dinah

    Escherichia coli is a normal inhabitant of the human gastrointestinal tract and can cause healthcare-associated infections. The organism is most frequently responsible for urinary tract infections and it is the bacterium most often implicated in the cause of diarrhoea in people travelling overseas. In recent years, a strain called Ecoli O157 has gained notoriety for causing foodborne infection, which can have severe health consequences, especially in young children. This article describes the range of different infections caused by Ecoli in healthcare settings and the community and discusses the characteristics of the different strains of the bacteria that explain variations in their pathogenicity.

  10. Genes and proteins of Escherichia coli (GenProtEc).

    Science.gov (United States)

    Riley, M; Space, D B

    1996-01-01

    GenProtEc is a database of Escherichia coli genes and their gene products, classified by type of function and physiological role and with citations to the literature for each. Also present are data on sequence similarities among E.coli proteins with PAM values, percent identity of amino acids, length of alignment and percent aligned. The database is available as a PKZip file by ftp from mbl.edu/pub/ecoli.exe. The program runs under MS-DOS on IMB-compatible machines. GenProtEc can also be accessed through the World Wide Web at URL http://mbl.edu/html/ecoli.html.

  11. FimH-mediated autoaggregation of Escherichia coli

    DEFF Research Database (Denmark)

    Schembri, Mark; Christiansen, G.; Klemm, Per

    2001-01-01

    Autoaggregation is a phenomenon thought to contribute to colonization of mammalian hosts by pathogenic bacteria. Type 1 fimbriae are surface organelles of Escherichia coli that mediate D-mannose-sensitive binding to various host surfaces. This binding is conferred by the minor fimbrial component...... FimH. In this study, we have used random mutagenesis to identify variants of the FimH adhesin that confer the ability of E. coli to autoaggregate and settle from liquid cultures. Three separate autoaggregating clones were identified, all of which contained multiple amino acid changes located within...

  12. Escherichia coli promoter sequences predict in vitro RNA polymerase selectivity.

    Science.gov (United States)

    Mulligan, M E; Hawley, D K; Entriken, R; McClure, W R

    1984-01-11

    We describe a simple algorithm for computing a homology score for Escherichia coli promoters based on DNA sequence alone. The homology score was related to 31 values, measured in vitro, of RNA polymerase selectivity, which we define as the product KBk2, the apparent second order rate constant for open complex formation. We found that promoter strength could be predicted to within a factor of +/-4.1 in KBk2 over a range of 10(4) in the same parameter. The quantitative evaluation was linked to an automated (Apple II) procedure for searching and evaluating possible promoters in DNA sequence files.

  13. Escherichia coli promoter sequences predict in vitro RNA polymerase selectivity.

    OpenAIRE

    Mulligan, M E; Hawley, D K; Entriken, R; McClure, W R

    1984-01-01

    We describe a simple algorithm for computing a homology score for Escherichia coli promoters based on DNA sequence alone. The homology score was related to 31 values, measured in vitro, of RNA polymerase selectivity, which we define as the product KBk2, the apparent second order rate constant for open complex formation. We found that promoter strength could be predicted to within a factor of +/-4.1 in KBk2 over a range of 10(4) in the same parameter. The quantitative evaluation was linked to ...

  14. Escherichia coli : host interactions in the pathogenesis of canine pyometra

    OpenAIRE

    Henriques, Sofia Correia Rosa de Barros

    2016-01-01

    Tese de Doutoramento em Ciências Veterinárias na Especialidade de Ciências Biológicas e Biomédicas Canine pyometra develops as a result of a complex interaction of etiological and physiopathological factors, such as the virulence and type of the bacteria and the individual host defence mechanisms. Since Escherichia coli is the most common bacterium isolated from uterus of bitches with pyometra, one main objective of this work was to characterize E. coli virulence potential, and...

  15. Prevalence of Antibiotic Resistance in Escherichia coli Isolated from Poultry Meat Supply in Isfahan

    Directory of Open Access Journals (Sweden)

    Farhad Safarpordehkordi

    2014-08-01

    Conclusions: Despite the high contamination rate of chicken meat with Escherichia coli, majority of isolates had high resistance to common antibiotics. Complete cooking of meat and avoid indiscriminate prescribing of antibiotics, preventing the occurrence of food poisoning due to resistant Escherichia coli.

  16. the occurrence of escherichia coli o157:h7 in market and abattoir

    African Journals Online (AJOL)

    user

    Escherichia coli O157:H7 is a newly emerging pathogen frequently associated with the consumption of foods of ... KEY WORDS: E. coli O157:H7, Pathogen, Abattoir, Market, and Infections ..... pathogen. Escherichia coli O157:H7 as a model of.

  17. Lon gene and photoprotection in Escherichia coli K-12

    Energy Technology Data Exchange (ETDEWEB)

    Waksman, G.; Thomas, G.; Favre, A. (Institut de Recherche en Biologie Moleculaire, Group de Photobiologie Moleculaire, Paris (France))

    1984-03-01

    Photoprotection, i.e. the increased resistance of the cells preilluminated with near ultraviolet light (300-380 nm) to the lethal action of 254nm radiations requires either an integrated prophage or a recA mutation in Escherichia coli K12 strains. Significant photoprotection occurs in an Escherichia coli K12 recA/sup +/ cell containing the lon allele responsible for filamentous growth after 254nm irradiation. The Fil phenotype can be suppressed by the sfiA or sfiB suppressor genes. Since the E. coli K12 recA/sup +/ lon sfiB strain exhibits no more photoprotection, it is concluded that in lon strains photoprotection is due to the abolition of the 254nm induced filamentation by the near ultraviolet treatment. In addition, near ultraviolet illumination of the cells leads to a severe restriction of the bulk protein synthesis. This effect is observed only in nuv/sup +/ cells that contain 4-thiouridine the chromophore responsible for photoprotection. It is proposed that in lon (lysogenic strains) photoprotection is due to prevention of the SOS response. During the growth lag, the low residual level of protein synthesis does not allow the induction of the SOS response and accordingly prevents filamentation (the lytic cycle).

  18. Inactivation of Escherichia coli in soil by solarization

    International Nuclear Information System (INIS)

    Wu, S.; Nishihara, M.; Kawasaki, Y.; Yokoyama, A.; Matsuura, K.; Koga, T.; Ueno, D.; Inoue, K.; Someya, T.

    2009-01-01

    Contamination of agricultural soil by fecal pathogenic bacteria poses a potential risk of infection to humans. For the biosafety control of field soil, soil solarization in an upland field was examined to determine the efficiency of solarization on the inactivation of Escherichia coli inoculated into soil as a model microorganism for human pathogenic bacteria. Soil solarization, carried out by sprinkling water and covering the soil surface with thin plastic sheets, greatly increased the soil temperature. The daily average temperature of the solarized soil was 4–10°C higher than that of the non-solarized soil and fluctuated between 31 and 38°C. The daily highest temperature reached more than 40°C for 8 days in total in the solarized soil during the second and third weeks of the experiment. Escherichia coli in the solarized soil became undetectable (< 0.08 c.f.u. g −1 dry soil) within 4 weeks as a result, whereas E. coli survived for more than 6 weeks in the non-solarized soil. Soil solarization, however, had little influence on the total direct count and total viable count of bacteria in the soil. These results indicate that soil solarization would be useful for the biosafety control of soil contaminated by human pathogens via immature compost or animal feces. (author)

  19. Environmental Escherichia coli: Ecology and public health implications - A review

    Science.gov (United States)

    Jang, Jeonghwan; Hur, Hor-Gil; Sadowsky, Michael J.; Byappanahalli, Muruleedhara; Yan, Tao; Ishii, Satoshi

    2017-01-01

    Escherichia coli is classified as a rod-shaped, Gram-negative bacterium in the family Enterobacteriaceae. The bacterium mainly inhabits the lower intestinal tract of warm-blooded animals, including humans, and is often discharged into the environment through feces or wastewater effluent. The presence of E. coli in environmental waters has long been considered as an indicator of recent fecal pollution. However, numerous recent studies have reported that some specific strains of E. coli can survive for long periods of time, and potentially reproduce, in extra-intestinal environments. This indicates that E. coli can be integrated into indigenous microbial communities in the environment. This naturalization phenomenon calls into question the reliability of E. coli as a fecal indicator bacterium (FIB). Recently, many studies reported that E. coli populations in the environment are affected by ambient environmental conditions affecting their long-term survival. Large-scale studies of population genetics provide the diversity and complexity of E. coli strains in various environments, affected by multiple environmental factors. This review examines the current knowledge on the ecology of E. coli strains in various environments in regards to its role as a FIB and as a naturalized member of indigenous microbial communities. Special emphasis is given on the growth of pathogenic E. coli in the environment, and the population genetics of environmental members of the genus Escherichia. The impact of environmental E. coli on water quality and public health is also discussed.

  20. Damage-induced DNA repair processes in Escherichia coli cells

    International Nuclear Information System (INIS)

    Slezarikova, V.

    1986-01-01

    The existing knowledge is summed up of the response of Escherichia coli cells to DNA damage due to various factors including ultraviolet radiation. So far, three inducible mechanisms caused by DNA damage are known, viz., SOS induction, adaptation and thermal shock induction. Greatest attention is devoted to SOS induction. Its mechanism is described and the importance of the lexA recA proteins is shown. In addition, direct or indirect role is played by other proteins, such as the ssb protein binding the single-strand DNA sections. The results are reported of a study of induced repair processes in Escherichia coli cells repeatedly irradiated with UV radiation. A model of induction by repeated cell irradiation discovered a new role of induced proteins, i.e., the elimination of alkali-labile points in the daughter DNA synthetized on a damaged model. The nature of the alkali-labile points has so far been unclear. In the adaptation process, regulation proteins are synthetized whose production is induced by the presence of alkylation agents. In the thermal shock induction, new proteins synthetize in cells, whose function has not yet been clarified. (E.S.)

  1. The lon gene and photoprotection in Escherichia coli K-12

    International Nuclear Information System (INIS)

    Waksman, G.; Thomas, G.; Favre, A.

    1984-01-01

    Photoprotection, i.e. the increased resistance of the cells preilluminated with near ultraviolet light (300-380 nm) to the lethal action of 254nm radiations requires either an integrated prophage or a recA mutation in Escherichia coli K12 strains. Significant photoprotection occurs in an Escherichia coli K12 recA + cell containing the lon allele responsible for filamentous growth after 254nm irradiation. The Fil phenotype can be suppressed by the sfiA or sfiB suppressor genes. Since the E. coli K12 recA + lon sfiB strain exhibits no more photoprotection, it is concluded that in lon strains photoprotection is due to the abolition of the 254nm induced filamentation by the near ultraviolet treatment. In addition, near ultraviolet illumination of the cells leads to a severe restriction of the bulk protein synthesis. This effect is observed only in nuv + cells that contain 4-thiouridine the chromophore responsible for photoprotection. It is proposed that in lon (lysogenic strains) photoprotection is due to prevention of the SOS response. During the growth lag, the low residual level of protein synthesis does not allow the induction of the SOS response and accordingly prevents filamentation (the lytic cycle). (author)

  2. Dynamics of Escherichia coli Chromosome Segregation during Multifork Replication

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck; Youngren, Brenda; Hansen, Flemming G.

    2007-01-01

    Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division, the chro......Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division......, the chromosomes contain multiple replication forks and must be segregated while this complex pattern of replication is still ongoing. Here, we show that replication and segregation continue in step, starting at the origin and progressing to the replication terminus. Thus, early-replicated markers on the multiple......-branched chromosomes continue to separate soon after replication to form separate protonucleoids, even though they are not segregated into different daughter cells until later generations. The segregation pattern follows the pattern of chromosome replication and does not follow the cell division cycle. No extensive...

  3. MUTATIONAL SYNERGISM BETWEEN RADIATIONS AND METHYLATED PURINES IN ESCHERICHIA COLI

    Science.gov (United States)

    Doneson, Ira N.; Shankel, Delbert M.

    1964-01-01

    Doneson, Ira N. (University of Kansas, Lawrence), and Delbert M. Shankel. Mutational synergism between radiations and methylalted purines in Escherichia coli. J. Bacteriol. 87:61–67. 1964.—A synergistic mutational effect was demonstrated between low doses of ultraviolet light and the methylated purines caffeine, theophylline, and theobromine. Caffeine produced the greatest effect and theobromine the least effect. The magnitude of the synergism was inversely related to the ultraviolet dosage. A large percentage of the synergistic effect could be “photoprevented” by exposure of the ultraviolet-treated cells to white light prior to exposure to the analogues. The consequence of the combined treatment occurred only when the chemical treatment followed the ultraviolet treatment. Furthermore, it was necessary to administer the chemical treatment soon after the ultraviolet treatment or the mutants were “lost.” When cells were treated with low dosages of ultraviolet light and of X irradiation (X ray), the result was merely additive, and combinations of X ray and chemical treatment yielded no synergism. Synchronous growth studies indicated that a particular growth stage of the organisms was most susceptible to the synergistic effect. The mutation studied was that of Escherichia coli B/r to high-level streptomycin resistance. PMID:14102875

  4. Viabilidad de Escherichia coli en presencia de diferentes contaminantes

    Directory of Open Access Journals (Sweden)

    Antonio Rivera T

    2006-04-01

    Full Text Available La contaminación en ríos condiciona la presencia de microorganismos adaptados al ecosistema entre ellos a patógenos de importancia en salud pública. Objetivo: Determinar la viabilidad de Escherichia coli en presencia de nitrato de plata, carbonato de amonio, fenol y formaldehído. Materiales y métodos: Se tomaron muestras de agua del río Alseseca, que luego se sembró en medios de cultivo selectivos para enterobacterias, seleccionándose las colonias del género Escherichia, las cuales fueron sembradas en el medio de orientación CHROMagar ECC. Las muestras de E. coli se evaluaron en presencia de nitrato de plata, carbonato de amonio, fenol y formaldehído. Resultados: El grupo experimental presentó viabilidad en presencia de los cuatro compuestos, el grupo control positivo presentó nula viabilidad, la comparación entre los grupos mostró diferencia significativa (p< 0,05. Conclusión: Los aislamientos de E. coli mostraron viabilidad, implicando riesgos para el ecosistemas y la salud, ya que el río Alseseca atraviesa por el municipio de Puebla donde existen núcleos poblacionales importantes.

  5. Growth of the modeling of Escherichia coli in milk

    International Nuclear Information System (INIS)

    Mbangu, N.; Malakasa, M.; Ekalakala, T.; N'dendje, B.; Abedi, M.; Muzembe, K.; Bandejile, M.

    2010-01-01

    Escherichia coli is a contaminant potential of milk. Collective toxinfections implying the bacterium and milk were announced of share the world. However, no identified work proposed a mathematical expression of the growth of the bacterium in milk. The interest of such a step is however undeniable. Under specified conditions, the mathematical formulation of the growth provides the means of considering the population bacterial when the analyses cannot be carried out. It also makes it possible to test the negatiable instruments of the unfavourable circumstances supposed suchas chain breakage of cold on the development of the microbial charge. This work established mathematical expressions of the growth of Escherichia coli in milk for part of its range of temperature of growth suboptimale i.e. between 25 and 35 Deg C. It was not possible to generalize these expressions for predictions on all the range of temperature suboptimal. This work also made it possible to highlight a deviation of the behavoir of the bacterium compared to the model of Ratkowsky without however that it is not possible to provide a univocal explanantion of it. Varoius assumptions were put forth referring to either a singularity of the behavior of the bacterium or a skew of the value of its minimal temperature of growth

  6. Research on killing Escherichia Coli by reactive oxygen species based on strong ionization discharging plasma

    International Nuclear Information System (INIS)

    Li, Y J; Tian, Y P; Zhang, Z T; Li, R H; Cai, L J; Gao, J Y

    2013-01-01

    Reactive oxygen species solution produced by strong ionization discharging plasma was used to kill Escherichia coli by spraying. Several effect factors such as pH value, solution temperature, spraying time and exposure time were observed in this study, and their effects on killing rate of Escherichia coli were discussed and analysed. Results show that the treating efficiency of ROS solution for Escherichia coli is higher in alkaline solution than that in acid solution. The killing rate of Escherichia coli increases while the spraying time and exposure time are longer and the temperature is lower. The effects of different factors on killing rate of Escherichia coli are as follows: spraying time > pH value > exposure time > solution temperature.

  7. Microbial electrolytic disinfection process for highly efficient Escherichia coli inactivation

    DEFF Research Database (Denmark)

    Zhou, Shaofeng; Huang, Shaobin; Li, Xiaohu

    2018-01-01

    extensively studied for recalcitrant organics removal, its application potential towards water disinfection (e.g., inactivation of pathogens) is still unknown. This study investigated the inactivation of Escherichia coli in a microbial electrolysis cell based bio-electro-Fenton system (renamed as microbial......Water quality deterioration caused by a wide variety of recalcitrant organics and pathogenic microorganisms has become a serious concern worldwide. Bio-electro-Fenton systems have been considered as cost-effective and highly efficient water treatment platform technology. While it has been......]OH was identified as one potential mechanism for disinfection. This study successfully demonstrated the feasibility of bio-electro-Fenton process for pathogens inactivation, which offers insight for the future development of sustainable, efficient, and cost-effective biological water treatment technology....

  8. Alterations induced in Escherichia Coli cells by gamma radiation

    International Nuclear Information System (INIS)

    Kappke, J.; Schelin, H.R.; Paschuk, S.A.; Denyak, V.; Silva, E.R. da; Jesus, E.F.O. de; Lopes, R.T.; Carlin, N.; Toledo, E.S.

    2007-01-01

    Modifications occurred in Escherichia coli cells exposed to gamma radiation ( 60 Co source) were investigated. The irradiations were done at the LIN-COPPE laboratory of the UFRJ and the analysis at the Biology Department of the UTFPR. The E. coli cells were irradiated with 30, 60, 90, 120, 150, 180, 210, 240, 300, 480, 600 e 750 Gy doses. The samples were analyzed with Gram-stain, biochemical tests in EPM, MIO and Lysine Broth, Simmons Cytrate Medium and Rhamnose Broth, antibiogram and isolation of auxotrophic mutants. It was observed that for the received doses the E. coli did not show morphological alterations in the tests. Some E. Coli cells showed to be able to deaminade the L-tryptophan or they changed their sensibility for amoxillin and cephaloonine after the irradiation. The existence of aauxotrophic mutants after irradiation was also verified. (author)

  9. Expression of maize prolamins in Escherichia Coli. [Zea mays L

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Szu-zhen; Esen, Asim

    1985-12-02

    A cDNA expression library of developing corn (Zea mays L.) endosperm has been constructed using plasmid pUC8 as vector and Escherichia coli strain DH1 as host. The expression library was screened with non-radioactive immunological probes to detect the expression of gamma-zein and alpha-zein. When anti-gamma-zein antibody was used as the probe, 23 colonies gave positive reactions. The lengths of cDNA inserts of the 23 colonies were found to be 250-900 base pairs. When anti-alpha zein antibody was used, however, fewer colonies gave positive reactions. The library was also screened by colony-hybridization with /sup 32/P-labeled DNA probes. Based on immunological and hybridization screening of the library and other evidence, it was conclude that alpha-zein was either toxic to E. coli cells or rapidly degraded whereas gamma-zein and its fragments were readily expressed. 21 references.

  10. Mechanisms of the radioprotective effect of cysteamine in Escherichia coli

    International Nuclear Information System (INIS)

    Korystov, Yu.N.; Vexler, F.B.

    1988-01-01

    The values of the oxygen effect (m) and the maximal protective effect of cysteamine (DMF*) were estimated for four Escherichia coli strains: AB1157 (wild type), AB1886 (uvrA), AB2463 (recA), and p3478 (polA). A correlation made between DMF* and m as well as the kinetics of the increase of DMF with oxygen depletion showed that the protective effect of cysteamine is realized by three mechanisms: (i) anoxia achieved by oxygen reduction, with the DMF varying from 2.2 to 4.2 for different E. coli strains (this protection is the major contribution to the entire mechanism); (ii) lowering of the indirect radiation effect; i.e., for 50 mM cysteamine DMF does not exceed 1.1; and (iii) increase of the efficiency of enzymatic repair. The latter effect of cysteamine is registered only with the wild-type E. coli, the DMF being not less than 1.4

  11. Impact of cranberry on Escherichia coli cellular surface characteristics

    International Nuclear Information System (INIS)

    Johnson, Brandy J.; Lin Baochuan; Dinderman, Michael A.; Rubin, Robert A.; Malanoski, Anthony P.; Ligler, Frances S.

    2008-01-01

    The anti-adhesive effects of cranberry have been attributed to both interactions of its components with the surface of bacterial cells and to inhibition of p-fimbriae expression. Previous reports also suggested that the presence of cranberry juice changed the Gram stain characteristics of Escherichia coli. Here, we show that the morphology of E. coli is changed when grown in the presence of juice or extract from Vaccinium macrocarpon (cranberry). Gene expression analysis indicates the down regulation of flagellar basal body rod and motor proteins. Consistent with this finding and previous reports, the SEM images indicate a decrease in the visible p-fimbriae. The iodine used in Gram-staining protocols was found to interact differently with the bacterial membrane when cells were cultured in spiked media. Slight alterations in the Gram stain protocol demonstrated that culturing in the presence of cranberry juice does not change the Gram stain characteristics contradicting other reports.

  12. Structure of Escherichia coli Hfq bound to polyriboadenylate RNA

    DEFF Research Database (Denmark)

    Link, Todd M; Valentin-Hansen, Poul; Brennan, Richard G

    2009-01-01

    (A) RNA, A(15). The structure reveals a unique RNA binding mechanism. Unlike uridine-containing sequences, which bind to the "proximal" face, the poly(A) tract binds to the "distal" face of Hfq using 6 tripartite binding motifs. Each motif consists of an adenosine specificity site (A site), which......Hfq is a small, highly abundant hexameric protein that is found in many bacteria and plays a critical role in mRNA expression and RNA stability. As an "RNA chaperone," Hfq binds AU-rich sequences and facilitates the trans annealing of small RNAs (sRNAs) to their target mRNAs, typically resulting...... in the down-regulation of gene expression. Hfq also plays a key role in bacterial RNA decay by binding tightly to polyadenylate [poly(A)] tracts. The structural mechanism by which Hfq recognizes and binds poly(A) is unknown. Here, we report the crystal structure of Escherichia coli Hfq bound to the poly...

  13. Two proline porters in Escherichia coli K-12.

    Science.gov (United States)

    Stalmach, M E; Grothe, S; Wood, J M

    1983-11-01

    Escherichia coli mutants defective at putP and putA lack proline transport via proline porter I and proline dehydrogenase activity, respectively. They retain a proline uptake system (proline porter II) that is induced during tryptophan-limited growth and are sensitive to the toxic L-proline analog, 3,4-dehydroproline. 3,4-Dehydroproline-resistant mutants derived from a putP putA mutant lack proline porter II. Auxotrophic derivatives derived from putP+ or putP bacteria can grow if provided with proline at low concentration (25 microM); those derived from the 3,4-dehydroproline-resistant mutants require high proline for growth (2.5 mM). We conclude that E. coli, like Salmonella typhimurium, possesses a second proline porter that is inactivated by mutations at the proP locus.

  14. Sickness behavior in dairy cows during Escherichia coli mastitis

    DEFF Research Database (Denmark)

    Fogsgaard, Katrine Kop; Røntved, Christine Maria; Sørensen, Peter

    2012-01-01

    The consequences of mastitis in terms of dairy cow behavior are relatively unknown. Future assessment of dairy cow welfare during mastitis will be facilitated by knowledge about the potential of mastitis to induce sickness behavior. Our aim was to examine behavior of dairy cows in the period from 2...... d before (d −2 and −1) to 3 d (d 0, 1, and 2) after experimental intramammary challenge with Escherichia coli. Effects of experimentally induced mastitis on behavior were examined in 20 primiparous Danish Holstein-Friesian cows, all 3 to 6 wk after calving and kept in tie stalls. After evening....... This knowledge can be useful for the development of welfare assessment protocols, early disease detection, and for future work aimed at understanding the behavioral needs of dairy cows suffering from mastitis....

  15. Purification and characterization of Escherichia coli MreB protein.

    Science.gov (United States)

    Nurse, Pearl; Marians, Kenneth J

    2013-02-01

    The actin homolog MreB is required in rod-shaped bacteria for maintenance of cell shape and is intimately connected to the holoenzyme that synthesizes the peptidoglycan layer. The protein has been reported variously to exist in helical loops under the cell surface, to rotate, and to move in patches in both directions around the cell surface. Studies of the Escherichia coli protein in vitro have been hampered by its tendency to aggregate. Here we report the purification and characterization of native E. coli MreB. The protein requires ATP hydrolysis for polymerization, forms bundles with a left-hand twist that can be as long as 4 μm, forms sheets in the presence of calcium, and has a critical concentration for polymerization of 1.5 μM.

  16. Purification and Characterization of Escherichia coli MreB Protein*

    Science.gov (United States)

    Nurse, Pearl; Marians, Kenneth J.

    2013-01-01

    The actin homolog MreB is required in rod-shaped bacteria for maintenance of cell shape and is intimately connected to the holoenzyme that synthesizes the peptidoglycan layer. The protein has been reported variously to exist in helical loops under the cell surface, to rotate, and to move in patches in both directions around the cell surface. Studies of the Escherichia coli protein in vitro have been hampered by its tendency to aggregate. Here we report the purification and characterization of native E. coli MreB. The protein requires ATP hydrolysis for polymerization, forms bundles with a left-hand twist that can be as long as 4 μm, forms sheets in the presence of calcium, and has a critical concentration for polymerization of 1.5 μm. PMID:23235161

  17. Induced clustering of Escherichia coli by acoustic fields.

    Science.gov (United States)

    Gutiérrez-Ramos, Salomé; Hoyos, Mauricio; Ruiz-Suárez, J C

    2018-03-16

    Brownian or self-propelled particles in aqueous suspensions can be trapped by acoustic fields generated by piezoelectric transducers usually at frequencies in the megahertz. The obtained confinement allows the study of rich collective behaviours like clustering or spreading dynamics in microgravity-like conditions. The acoustic field induces the levitation of self-propelled particles and provides secondary lateral forces to capture them at nodal planes. Here, we give a step forward in the field of confined active matter, reporting levitation experiments of bacterial suspensions of Escherichia coli. Clustering of living bacteria is monitored as a function of time, where different behaviours are clearly distinguished. Upon the removal of the acoustic signal, bacteria rapidly spread, impelled by their own swimming. Nevertheless, long periods of confinement result in irreversible bacteria entanglements that could act as seeds for levitating bacterial aggregates.

  18. Antibiotic treatment of verocytotoxin-producing Escherichia coli (VTEC) infection

    DEFF Research Database (Denmark)

    Agger, Morten; Scheutz, Flemming; Villumsen, Steen

    2015-01-01

    OBJECTIVES: A consensus has existed on not to treat verocytotoxin-producing Escherichia coli (VTEC)-infected individuals with antibiotics because of possible subsequent increased risk of developing haemolytic uraemic syndrome (HUS). The aim of this systematic review is to clarify the risk...... associated with antibiotic treatment during acute VTEC infection and in chronic VTEC carrier states. METHODS: A systematic search in PubMed identified 1 meta-analysis, 10 clinical studies and 22 in vitro/in vivo studies. RESULTS: Four clinical studies found an increased risk of HUS, four studies found...... no altered risk of HUS and two studies found a protective effect of antibiotics. In vitro and clinical studies suggest that DNA synthesis inhibitors should be avoided, whereas evidence from in vitro studies indicates that certain protein and cell wall synthesis inhibitors reduce the release of toxins from...

  19. ROS mediated selection for increased NADPH availability in Escherichia coli.

    Science.gov (United States)

    Reynolds, Thomas S; Courtney, Colleen M; Erickson, Keesha E; Wolfe, Lisa M; Chatterjee, Anushree; Nagpal, Prashant; Gill, Ryan T

    2017-11-01

    The economical production of chemicals and fuels by microbial processes remains an intense area of interest in biotechnology. A key limitation in such efforts concerns the availability of key co-factors, in this case NADPH, required for target pathways. Many of the strategies pursued for increasing NADPH availability in Escherichia coli involve manipulations to the central metabolism, which can create redox imbalances and overall growth defects. In this study we used a reactive oxygen species based selection to search for novel methods of increasing NADPH availability. We report a loss of function mutation in the gene hdfR appears to increase NADPH availability in E. coli. Additionally, we show this excess NADPH can be used to improve the production of 3HP in E. coli. © 2017 Wiley Periodicals, Inc.

  20. Regulation of gene expression in Escherichia coli and its bacteriophage

    International Nuclear Information System (INIS)

    Higgins, C.F.

    1986-01-01

    This chapter reviews the study of prokaryotic gene expression beginning with a look at the regulation of the lactose operon and the mechanism of attenuation in the tryptophan operon to the more recent development of recombinant DNA technology. The chapter deals almost entirely with escherichia coli and its bacteriophage. The only experimental technique which the authors explore in some detail is the construction and use of gene and operon fusions which have revolutionized the study of gene expression. Various mechanisms by which E. Coli regulate the cellular levels of individual messenger-RNA species are described. Translational regulation of the cellular levels of messenger-RNA include signals encoded within the messenger-RNA molecule itself and regulatory molecules which interact with the messenger-RNA and alter it translational efficiency

  1. Carbon and energy metabolism of atp mutants of Escherichia coli

    DEFF Research Database (Denmark)

    Jensen, Peter Ruhdal; Michelsen, Ole

    1992-01-01

    strain is not able to utilize the resulting proton motive force for ATP synthesis. Indeed, the ratio of ATP concentration to ADP concentration was decreased from 19 in the wild type to 7 in the atp mutant, and the membrane potential of the atp deletion strain was increased by 20%, confirming......The membrane-bound H+-ATPase plays a key role in free-energy transduction of biological systems. We report how the carbon and energy metabolism of Escherichia coli changes in response to deletion of the atp operon that encodes this enzyme. Compared with the isogenic wild-type strain, the growth...... rate and growth yield were decreased less than expected for a shift from oxidative phosphorylation to glycolysis alone as a source of ATP. Moreover, the respiration rate of a atp deletion strain was increased by 40% compared with the wild-type strain. This result is surprising, since the atp deletion...

  2. Sedimentation and gravitational instability of Escherichia coli Suspension

    Science.gov (United States)

    Salin, Dominique; Douarche, Carine

    2017-11-01

    The successive runs and tumbles of Escherichia coli bacteria provide an active matter suspension of rod-like particles with a large swimming, Brownian like, diffusion. As opposed to inactive elongated particles, this diffusion prevents clustering of the particles and hence instability in the gravity field. We measure the time dependent E . coli concentration profile during their sedimentation. After some hours, due to the dioxygen consumption, a motile / non-motile front forms leading to a Rayleigh-Taylor type gravitational instability. Analysing both sedimentation and instability in the framework of active particle suspensions, we can measure the relevant bacteria hydrodynamic characteristics such as its single particle sedimentation velocity and its hindrance volume. Comparing these quantities to the ones of equivalent passive particles (ellipsoid, rod) we tentatively infer the effective shape and size of the bacteria involved in its buoyancy induced advection and diffusion. Laboratoire FAST University Paris Saclay France.

  3. DNA turnover and strand breaks in Escherichia coli

    International Nuclear Information System (INIS)

    Hanawalt, P.; Grivell, A.; Nakayama, H.

    1975-01-01

    The extent of DNA turnover has been measured in a dnaB mutant of Escherichia coli, temperature sensitive for semiconservative DNA replication. At the nonpermissive temperature about 0.02 percent of the deoxynucleotides in DNA are exchanged per generation period. This turnover rate is markedly depressed in the presence of rifampicin. During thymine starvation strand breaks accumulate in the DNA of E. coli strains that are susceptible to thymineless death. Rifampicin suppresses the appearance of these breaks, consistent with our hypothesis that transcription may be accompanied by repairable single-strand breaks in DNA. DNA turnover is enhanced severalfold in strands containing 5-bromodeoxyuridine in place of thymidine, possibly because the analog (or the deoxyuridine, following debromination) is sometimes recognized and excised

  4. The Resistome: A Comprehensive Database of Escherichia coli Resistance Phenotypes.

    Science.gov (United States)

    Winkler, James D; Halweg-Edwards, Andrea L; Erickson, Keesha E; Choudhury, Alaksh; Pines, Gur; Gill, Ryan T

    2016-12-16

    The microbial ability to resist stressful environmental conditions and chemical inhibitors is of great industrial and medical interest. Much of the data related to mutation-based stress resistance, however, is scattered through the academic literature, making it difficult to apply systematic analyses to this wealth of information. To address this issue, we introduce the Resistome database: a literature-curated collection of Escherichia coli genotypes-phenotypes containing over 5,000 mutants that resist hundreds of compounds and environmental conditions. We use the Resistome to understand our current state of knowledge regarding resistance and to detect potential synergy or antagonism between resistance phenotypes. Our data set represents one of the most comprehensive collections of genomic data related to resistance currently available. Future development will focus on the construction of a combined genomic-transcriptomic-proteomic framework for understanding E. coli's resistance biology. The Resistome can be downloaded at https://bitbucket.org/jdwinkler/resistome_release/overview .

  5. Bactericidal activity of ciprofloxacin upon Escherichia coli and Acinetobacter baumanni.

    Science.gov (United States)

    Zemelman, R; Vejar, C; Bello, H; Domínguez, M; González, G

    1992-01-01

    The mechanisms of bactericidal activity of ciprofloxacin (mechanisms A and B) upon cells of a strain of Escherichia coli and one strain of Acinetobacter baumannii were investigated under different conditions. The killing of E. coli cells by ciprofloxacin was significantly reduced by chloramphenicol, but this antibiotic showed almost no activity upon killing of A. baumannii cells by this quinolone. Similar results were obtained when rifampicin was added to ciprofloxacin. Bactericidal activity of ciprofloxacin upon nondividing cells of E. coli was lower and that upon non-dividing cells of A. baumannii was not affected when compared with activity of ciprofloxacin upon dividing cells of both microorganisms. These results demonstrate that the antibacterial activity of ciprofloxacin upon A. baumannii is independent of protein and ARN synthesis, a fact which suggests that this quinolone exerts only bactericidal mechanism B upon A. baumannii. This finding might explain, at least in part, the lower susceptibility of this microorganism to ciprofloxacin.

  6. Expression and purification of recombinant hemoglobin in Escherichia coli

    DEFF Research Database (Denmark)

    Natarajan, Chandrasekhar; Jiang, Xiaoben; Fago, Angela

    2011-01-01

    BACKGROUND: Recombinant DNA technologies have played a pivotal role in the elucidation of structure-function relationships in hemoglobin (Hb) and other globin proteins. Here we describe the development of a plasmid expression system to synthesize recombinant Hbs in Escherichia coli, and we describe...... a protocol for expressing Hbs with low intrinsic solubilities. Since the alpha- and beta-chain Hbs of different species span a broad range of solubilities, experimental protocols that have been optimized for expressing recombinant human HbA may often prove unsuitable for the recombinant expression......-translational modifications. CONCLUSION/SIGNIFICANCE: Our protocol should prove useful for the experimental study of recombinant Hbs in many non-human animals. One of the chief advantages of our protocol is that we can express soluble recombinant Hb without co-expressing molecular chaperones, and without the need...

  7. Incidence of Escherichia coli in black walnut meats.

    Science.gov (United States)

    Meyer, M T; Vaughn, R H

    1969-11-01

    Examination of commercially shelled black walnut meats showed inconsistent numbers of total aerobic bacteria, coliforms, and Escherichia coli; variation occurred among different meat sizes and within each meat size. The incidence of E. coli on meats of commercially hulled black walnuts depended on the physical condition of the nuts. Apparently tightly sealed ones contained only a few or none, whereas those with visibly separated sutures and spoiled meats yielded the most. This contamination was in part correlated to a hulling operation. Large numbers of E. coli on the husk of the walnuts contaminated the hulling water, subsequently also contaminating the meats by way of separated sutures. Chlorination of the hulling wash water was ineffective. Attempts were made to decontaminate the walnut meats without subsequent deleterious changes in flavor or texture. A treatment in coconut oil at 100 C followed by removal of excess surface oil by centrifugation was best.

  8. Peptide nucleic acid (PNA) antisense effects in Escherichia coli

    DEFF Research Database (Denmark)

    Good, L; Nielsen, P E

    1999-01-01

    Antisense peptide nucleic acid (PNA) can be used to control cell growth, gene expression and growth phenotypes in the bacteria Escherichia coli. PNAs targeted to the RNA components of the ribosome can inhibit translation and cell growth, and PNAs targeted to mRNA can limit gene expression with gene...... and sequence specificity. In an E. coli cell extract, efficient inhibition is observed when using PNA concentrations in the nanomolar range, whereas micromolar concentrations are required for inhibition in growing cells. A mutant strain of E. coli that is more permeable to antibiotics also is more susceptible...... to antisense PNAs than the wild type. This chapter details methods for testing the antisense activities of PNA in E. coli. As an example of the specific antisense inhibition possible, we show the effects of an anti-beta-galactosidase PNA in comparison to control PNAs. With improvements in cell uptake...

  9. Impact of antibiotic restriction on resistance levels of Escherichia coli

    DEFF Research Database (Denmark)

    Boel, Jonas; Andreasen, Viggo; Jarløv, Jens Otto

    2016-01-01

    as a retrospective controlled interrupted time series (ITS) at two university teaching hospitals, intervention and control, with 736 and 552 beds, respectively. The study period was between January 2008 and September 2014. We used ITS analysis to determine significant changes in antibiotic use and resistance levels......% CI -177, -126)] and fluoroquinolones [-44.5 DDDs/1000 bed-days (95% CI -58.9, -30.1)]. Resistance of E. coli showed a significant change in slope for cefuroxime [-0.13 percentage points/month (95% CI -0.21, -0.057)] and ciprofloxacin [-0.15 percentage points/month (95% CI -0.26, -0.038)]. CONCLUSIONS......OBJECTIVES: We evaluated the effect of an antibiotic stewardship programme (ASP) on the use of antibiotics and resistance levels of Escherichia coli using a method that allowed direct comparison between an intervention hospital and a control hospital. METHODS: The study was conducted...

  10. Analysis of genes involved in glycogen degradation in Escherichia coli.

    Science.gov (United States)

    Strydom, Lindi; Jewell, Jonathan; Meier, Michael A; George, Gavin M; Pfister, Barbara; Zeeman, Samuel; Kossmann, Jens; Lloyd, James R

    2017-02-01

    Escherichia coli accumulate or degrade glycogen depending on environmental carbon supply. Glycogen phosphorylase (GlgP) and glycogen debranching enzyme (GlgX) are known to act on the glycogen polymer, while maltodextrin phosphorylase (MalP) is thought to remove maltodextrins released by GlgX. To examine the roles of these enzymes in more detail, single, double and triple mutants lacking all their activities were produced. GlgX and GlgP were shown to act directly on the glycogen polymer, while MalP most likely catabolised soluble malto-oligosaccharides. Interestingly, analysis of a triple mutant lacking all three enzymes indicates the presence of another enzyme that can release maltodextrins from glycogen. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Detection of Escherichia albertii from chicken meat and giblets.

    Science.gov (United States)

    Maeda, Eriko; Murakami, Koichi; Sera, Nobuyuki; Ito, Kenitiro; Fujimoto, Shuji

    2015-07-01

    Escherichia albertii occasionally causes food-borne outbreaks of gastroenteritis in humans; however, little is known about the vehicle of transmission. To screen retail chicken products for the presence of E. albertii, 104 retail chicken products were investigated. Portions of enrichment cultures that were PCR-positive for E. albertii (n=3) were sub-cultured on agar medium. Only 2 strains obtained from 2 chicken giblet samples were identified as E. albertii by multi locus sequence typing. Antimicrobial susceptibility testing showed that 1 strain was resistant to streptomycin and sulfisoxazole. Both strains harbored the virulence genes cdt and eae. This study is the first description of E. albertii isolation from retail food, suggesting that chicken products are a potential vehicle of E. albertii transmission.

  12. Production of caffeoylmalic acid from glucose in engineered Escherichia coli.

    Science.gov (United States)

    Li, Tianzhen; Zhou, Wei; Bi, Huiping; Zhuang, Yibin; Zhang, Tongcun; Liu, Tao

    2018-07-01

    To achieve biosynthesis of caffeoylmalic acid from glucose in engineered Escherichia coli. We constructed the biosynthetic pathway of caffeoylmalic acid in E. coli by co-expression of heterologous genes RgTAL, HpaBC, At4CL2 and HCT2. To enhance the production of caffeoylmalic acid, we optimized the tyrosine metabolic pathway of E. coli to increase the supply of the substrate caffeic acid. Consequently, an E. coli-E. coli co-culture system was used for the efficient production of caffeoylmalic acid. The final titer of caffeoylmalic acid reached 570.1 mg/L. Microbial production of caffeoylmalic acid using glucose has application potential. In addition, microbial co-culture is an efficient tool for producing caffeic acid esters.

  13. Is Escherichia coli urinary tract infection a zoonosis?

    DEFF Research Database (Denmark)

    Jacobsen, L.; Garneau, P.; Bruant, G.

    2012-01-01

    Recently, it has been suggested that the Escherichia coli causing urinary tract infection (UTI) may come from meat and animals. The purpose was to investigate if a clonal link existed between E. coli from animals, meat and UTI patients. Twenty-two geographically and temporally matched B2 E. coli...... from UTI patients, community-dwelling humans, broiler chicken meat, pork, and broiler chicken, previously identified to exhibit eight virulence genotypes by microarraydetection of approximately 300 genes, were investigated for clonal relatedness by PFGE. Nine isolates were selected and tested...... for in vivo virulence in the mouse model of ascending UTI. UTI and community-dwelling human strains were closely clonally related to meat strains. Several human derived strains were also clonally interrelated. All nine isolates regardless of origin were virulent in the UTI model with positive urine, bladder...

  14. ESBL-Producing Escherichia coli

    DEFF Research Database (Denmark)

    Hertz, Frederik Boetius

    Urinary tract infection (UTI) is one the most common bacterial infections and is regularly treated in primary health care. The most common cause of UTI is extraintestinal pathogenic Escherichia coli (ExPEC) already present in the intestinal microflora, often as the dominating strain. Resistance...... in E.coli is increasing and especially isolates producing Extended-Spectrum Beta-Lactamases (ESBL) have been reported worldwide. Treatment of UTI is usually initiated by the general practitioners and a significant proportion of clinical isolates are now resistant to first line antibiotics. The global...... to investigate (i) antibiotics involved in selection of ESBL-producing E.coli, in an experimental mouse model in vivo, (ii) risk factors for UTI with ESBL-producing E.coli and (iii) to describe the phylogenetic composition of E.coli populations with different resistance patterns. We found that different...

  15. Biocatalytically active silCoat-composites entrapping viable Escherichia coli.

    Science.gov (United States)

    Findeisen, A; Thum, O; Ansorge-Schumacher, M B

    2014-02-01

    Application of whole cells in industrial processes requires high catalytic activity, manageability, and viability under technical conditions, which can in principle be accomplished by appropriate immobilization. Here, we report the identification of carrier material allowing exceptionally efficient adsorptive binding of Escherichia coli whole cells hosting catalytically active carbonyl reductase from Candida parapsilosis (CPCR2). With the immobilizates, composite formation with both hydrophobic and hydrophilized silicone was achieved, yielding advanced silCoat-material and HYsilCoat-material, respectively. HYsilCoat-whole cells were viable preparations with a cell loading up to 400 mg(E. coli) · g(-1)(carrier) and considerably lower leaching than native immobilizates. SilCoat-whole cells performed particularly well in neat substrate exhibiting distinctly increased catalytic activity.

  16. Transport of Escherichia coli phage through saturated porous media considering managed aquifer recharge.

    Science.gov (United States)

    Zhang, Wenjing; Li, Shuo; Wang, Shuang; Lei, Liancheng; Yu, Xipeng; Ma, Tianyi

    2018-03-01

    Virus is one of the most potentially harmful microorganisms in groundwater. In this paper, the effects of hydrodynamic and hydrogeochemical conditions on the transportation of the colloidal virus considering managed aquifer recharge were systematically investigated. Escherichia coli phage, vB_EcoM-ep3, has a broad host range and was able to lyse pathogenic Escherichia coli. Bacteriophage with low risk to infect human has been found extensively in the groundwater environment, so it is considered as a representative model of groundwater viruses. Laboratory studies were carried out to analyze the transport of the Escherichia coli phage under varying conditions of pH, ionic strength, cation valence, flow rate, porous media, and phosphate buffer concentration. The results indicated that decreasing the pH will increase the adsorption of Escherichia coli phage. Increasing the ionic strength, either Na + or Ca 2+ , will form negative condition for the migration of Escherichia coli phage. A comparison of different cation valence tests indicated that changes in transport and deposition were more pronounced with divalent Ca 2+ than monovalent Na + . As the flow rate increases, the release of Escherichia coli phage increases and the retention of Escherichia coli phage in the aquifer medium reduces. Changes in porous media had a significant effect on Escherichia coli phage migration. With increase of phosphate buffer concentration, the suspension stability and migration ability of Escherichia coli phage are both increased. Based on laboratory-scale column experiments, a one-dimensional transport model was established to quantitatively describe the virus transport in saturated porous medium.

  17. GENETIC CONTROL OF RESTRICTION AND MODIFICATION IN ESCHERICHIA COLI1

    Science.gov (United States)

    Boyer, Herbert

    1964-01-01

    Boyer, Herbert (Yale University, New Haven, Conn.). Genetic control of restriction and modification in Escherichia coli. J. Bacteriol. 88:1652–1660. 1964.—Bacterial crosses with K-12 strains of Escherichia coli as Hfr donors (Hfr Hayes, Hfr Cavalli, and Hfr P4X-6) and B/r strains of E. coli as F− recipients were found to differ from crosses between K-12 Hfr donors and K-12 F− recipients in two ways: (i) recombinants (leu, pro, lac, and gal) did not appear at discrete time intervals but did appear simultaneously 30 min after matings were initiated, and (ii) the linkage of unselected markers to selected markers was reduced. Integration of a genetic region linked to the threonine locus of K-12 into the B/r genome resulted in a hybrid which no longer gave anomalous results in conjugation experiments. A similar region of the B strain was introduced into the K-12 strain, which then behaved as a typical B F− recipient. These observations are interpreted as the manifestation of host-controlled modification and restriction on the E. coli chromosome. This was verified by experiments on the restriction and modification of the bacteriophage lambda, F-lac, F-gal, and sex-factor, F1. It was found that the genetic region that controlled the mating responses of the K-12 and B/r strains also controlled the modification and restriction properties of these two strains. The genes responsible for the restricting and modifying properties of the K-12 and B strains of E. coli were found to be allelic, linked to each other, and linked to the threonine locus. PMID:14240953

  18. Photoreactivable sector of lethal damage in ultraviolet-irradiated Escherichia coli cells

    International Nuclear Information System (INIS)

    Balgavy, P.

    1976-01-01

    The photoreactivable sector of lethal damage in Escherichia coli Bsub(s-1), Escherichia coli B/r Hcr - and Escherichia coli B/r Hcr + cells after ultraviolet irradiation at 254 nm is 0.823 +- 0.004, 0.70 +- 0.01 and 0.53 +- 0.06, respectively, at 99% confidence limits. For the low values of the photoreactivable sector in the B/r Hcr - and B/r Hcr + strains are likely to be responsible dark repair processes which eliminate lethal damage, brought about by pyrimidine dimers, preferably in comparison with lethal damage caused by photoproducts of another type. (author)

  19. Resonance energy transfer study on the proximity relationship between the GTP binding site and the rifampicin binding site of Escherichia coli RNA polymerase

    International Nuclear Information System (INIS)

    Kumar, K.P.; Chatterji, D.

    1990-01-01

    Terbium(III) upon complexation with guanosine 5'-triphosphate showed remarkable enhancement of fluorescence emission at 488 and 545 nm when excited at 295 nm. Analysis of the binding data yielded a value for the mean K d between Tb(III) and GTP of 0.2 μM, with three binding sites for TB(III) on GTP. 31 P and 1 H NMR measurements revealed that Tb(III) mainly binds the phosphate moiety of GTP. Fluorescence titration of the emission signals of the TbGTP complex with varying concentrations of Escherichia coli RNA polymerase resulted in a K d values of 4 μM between the TbGTP and the enzyme. It was observed that TbGTP can be incorporated in the place of GTP during E. coli RNA polymerase catalyzed abortive synthesis of dinucleotide tetraphosphate at T7A2 promoter. Both the substrate TbGTP and the inhibitor of the initiation of transcription rifampicin bind to the β-subunit of E. coli RNA polymerase. This allows the measurement of the fluorescence excited-state energy transfer from the donor TbGTP-RNA polymerase to the acceptor rifampicin. Both emission bands of Tb(III) overlap with the rifampicin absorption, and the distances at 50% efficiency of energy transfer were calculated to be 28 and 24 angstrom for the 488- and 545-nm emission bands, respectively. The distance between the substrate binding site and the rifampicin binding site on the β-subunit of E. coli RNA polymerase was measured to be around 30 angstrom. This suggest that the nature of inhibition of transcription by rifampicin is essentially noncompetitive with the substrate

  20. Engineering and Validation of a Vector for Concomitant Expression of Rare Transfer RNA (tRNA and HIV-1 nef Genes in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Siti Aisyah Mualif

    Full Text Available Relative ease in handling and manipulation of Escherichia coli strains make them primary candidate to express proteins heterologously. Overexpression of heterologous genes that contain codons infrequently used by E. coli is related with difficulties such as mRNA instability, early termination of transcription and/or translation, deletions and/or misincorporation, and cell growth inhibition. These codon bias -associated problems are addressed by co-expressing ColE1-compatible, rare tRNA expressing helper plasmids. However, this approach has inadequacies, which we have addressed by engineering an expression vector that concomitantly expresses the heterologous protein of interest, and rare tRNA genes in E. coli. The expression vector contains three (argU, ileY, leuW rare tRNA genes and a useful multiple cloning site for easy in-frame cloning. To maintain the overall size of the parental plasmid vector, the rare tRNA genes replaced the non-essential DNA segments in the vector. The cloned gene is expressed under the control of T7 promoter and resulting recombinant protein has a C-terminal 6His tag for IMAC-mediated purification. We have evaluated the usefulness of this expression vector by expressing three HIV-1 genes namely HIV-1 p27 (nef, HIV-1 p24 (ca, and HIV-1 vif in NiCo21(DE3 E.coli and demonstrated the advantages of using expression vector that concomitantly expresses rare tRNA and heterologous genes.

  1. Isolation, genotyping, and antimicrobial resistance of zoonotic shiga toxin-producing escherichia coli

    Science.gov (United States)

    Shiga toxin-producing Escherichia coli (STEC) is an enteric pathogen linked to outbreaks of human gastroenteritis with diverse clinical spectra. Traditional culture and isolation methods, including selective enrichment and differential plating, have enabled the effective recovery of STEC. Ruminants ...

  2. Effects of substitutions at position 180 in the Escherichia coli RNA ...

    Indian Academy of Sciences (India)

    Escherichia coli RNA polymerase, two mutant variants of the protein with substitutions for either alanine or glutamic .... promoter signals utilized for in vitro transcription assays and ..... free recombinant protein using a self-cleavable affinity tag.

  3. A Canadian Multicentre Case-Control Study of Sporadic Escherichia coli 0157:H7 Infection

    Directory of Open Access Journals (Sweden)

    Donna Holton

    1999-01-01

    Full Text Available OBJECTIVE: To evaluate further risk factors for Escherichia coli 0157:H7 infection including consumer preferences related to the consumption of ground beef and the role of person-to-person transmission of this infection.

  4. COMPARISON OF ESCHERICHIA COLI, TOTAL COLIFORM, AND FECAL COLIFORM POPULATIONS AS INDICATORS OF WASTEWATER TREATMENT EFFICIENCY

    Science.gov (United States)

    Escherichia coli, total coliform, and fecal coliform data were collected from two wastewater treatment facilities, a subsurface constructed wetlands, and the receiving stream. Results are presented from individual wastewater treatment process streams, final effluent and river sit...

  5. Enterococcus and Escherichia coli fecal source apportionment with microbial source tracking genetic markers - is it feasible?

    Science.gov (United States)

    Fecal pollution is measured in surface waters using culture-based measurements of enterococci and Escherichia coli bacteria. Source apportionment of these two fecal indicator bacteria is an urgent need for prioritizing remediation efforts and quantifying health risks associated...

  6. THE WIDESPREAD OCCURRENCE OF THE ENTEROHEMOLYSIN GENE EHLYA AMONG ENVIRONMENTAL STRAINS OF ESCHERICHIA COLI

    Science.gov (United States)

    The putative virulence factor enterohemolysin, encoded for by the ehlyA gene, has been closely associated with the pathogenic enterohemorrhagic Escherichia coli (EHEC) group. E. coli isolates from effluents from seven geographically dispersed municipal ...

  7. Hemagglutinin Typing as an Aid in Identification of Biochemically Atypical Escherichia coli Strains

    OpenAIRE

    Crichton, Pamela B.; Ip, S. M.; Old, D. C.

    1981-01-01

    Tests for the presence of mannose-sensitive and mannose-resistant, eluting hemagglutinins and fimbriae were helpful in indicating whether biochemically atypical strains of the tribe Escherichieae might be escherichiae or shigellae.

  8. Hemagglutinin Typing as an Aid in Identification of Biochemically Atypical Escherichia coli Strains

    Science.gov (United States)

    Crichton, Pamela B.; Ip, S. M.; Old, D. C.

    1981-01-01

    Tests for the presence of mannose-sensitive and mannose-resistant, eluting hemagglutinins and fimbriae were helpful in indicating whether biochemically atypical strains of the tribe Escherichieae might be escherichiae or shigellae. PMID:7334072

  9. Low resolution solution structure of the Apo form of Escherichia coli haemoglobin protease Hbp.

    NARCIS (Netherlands)

    scott, D.J.; Grossman, J.G.; Tame, J.R.H.; Byron, O.; Wilson, K.S.; Otto, B.R.

    2002-01-01

    We have studied the solution properties of the apo form of the haemoglobin protease or "haemoglobinase", Hbp, a principal component of an important iron acquisition system in pathogenic Escherichia coli. Experimental determination of secondary structure content from circular dichroism (CD)

  10. The Escherichia coli antiterminator protein BglG stabilizes the 5 ...

    Indian Academy of Sciences (India)

    Unknown

    Keywords. Antitermination; mRNA stability; RNA binding protein ... factor, Rho, and the pBR322 copy number protein, Rop, have been .... Transcription analysis using the oligo- ..... Retarded RNA turnover in Escherichia coli a means of main-.

  11. Antibacterial activity of Tribulus terrestris methanol extract against clinical isolates of Escherichia coli

    Directory of Open Access Journals (Sweden)

    Batoei Sara

    2016-06-01

    Full Text Available Introduction:Tribulus terrestris L. is traditionally used for treatment of urinary tract infections. Escherichia coli, as the most prominent agent of urinary tract infections, can be sensitive to T. terrestris extract.

  12. Identification of Genes Important for Growth of Asymptomatic Bacteriuria Escherichia coli in Urine

    DEFF Research Database (Denmark)

    Vejborg, Rebecca Munk; de Evgrafov, Mari Cristina Rodriguez; Phan, Minh Duy

    2012-01-01

    Escherichia coli is the most important etiological agent of urinary tract infections (UTIs). Unlike uropathogenic E. coli, which causes symptomatic infections, asymptomatic bacteriuria (ABU) E. coli strains typically lack essential virulence factors and colonize the bladder in the absence...

  13. Characterization of the R7T7 LWR reference glass

    International Nuclear Information System (INIS)

    Pacaud, F.; Fillet, C.; Baudin, G.; Bastien-Thiry, H.

    1990-01-01

    Characterization describes the glass properties by means of standard tests with no attempt to assess its long-term behavior. Characterization involved complementary comparative investigations of nonradioactive laboratory glass specimens, radioactive glass specimens prepared in laboratory hot cells, and nonradioactive industrial glass samples fabricated in the full-scale continuous vitrification prototype facility (specimens were taken from the casting stream and core-samples were taken from a 200 kg glass block after cooling in the canister). Additional measurements are planned on actual radioactive glass samples fabricated in the R7 facility at La Hague. The results are indicated for each of the properties studied: physical, thermal and mechanical properties; structure and homogeneity examination; thermal stability and crystallization; resistance to chemical corrosion; irradiation resistance and volatilization. Comparative examination of glass samples of different origins showed consistent properties

  14. PODAAC-AQR40-3T7CS

    Data.gov (United States)

    National Aeronautics and Space Administration — The version 4.0 Aquarius-CAP Level 3 product contains the third release of the gridded/mapped 7day running mean SSS data from AQUARIUS/SAC-D for the JPL CAP algorithm

  15. Genome Sequences of Two Copper-Resistant Escherichia coli Strains Isolated from Copper-Fed Pigs

    DEFF Research Database (Denmark)

    Lüthje, Freja L.; Hasman, Henrik; Aarestrup, Frank Møller

    2014-01-01

    The draft genome sequences of two copper-resistant Escherichia coli strains were determined. These had been isolated from copper-fed pigs and contained additional putative operons conferring copper and other metal and metalloid resistances.......The draft genome sequences of two copper-resistant Escherichia coli strains were determined. These had been isolated from copper-fed pigs and contained additional putative operons conferring copper and other metal and metalloid resistances....

  16. Escherichia coli: a brief review of diarrheagenic pathotypes and their role in diarrheal diseases in Iran

    Science.gov (United States)

    Jafari, A; Aslani, MM; Bouzari, S

    2012-01-01

    Diarrheagenic Escherichia coli have developed different strategies for establishment of infection in their host. Understanding these pathogenic mechanisms has led to the development of specific diagnostic tools for identification and categorization of E. coli strains into different pathotypes. This review aims to provide an overview of the various categories of diarrheagenic Escherichia coli and the data obtained in Iran pertaining to these pathotypes. PMID:23066484

  17. Purification and Characterization of Recombinant Vaccinia L1R Protein from Escherichia coli

    Science.gov (United States)

    2016-08-01

    RECOMBINANT VACCINIA L1R PROTEIN FROM ESCHERICHIA COLI 1. INTRODUCTION 1.1 Background Vaccinia virus (VACV) is the active component of the...the preparation of the recombinant VACV L1R protein fragment by denaturing , refolding, and purifying material expressed into inclusion bodies in...PURIFICATION AND CHARACTERIZATION OF RECOMBINANT VACCINIA L1R PROTEIN FROM ESCHERICHIA COLI ECBC-TR-1370

  18. Heterologous expression of enterocin A, a bacteriocin from Enterococcus faecium, fused to a cellulose-binding domain in Escherichia coli results in a functional protein with inhibitory activity against Listeria.

    Science.gov (United States)

    Klocke, Michael; Mundt, Kerstin; Idler, Frank; Jung, Sabrina; Backhausen, Jan E

    2005-06-01

    The genes for the bacteriocins enterocin A and B were isolated from Enterococcus faecium ATB 197a. Using the pET37b(+) vector, the enterocin genes were fused to an Escherichia coli specific export signal sequence, a cellulose-binding domain (CBD(cenA)) and a S-tag under the control of a T7lac promotor. The constructs were subsequently cloned into E. coli host cells. The expression of the recombinant enterocins had different effects on both the host cells and other Gram-positive bacteria. The expression of entA in Esc. coli led to the synthesis and secretion of functional active enterocin A fusion proteins, which were active against some Gram-positive indicator bacteria, but did not influence the viability of the host cells. In contrast, the expression of enterocin B fusion proteins led to a reduced viability of the host cells, indicating a misfolding of the protein or interference with the cellular metabolism of Esc. coli. Indicator strains of Gram-positive bacteria were not inhibited by purified enterocin B fusion proteins. However, recombinant enterocin B displayed inhibitory activity after the proteolytic cleavage of the fused peptides.

  19. Immobilizing live Escherichia coli for AFM studies of surface dynamics

    International Nuclear Information System (INIS)

    Lonergan, N.E.; Britt, L.D.; Sullivan, C.J.

    2014-01-01

    Atomic force microscopy (AFM) is a probe-based technique that permits high resolution imaging of live bacterial cells. However, stably immobilizing cells to withstand the probe-based lateral forces remains an obstacle in AFM mediated studies, especially those of live, rod shaped bacteria in nutrient media. Consequently, AFM has been under-utilized in the research of bacterial surface dynamics. The aim of the current study was to immobilize a less adherent Escherichia coli strain in a method that both facilitates AFM imaging in nutrient broth and preserves overall cell viability. Immobilization reagents and buffers were systematically evaluated and the cell membrane integrity was monitored in all sample preparations. As expected, the biocompatible gelatin coated surfaces facilitated stable cell attachment in lower ionic strength buffers, yet poorly immobilized cells in higher ionic strength buffers. In comparison, poly-L-lysine surfaces bound cells in both low and high ionic strength buffers. The benefit of the poly-L-lysine binding capacity was offset by the compromised membrane integrity exhibited by cells on poly-L-lysine surfaces. However, the addition of divalent cations and glucose to the immobilization buffer was found to mitigate this unfavorable effect. Ultimately, immobilization of E. coli cells on poly-L-lysine surfaces in a lower ionic strength buffer supplemented with Mg 2+ and Ca 2+ was determined to provide optimal cell attachment without compromising the overall cell viability. Cells immobilized in this method were stably imaged in media through multiple division cycles. Furthermore, permeability assays indicated that E. coli cells recover from the hypoosmotic stress caused by immobilization in low ionic strength buffers. Taken together, this data suggests that stable immobilization of viable cells on poly-L-lysine surfaces can be accomplished in lower ionic strength buffers that are supplemented with divalent cations for membrane stabilization while

  20. High mutation rates limit evolutionary adaptation in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Kathleen Sprouffske

    2018-04-01

    Full Text Available Mutation is fundamental to evolution, because it generates the genetic variation on which selection can act. In nature, genetic changes often increase the mutation rate in systems that range from viruses and bacteria to human tumors. Such an increase promotes the accumulation of frequent deleterious or neutral alleles, but it can also increase the chances that a population acquires rare beneficial alleles. Here, we study how up to 100-fold increases in Escherichia coli's genomic mutation rate affect adaptive evolution. To do so, we evolved multiple replicate populations of asexual E. coli strains engineered to have four different mutation rates for 3000 generations in the laboratory. We measured the ability of evolved populations to grow in their original environment and in more than 90 novel chemical environments. In addition, we subjected the populations to whole genome population sequencing. Although populations with higher mutation rates accumulated greater genetic diversity, this diversity conveyed benefits only for modestly increased mutation rates, where populations adapted faster and also thrived better than their ancestors in some novel environments. In contrast, some populations at the highest mutation rates showed reduced adaptation during evolution, and failed to thrive in all of the 90 alternative environments. In addition, they experienced a dramatic decrease in mutation rate. Our work demonstrates that the mutation rate changes the global balance between deleterious and beneficial mutational effects on fitness. In contrast to most theoretical models, our experiments suggest that this tipping point already occurs at the modest mutation rates that are found in the wild.

  1. Improvements In Ethanologenic Escherichia Coli and Klebsiella Oxytoca

    Energy Technology Data Exchange (ETDEWEB)

    Dr. David Nunn

    2010-09-30

    The current Verenium cellulosic ethanol process is based on the dilute-acid pretreatment of a biomass feedstock, followed by a two-stage fermentation of the pentose sugar-containing hydrolysate by a genetically modified ethanologenic Escherichia coli strain and a separate simultaneous saccharification-fermentation (SSF) of the cellulosic fraction by a genetically modified ethanologenic Klebsiella oxytoca strain and a fungal enzyme cocktail. In order to reduce unit operations and produce a fermentation beer with higher ethanol concentrations to reduce distillation costs, we have proposed to develop a simultaneous saccharification co-fermentation (SScF) process, where the fermentation of the pentose-containing hydrolysate and cellulosic fraction occurs within the same fermentation vessel. In order to accomplish this goal, improvements in the ethanologens must be made to address a number of issues that arise, including improved hydrolysate tolerance, co-fermentation of the pentose and hexose sugars and increased ethanol tolerance. Using a variety of approaches, including transcriptomics, strain adaptation, metagenomics and directed evolution, this work describes the efforts of a team of scientists from Verenium, University of Florida, Massachusetts Institute of Technology and Genomatica to improve the E. coli and K. oxytoca ethanologens to meet these requirements.

  2. Attachment of Escherichia coli and enterococci to particles in runoff.

    Science.gov (United States)

    Soupir, Michelle L; Mostaghimi, Saied; Dillaha, Theo

    2010-01-01

    Association of Escherichia coli and enterococci with particulates present in runoff from erodible soils has important implications for modeling the fate and transport of bacteria from agricultural sources and in the selection of management practices to reduce bacterial movement to surface waters. Three soils with different textures were collected from the Ap horizon (silty loam, silty clay loam, and loamy fine sand), placed in portable box plots, treated with standard cowpats, and placed under a rainfall simulator. Rainfall was applied to the plots until saturation-excess flow occurred for 30 min, and samples were collected 10, 20, and 30 min after initiation of the runoff event. The attachment of E. coli and enterococci to particles present in runoff was determined by a screen filtration and centrifugation procedure. Percentage of E. coli and enterococci attached to particulates in runoff ranged from 28 to 49%, with few statistically significant differences in attachment among the three soils. Similar partitioning release patterns were observed between E. coli and enterococci from the silty loam (r = 0.57) and silty clay loam soils (r = 0.60). At least 60% of all attached E. coli and enterococci were associated particles within an 8- to 62-microm particle size category. The results indicate that the majority of fecal bacteria attach to and are transported with manure colloids in sediment-laden flow regardless of the soil texture.

  3. Enzyme organization in the proline biosynthetic pathway of Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Gamper, H; Moses, V

    1974-01-01

    The conversion of glutamic acid to proline by an Escherichia coli extract was studied. The activity was dependent upon the presence of ATP and NADPH and was largely unaffected by the presence of NH/sub 3/ or imidazole. The first two pathway enzymes appear to exist as a complex which stabilizes a labile intermediate postulated as ..gamma..-glutamyl phosphate. Attempted synthesis of this compound was unsuccessful due to its spontaneous cyclization to 2-pyrrolidone 5-carboxylate. Dissociation of the enzyme complex upon dilution of the extract is presumed responsible for an experimentally observed dilution effect. E. coli pro/sub A//sup -/ and pro/sub B//sup -/ auxotroph extracts failed to complement one another in the biosynthesis of proline. This is attributed to the lack of a dynamic equilibrium between the complex and its constituent enzymes. In vivo studies with E. coli showed no evidence for metabolic channeling in the final reaction of proline synthesis, the reduction of ..delta../sup 1/-pyrroline 5-carboxylate.

  4. Incidence of Escherichia coli O157:H7 in Thailand

    International Nuclear Information System (INIS)

    Sukhumungoon, P.

    2015-01-01

    Entero hemorrhagic Escherichia coli (EHEC) especially serotype O157:H7 is one of the important food-borne pathogens because it is able to produce crucial toxins Shiga. However, the outbreak of this organism in Thailand has not been reported. Antibody to O157 antigen was detected in some Thai populations and Shiga toxin-producing E. coli were detected in low numbers of clinical specimens. Interestingly, some E. coli that showed positive to O157 fimbriae probe and lack of virulence gene were isolated from certain patients and one isolate of E. coli O157:H7 which possessed stx1, stx2v was detected in a normal child. In addition, the incidence of E. coli O157:H7 strains were monitored by the samples from cattle and retail beef in Thailand although their inability to produce toxins or produce in a low concentration was demonstrated. This review discusses the incidences of E. coli O157 in clinical and environmental samples of Thailand including the transmission possibility of this bacterium across the Thai border through food trade. (author)

  5. Oxidative Stress in Shiga Toxin Production by Enterohemorrhagic Escherichia coli

    Directory of Open Access Journals (Sweden)

    Katarzyna Licznerska

    2016-01-01

    Full Text Available Virulence of enterohemorrhagic Escherichia coli (EHEC strains depends on production of Shiga toxins. These toxins are encoded in genomes of lambdoid bacteriophages (Shiga toxin-converting phages, present in EHEC cells as prophages. The genes coding for Shiga toxins are silent in lysogenic bacteria, and prophage induction is necessary for their efficient expression and toxin production. Under laboratory conditions, treatment with UV light or antibiotics interfering with DNA replication are commonly used to induce lambdoid prophages. Since such conditions are unlikely to occur in human intestine, various research groups searched for other factors or agents that might induce Shiga toxin-converting prophages. Among other conditions, it was reported that treatment with H2O2 caused induction of these prophages, though with efficiency significantly lower relative to UV-irradiation or mitomycin C treatment. A molecular mechanism of this phenomenon has been proposed. It appears that the oxidative stress represents natural conditions provoking induction of Shiga toxin-converting prophages as a consequence of H2O2 excretion by either neutrophils in infected humans or protist predators outside human body. Finally, the recently proposed biological role of Shiga toxin production is described in this paper, and the “bacterial altruism” and “Trojan Horse” hypotheses, which are connected to the oxidative stress, are discussed.

  6. On the control of ribosomal protein biosynthesis in Escherichia coli

    International Nuclear Information System (INIS)

    Pichon, J.; Marvaldi, J.; Coeroli, C.; Cozzone, A.; Marchis-Mouren, G.

    1977-01-01

    The rate of individual ribosomal protein synthesis relative to total protein synthesis has been determined in Escherichia coli rel + and rel - cells, under valyl-tRNA deprivation. These strains have a temperature-sensitive valyl-tRNA synthetase. Starvation was obtained following transfer of the cells to non-permissive temperature. Ribosomal proteins were obtained by treatment of either total lysates of freeze-thawed lysozyme spheroplasts or ammonium sulphate precipitate of ribosomes, with acetic acid. Differential labelling of the ribosomal proteins was observed in both strains: proteins from the rel + strain appear more labelled than those from the rel - strain, the rate of labelling of individual proteins being about the same in both strains. Moreover ribosomal proteins were found as stable during starvation as total protein. It is thus concluded that in starving cells individual ribosomal proteins are not synthesized at equal rates. This indicates that the synthesis of ribosomal proteins is not only under the control of the rel gene

  7. Response of Escherichia coli to Prolonged Berberine Exposure.

    Science.gov (United States)

    Budeyri Gokgoz, Nilay; Avci, Fatma Gizem; Yoneten, Kubra Karaosmanoglu; Alaybeyoglu, Begum; Ozkirimli, Elif; Sayar, Nihat Alpagu; Kazan, Dilek; Sariyar Akbulut, Berna

    2017-07-01

    Berberine is a plant-derived alkaloid possessing antimicrobial activity; unfortunately, its efflux through multidrug resistance pumps reduces its efficacy. Cellular life span of Escherichia coli is generally shorter with prolonged berberine exposure; nevertheless, about 30% of the cells still remain robust following this treatment. To elucidate its mechanism of action and to identify proteins that could be involved in development of antimicrobial resistance, protein profiles of E. coli cells treated with berberine for 4.5 and 8 hours were compared with control cells. A total of 42 proteins were differentially expressed in cells treated with berberine for 8 hours when compared to control cells. In both 4.5 and 8 hours of berberine-treated cells, carbohydrate and peptide uptake regimens remained unchanged, although amino acid maintenance regimen switched from transport to synthesis. Defect in cell division persisted and this condition was confirmed by images obtained from scanning electron microscopy. Universal stress proteins were not involved in stress response. The significant increase in the abundance of elongation factors could suggest the involvement of these proteins in protection by exhibiting chaperone activities. Furthermore, the involvement of the outer membrane protein OmpW could receive special attention as a protein involved in response to antimicrobial agents, since the expression of only this porin protein was upregulated after 8 hours of exposure.

  8. Overexpression of SOS genes in ciprofloxacin resistant Escherichia coli mutants.

    Science.gov (United States)

    Pourahmad Jaktaji, Razieh; Pasand, Shirin

    2016-01-15

    Fluoroquinolones are important antibiotics for the treatment of urinary tract infections caused by Escherichia coli. Mutational studies have shown that ciprofloxacin, a member of fluoroquinolones induces SOS response and mutagenesis in pathogenic bacteria which in turn develop antibiotic resistance. However, inhibition of SOS response can increase recombination activity which in turn leads to genetic variation. The aim of this study was to measure 5 SOS genes expressions in nine E. coli mutants with different MICs for ciprofloxacin following exposure to ciprofloxacin. Gene expression was assessed by quantitative real time PCR. Gene alteration assessment was conducted by PCR amplification and DNA sequencing. Results showed that the expression of recA was increased in 5 mutants. This overexpression is not related to gene alteration, and enhances the expression of polB and umuCD genes encoding nonmutagenic and mutagenic polymerases, respectively. The direct relationship between the level of SOS expression and the level of resistance to ciprofloxacin was also indicated. It was concluded that novel therapeutic strategy that inhibits RecA activity would enhance the efficiency of common antibiotics against pathogenic bacteria. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. CHARACTERIZATION AND ANTIBIOGRAM OF ENTEROPATHOGENIC ESCHERICHIA COLI ISOLATED FROM POULTRY

    Directory of Open Access Journals (Sweden)

    M. Sarkar

    2013-12-01

    Full Text Available One hundred sixty two samples from different poultry farms of West Bengal, India were screened for the presence of pathogenic Escherichia coli and 109 (67.3% were found positive. Out of forty six faecal samples from ailing birds suffering from acute colibacillosis, thirty one i.e. 67.2% were positive whereas postmortem sample of intestines (62 and liver tissues (54 revealed approx 72.6% and 61.1% positivity for E. coli. Biochemical characteristic of the isolates were positive to indole, MR, nitrate and non-reactive to VP, citrate & urease test. In serotyping of the E. coli isolates mostly revealed O2, O8, O9, O19, O37, O47, O55, O69, O86, O101, O103, O109, O133, O151 and O173. The serotypes viz. O2, O8, O9, O55, O101 and O133 showed acute pathogenicity in swiss mice followed by O19, O37, O47, O69, O86, O103, O109, O151 and O173 as moderately pathogenic serotypes. Among the antimicrobial drugs tested, the sensitive drugs were cefixime (93.6%, enrofloxacin (91.8%, nitrofurantoin (88.1% and azithromycin (85.3%. The resistant drugs were tetracycline (100%, nalidixic acid (97.2%, metronidazole (92.6%, penicillin G (88.9%, gatifloxacin (77.9% and bacitracin (76.2% .

  10. Mutagenic DNA repair in Escherichia coli. Pt. 2

    International Nuclear Information System (INIS)

    Doubleday, O.P.; Bridges, B.A.; Green, M.H.L.

    1975-01-01

    The photoreversibility of UV-induced mutations to Trp + in strain Escherichia coli WP2 uvr A trp (unable to excise pyrimidine dimers) was lost at different rates during incubation in different media. In Casamino acids medium after a short initial lag, photoreversibility was lost over about one generation time; in minimal medium with tryptophan, photoreversibility persisted for more than two generations; in Casamino acids medium with pantoyl lactone photoreversibility was lost extremely slowly. The rate of loss of photoreversibility was unaffected by UV dose in either Casamino acids medium or in minimal medium. The same eventual number of induced mutants was obtained when cells were incubated for two generations in any of the three media before being transferred to selective plates supplemented with Casamino acids. Thus in each the proportion of cells capable of giving rise to a mutant was the same and only the rate at which these cells did so during post-irradiation growth varied, suggesting that there might be a specific fraction of pyrimidine dimers at a given site capable of initiating a mutagenic repair event, and that the size of this fraction is dose dependent. Segregation experiments have shown that error-prone repair appears to occur once only and is not repeated in subsequent replication cycles, in contrast to (presumed error-free) recombination repair. The results are discussed in the light of current models of UV mutagenesis. (orig.) [de

  11. Ecological and genetic determinants of plasmid distribution in Escherichia coli.

    Science.gov (United States)

    Medaney, Frances; Ellis, Richard J; Raymond, Ben

    2016-11-01

    Bacterial plasmids are important carriers of virulence and antibiotic resistance genes. Nevertheless, little is known of the determinants of plasmid distribution in bacterial populations. Here the factors affecting the diversity and distribution of the large plasmids of Escherichia coli were explored in cattle grazing on semi-natural grassland, a set of populations with low frequencies of antibiotic resistance genes. Critically, the population genetic structure of bacterial hosts was chararacterized. This revealed structured E. coli populations with high diversity between sites and individuals but low diversity within cattle hosts. Plasmid profiles, however, varied considerably within the same E. coli genotype. Both ecological and genetic factors affected plasmid distribution: plasmid profiles were affected by site, E. coli diversity, E. coli genotype and the presence of other large plasmids. Notably 3/26 E. coli serotypes accounted for half the observed plasmid-free isolates indicating that within species variation can substantially affect carriage of the major conjugative plasmids. The observed population structure suggest that most of the opportunities for within species plasmid transfer occur between different individuals of the same genotype and support recent experimental work indicating that plasmid-host coevolution, and epistatic interactions on fitness costs are likely to be important in determining occupancy. © 2016 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

  12. Endonuclease IV of Escherichia coli is induced by paraquat

    International Nuclear Information System (INIS)

    Chan, E.; Weiss, B.

    1987-01-01

    The addition of paraquat (methyl viologen) to a growing culture of Escherichia coli K-12 led within 1 hr to a 10- to 20-fold increase in the level of endonuclease IV, a DNase for apurinic/apyrimidinic sites. The induction was blocked by chloramphenicol. Increases of 3-fold or more were also seen with plumbagin, menadione, and phenazine methosulfate. H 2 O 2 produced no more than a 2-fold increase in endonuclease IV activity. The following agents had no significant effect: streptonigrin, nitrofurantoin, tert-butyl hydroperoxide, γ rays, 260-nm UV radiation, methyl methanesulfonate, mitomycin C, and ascorbate. Paraquat, plumbagin, menadione, and phenazine methosulfate are known to generate superoxide radical anions via redox cycling in vivo. A mutant lacking superoxide dismutase was unusually sensitive to induction by paraquat. In addition, endonuclease IV could be induced by merely growing the mutant in pure O 2 . The levels of endonuclease IV in uninduced or paraquat-treated cells were unaffected by mutations of oxyR, a H 2 O 2 -inducible gene that governs an oxidative-stress regulon. The results indicate that endonuclease IV is an inducible DNA-repair enzyme and that its induction can be mediated via the production of superoxide radicals

  13. Endonuclease IV of Escherichia coli is induced by paraquat

    Energy Technology Data Exchange (ETDEWEB)

    Chan, E.; Weiss, B.

    1987-05-01

    The addition of paraquat (methyl viologen) to a growing culture of Escherichia coli K-12 led within 1 hr to a 10- to 20-fold increase in the level of endonuclease IV, a DNase for apurinic/apyrimidinic sites. The induction was blocked by chloramphenicol. Increases of 3-fold or more were also seen with plumbagin, menadione, and phenazine methosulfate. H/sub 2/O/sub 2/ produced no more than a 2-fold increase in endonuclease IV activity. The following agents had no significant effect: streptonigrin, nitrofurantoin, tert-butyl hydroperoxide, ..gamma.. rays, 260-nm UV radiation, methyl methanesulfonate, mitomycin C, and ascorbate. Paraquat, plumbagin, menadione, and phenazine methosulfate are known to generate superoxide radical anions via redox cycling in vivo. A mutant lacking superoxide dismutase was unusually sensitive to induction by paraquat. In addition, endonuclease IV could be induced by merely growing the mutant in pure O/sub 2/. The levels of endonuclease IV in uninduced or paraquat-treated cells were unaffected by mutations of oxyR, a H/sub 2/O/sub 2/-inducible gene that governs an oxidative-stress regulon. The results indicate that endonuclease IV is an inducible DNA-repair enzyme and that its induction can be mediated via the production of superoxide radicals.

  14. Adenosine diphosphate sugar pyrophosphatase prevents glycogen biosynthesis in Escherichia coli

    Science.gov (United States)

    Moreno-Bruna, Beatriz; Baroja-Fernández, Edurne; Muñoz, Francisco José; Bastarrica-Berasategui, Ainara; Zandueta-Criado, Aitor; Rodríguez-López, Milagros; Lasa, Iñigo; Akazawa, Takashi; Pozueta-Romero, Javier

    2001-01-01

    An adenosine diphosphate sugar pyrophosphatase (ASPPase, EC 3.6.1.21) has been characterized by using Escherichia coli. This enzyme, whose activities in the cell are inversely correlated with the intracellular glycogen content and the glucose concentration in the culture medium, hydrolyzes ADP-glucose, the precursor molecule of glycogen biosynthesis. ASPPase was purified to apparent homogeneity (over 3,000-fold), and sequence analyses revealed that it is a member of the ubiquitously distributed group of nucleotide pyrophosphatases designated as “nudix” hydrolases. Insertional mutagenesis experiments leading to the inactivation of the ASPPase encoding gene, aspP, produced cells with marginally low enzymatic activities and higher glycogen content than wild-type bacteria. aspP was cloned into an expression vector and introduced into E. coli. Transformed cells were shown to contain a dramatically reduced amount of glycogen, as compared with the untransformed bacteria. No pleiotropic changes in the bacterial growth occurred in both the aspP-overexpressing and aspP-deficient strains. The overall results pinpoint the reaction catalyzed by ASPPase as a potential step of regulating glycogen biosynthesis in E. coli. PMID:11416161

  15. Characterization of the YdeO regulon in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Yuki Yamanaka

    Full Text Available Enterobacteria are able to survive under stressful conditions within animals, such as acidic conditions in the stomach, bile salts during transfer to the intestine and anaerobic conditions within the intestine. The glutamate-dependent (GAD system plays a major role in acid resistance in Escherichia coli, and expression of the GAD system is controlled by the regulatory cascade consisting of EvgAS > YdeO > GadE. To understand the YdeO regulon in vivo, we used ChIP-chip to interrogate the E. coli genome for candidate YdeO binding sites. All of the seven operons identified by ChIP-chip as being potentially regulated by YdeO were confirmed as being under the direct control of YdeO using RT-qPCR, EMSA, DNaseI-footprinting and reporter assays. Within this YdeO regulon, we identified four stress-response transcription factors, DctR, NhaR, GadE, and GadW and enzymes for anaerobic respiration. Both GadE and GadW are involved in regulation of the GAD system and NhaR is an activator for the sodium/proton antiporter gene. In conjunction with co-transcribed Slp, DctR is involved in protection against metabolic endoproducts under acidic conditions. Taken all together, we suggest that YdeO is a key regulator of E. coli survival in both acidic and anaerobic conditions.

  16. Mechanosensing regulates virulence in Escherichia coli O157:H7.

    Science.gov (United States)

    Islam, Md Shahidul; Krachler, Anne Marie

    2016-01-01

    Enterohemorrhagic Escherichia coli O157:H7 is a food-borne pathogen transmitted via the fecal-oral route, and can cause bloody diarrhea and hemolytic uremic syndrome (HUS) in the human host. Although a range of colonization factors, Shiga toxins and a type III secretion system (T3SS) all contribute to disease development, the locus of enterocyte effacement (LEE) encoded T3SS is responsible for the formation of lesions in the intestinal tract. While a variety of chemical cues in the host environment are known to up-regulate LEE expression, we recently demonstrated that changes in physical forces at the site of attachment are required for localized, full induction of the system and thus spatial regulation of virulence in the intestinal tract. Here, we discuss our findings in the light of other recent studies describing mechanosensing of the host and force-dependent induction of virulence mechanisms. We discuss potential mechanisms of mechanosensing and mechanotransduction, and the level of conservation across bacterial species.

  17. Escherichia coli PII protein: purification, crystallization and oligomeric structure.

    Science.gov (United States)

    Vasudevan, S G; Gedye, C; Dixon, N E; Cheah, E; Carr, P D; Suffolk, P M; Jeffrey, P D; Ollis, D L

    1994-01-17

    The Escherichia coli signal transduction protein PII, product of the glnB gene, was overproduced and purified. The predicted molecular weight of the protein based on the correct nucleotide sequence is 12,427 and is very close to the value 12,435 obtained by matrix-assisted laser desorption mass spectrometry. Hexagonal crystals of the unuridylylated form of PII with dimensions 0.2 x 0.2 x 0.3 mm were grown and analysed by X-ray diffraction. The crystals belong to space group P6(3) with a = b = 61.6 A, c = 56.3 A and Vm of 2.5 for one subunit in the asymmetric unit. A low-resolution electron density map showed electron density concentrated around a three-fold axis, suggesting the molecule to be a trimer. A sedimentation equilibrium experiment of the meniscus depletion type was used to estimate a molecular weight of 35,000 +/- 1,000 for PII in solution. This result is consistent with the native protein being a homotrimer.

  18. Global functional atlas of Escherichia coli encompassing previously uncharacterized proteins.

    Directory of Open Access Journals (Sweden)

    Pingzhao Hu

    2009-04-01

    Full Text Available One-third of the 4,225 protein-coding genes of Escherichia coli K-12 remain functionally unannotated (orphans. Many map to distant clades such as Archaea, suggesting involvement in basic prokaryotic traits, whereas others appear restricted to E. coli, including pathogenic strains. To elucidate the orphans' biological roles, we performed an extensive proteomic survey using affinity-tagged E. coli strains and generated comprehensive genomic context inferences to derive a high-confidence compendium for virtually the entire proteome consisting of 5,993 putative physical interactions and 74,776 putative functional associations, most of which are novel. Clustering of the respective probabilistic networks revealed putative orphan membership in discrete multiprotein complexes and functional modules together with annotated gene products, whereas a machine-learning strategy based on network integration implicated the orphans in specific biological processes. We provide additional experimental evidence supporting orphan participation in protein synthesis, amino acid metabolism, biofilm formation, motility, and assembly of the bacterial cell envelope. This resource provides a "systems-wide" functional blueprint of a model microbe, with insights into the biological and evolutionary significance of previously uncharacterized proteins.

  19. Global functional atlas of Escherichia coli encompassing previously uncharacterized proteins.

    Science.gov (United States)

    Hu, Pingzhao; Janga, Sarath Chandra; Babu, Mohan; Díaz-Mejía, J Javier; Butland, Gareth; Yang, Wenhong; Pogoutse, Oxana; Guo, Xinghua; Phanse, Sadhna; Wong, Peter; Chandran, Shamanta; Christopoulos, Constantine; Nazarians-Armavil, Anaies; Nasseri, Negin Karimi; Musso, Gabriel; Ali, Mehrab; Nazemof, Nazila; Eroukova, Veronika; Golshani, Ashkan; Paccanaro, Alberto; Greenblatt, Jack F; Moreno-Hagelsieb, Gabriel; Emili, Andrew

    2009-04-28

    One-third of the 4,225 protein-coding genes of Escherichia coli K-12 remain functionally unannotated (orphans). Many map to distant clades such as Archaea, suggesting involvement in basic prokaryotic traits, whereas others appear restricted to E. coli, including pathogenic strains. To elucidate the orphans' biological roles, we performed an extensive proteomic survey using affinity-tagged E. coli strains and generated comprehensive genomic context inferences to derive a high-confidence compendium for virtually the entire proteome consisting of 5,993 putative physical interactions and 74,776 putative functional associations, most of which are novel. Clustering of the respective probabilistic networks revealed putative orphan membership in discrete multiprotein complexes and functional modules together with annotated gene products, whereas a machine-learning strategy based on network integration implicated the orphans in specific biological processes. We provide additional experimental evidence supporting orphan participation in protein synthesis, amino acid metabolism, biofilm formation, motility, and assembly of the bacterial cell envelope. This resource provides a "systems-wide" functional blueprint of a model microbe, with insights into the biological and evolutionary significance of previously uncharacterized proteins.

  20. Escherichia coli ST131, an Intriguing Clonal Group

    Science.gov (United States)

    Bertrand, Xavier; Madec, Jean-Yves

    2014-01-01

    SUMMARY In 2008, a previously unknown Escherichia coli clonal group, sequence type 131 (ST131), was identified on three continents. Today, ST131 is the predominant E. coli lineage among extraintestinal pathogenic E. coli (ExPEC) isolates worldwide. Retrospective studies have suggested that it may originally have risen to prominence as early as 2003. Unlike other classical group B2 ExPEC isolates, ST131 isolates are commonly reported to produce extended-spectrum β-lactamases, such as CTX-M-15, and almost all are resistant to fluoroquinolones. Moreover, ST131 E. coli isolates are considered to be truly pathogenic, due to the spectrum of infections they cause in both community and hospital settings and the large number of virulence-associated genes they contain. ST131 isolates therefore seem to contradict the widely held view that high levels of antimicrobial resistance are necessarily associated with a fitness cost leading to a decrease in pathogenesis. Six years after the first description of E. coli ST131, this review outlines the principal traits of ST131 clonal group isolates, based on the growing body of published data, and highlights what is currently known and what we need to find out to provide public health authorities with better information to help combat ST131. PMID:24982321

  1. Recent Advances in Understanding Enteric Pathogenic Escherichia coli

    Science.gov (United States)

    Croxen, Matthew A.; Law, Robyn J.; Scholz, Roland; Keeney, Kristie M.; Wlodarska, Marta

    2013-01-01

    SUMMARY Although Escherichia coli can be an innocuous resident of the gastrointestinal tract, it also has the pathogenic capacity to cause significant diarrheal and extraintestinal diseases. Pathogenic variants of E. coli (pathovars or pathotypes) cause much morbidity and mortality worldwide. Consequently, pathogenic E. coli is widely studied in humans, animals, food, and the environment. While there are many common features that these pathotypes employ to colonize the intestinal mucosa and cause disease, the course, onset, and complications vary significantly. Outbreaks are common in developed and developing countries, and they sometimes have fatal consequences. Many of these pathotypes are a major public health concern as they have low infectious doses and are transmitted through ubiquitous mediums, including food and water. The seriousness of pathogenic E. coli is exemplified by dedicated national and international surveillance programs that monitor and track outbreaks; unfortunately, this surveillance is often lacking in developing countries. While not all pathotypes carry the same public health profile, they all carry an enormous potential to cause disease and continue to present challenges to human health. This comprehensive review highlights recent advances in our understanding of the intestinal pathotypes of E. coli. PMID:24092857

  2. Adherent-invasive Escherichia coli in inflammatory bowel disease.

    Science.gov (United States)

    Palmela, Carolina; Chevarin, Caroline; Xu, Zhilu; Torres, Joana; Sevrin, Gwladys; Hirten, Robert; Barnich, Nicolas; Ng, Siew C; Colombel, Jean-Frederic

    2018-03-01

    Intestinal microbiome dysbiosis has been consistently described in patients with IBD. In the last decades, Escherichia coli , and the adherent-invasive E coli (AIEC) pathotype in particular, has been implicated in the pathogenesis of IBD. Since the discovery of AIEC, two decades ago, progress has been made in unravelling these bacteria characteristics and its interaction with the gut immune system. The mechanisms of adhesion of AIEC to intestinal epithelial cells (via FimH and cell adhesion molecule 6) and its ability to escape autophagy when inside macrophages are reviewed here. We also explore the existing data on the prevalence of AIEC in patients with Crohn's disease and UC, and the association between the presence of AIEC and disease location, activity and postoperative recurrence. Finally, we highlight potential therapeutic strategies targeting AIEC colonisation of gut mucosa, including the use of phage therapy, bacteriocins and antiadhesive molecules. These strategies may open new avenues for the prevention and treatment of IBD in the future. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  3. Fast, multiphase volume adaptation to hyperosmotic shock by Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Teuta Pilizota

    Full Text Available All living cells employ an array of different mechanisms to help them survive changes in extra cellular osmotic pressure. The difference in the concentration of chemicals in a bacterium's cytoplasm and the external environment generates an osmotic pressure that inflates the cell. It is thought that the bacterium Escherichia coli use a number of interconnected systems to adapt to changes in external pressure, allowing them to maintain turgor and live in surroundings that range more than two-hundred-fold in external osmolality. Here, we use fluorescence imaging to make the first measurements of cell volume changes over time during hyperosmotic shock and subsequent adaptation on a single cell level in vivo with a time resolution on the order of seconds. We directly observe two previously unseen phases of the cytoplasmic water efflux upon hyperosmotic shock. Furthermore, we monitor cell volume changes during the post-shock recovery and observe a two-phase response that depends on the shock magnitude. The initial phase of recovery is fast, on the order of 15-20 min and shows little cell-to-cell variation. For large sucrose shocks, a secondary phase that lasts several hours adds to the recovery. We find that cells are able to recover fully from shocks as high as 1 Osmol/kg using existing systems, but that for larger shocks, protein synthesis is required for full recovery.

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

    International Nuclear Information System (INIS)

    Amaral, L.; Lee, Y.; Schwarz, U.; Lorian, V.

    1986-01-01

    The binding of 35 S-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

  5. Expanded flux variability analysis on metabolic network of Escherichia coli

    Institute of Scientific and Technical Information of China (English)

    CHEN Tong; XIE ZhengWei; OUYANG Qi

    2009-01-01

    Flux balance analysis,based on the mass conservation law in a cellular organism,has been extensively employed to study the interplay between structures and functions of cellular metabolic networks.Consequently,the phenotypes of the metabolism can be well elucidated.In this paper,we introduce the Expanded Flux Variability Analysis (EFVA) to characterize the intrinsic nature of metabolic reactions,such as flexibility,modularity and essentiality,by exploring the trend of the range,the maximum and the minimum flux of reactions.We took the metabolic network of Escherichia coli as an example and analyzed the variability of reaction fluxes under different growth rate constraints.The average variability of all reactions decreases dramatically when the growth rate increases.Consider the noise effect on the metabolic system,we thus argue that the microorganism may practically grow under a suboptimal state.Besides,under the EFVA framework,the reactions are easily to be grouped into catabolic and anabolic groups.And the anabolic groups can be further assigned to specific biomass constitute.We also discovered the growth rate dependent essentiality of reactions.

  6. Pulsed-Plasma Disinfection of Water Containing Escherichia coli

    Science.gov (United States)

    Satoh, Kohki; MacGregor, Scott J.; Anderson, John G.; Woolsey, Gerry A.; Fouracre, R. Anthony

    2007-03-01

    The disinfection of water containing the microorganism, Escherichia coli (E. coli) by exposure to a pulsed-discharge plasma generated above the water using a multineedle electrode (plasma-exposure treatment), and by sparging the off-gas of the pulsed plasma into the water (off-gas-sparging treatment), is performed in the ambient gases of air, oxygen, and nitrogen. For the off-gas-sparging treatment, bactericidal action is observed only when oxygen is used as the ambient gas, and ozone is found to generate the bactericidal action. For the plasma-exposure treatment, the density of E. coli bacteria decreases exponentially with plasma-exposure time for all the ambient gases. It may be concluded that the main contributors to E. coli inactivation are particle species produced by the pulsed plasma. For the ambient gases of air and nitrogen, the influence of acidification of the water in the system, as a result of pulsed-plasma exposure, may also contribute to the decay of E. coli density.

  7. Ensemble modeling for aromatic production in Escherichia coli.

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    Matthew L Rizk

    2009-09-01

    Full Text Available Ensemble Modeling (EM is a recently developed method for metabolic modeling, particularly for utilizing the effect of enzyme tuning data on the production of a specific compound to refine the model. This approach is used here to investigate the production of aromatic products in Escherichia coli. Instead of using dynamic metabolite data to fit a model, the EM approach uses phenotypic data (effects of enzyme overexpression or knockouts on the steady state production rate to screen possible models. These data are routinely generated during strain design. An ensemble of models is constructed that all reach the same steady state and are based on the same mechanistic framework at the elementary reaction level. The behavior of the models spans the kinetics allowable by thermodynamics. Then by using existing data from the literature for the overexpression of genes coding for transketolase (Tkt, transaldolase (Tal, and phosphoenolpyruvate synthase (Pps to screen the ensemble, we arrive at a set of models that properly describes the known enzyme overexpression phenotypes. This subset of models becomes more predictive as additional data are used to refine the models. The final ensemble of models demonstrates the characteristic of the cell that Tkt is the first rate controlling step, and correctly predicts that only after Tkt is overexpressed does an increase in Pps increase the production rate of aromatics. This work demonstrates that EM is able to capture the result of enzyme overexpression on aromatic producing bacteria by successfully utilizing routinely generated enzyme tuning data to guide model learning.

  8. Escherichia coli Shiga Toxin Mechanisms of Action in Renal Disease

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    Tom G. Obrig

    2010-12-01

    Full Text Available Shiga toxin-producing Escherichia coli is a contaminant of food and water that in humans causes a diarrheal prodrome followed by more severe disease of the kidneys and an array of symptoms of the central nervous system. The systemic disease is a complex referred to as diarrhea-associated hemolytic uremic syndrome (D+HUS. D+HUS is characterized by thrombocytopenia, microangiopathic hemolytic anemia, and acute renal failure. This review focuses on the renal aspects of D+HUS. Current knowledge of this renal disease is derived from a combination of human samples, animal models of D+HUS, and interaction of Shiga toxin with isolated renal cell types. Shiga toxin is a multi-subunit protein complex that binds to a glycosphingolipid receptor, Gb3, on select eukaryotic cell types. Location of Gb3 in the kidney is predictive of the sites of action of Shiga toxin. However, the toxin is cytotoxic to some, but not all cell types that express Gb3. It also can cause apoptosis or generate an inflammatory response in some cells. Together, this myriad of results is responsible for D+HUS disease.

  9. Experimental Evolution of Escherichia coli Harboring an Ancient Translation Protein.

    Science.gov (United States)

    Kacar, Betül; Ge, Xueliang; Sanyal, Suparna; Gaucher, Eric A

    2017-03-01

    The ability to design synthetic genes and engineer biological systems at the genome scale opens new means by which to characterize phenotypic states and the responses of biological systems to perturbations. One emerging method involves inserting artificial genes into bacterial genomes and examining how the genome and its new genes adapt to each other. Here we report the development and implementation of a modified approach to this method, in which phylogenetically inferred genes are inserted into a microbial genome, and laboratory evolution is then used to examine the adaptive potential of the resulting hybrid genome. Specifically, we engineered an approximately 700-million-year-old inferred ancestral variant of tufB, an essential gene encoding elongation factor Tu, and inserted it in a modern Escherichia coli genome in place of the native tufB gene. While the ancient homolog was not lethal to the cell, it did cause a twofold decrease in organismal fitness, mainly due to reduced protein dosage. We subsequently evolved replicate hybrid bacterial populations for 2000 generations in the laboratory and examined the adaptive response via fitness assays, whole genome sequencing, proteomics, and biochemical assays. Hybrid lineages exhibit a general adaptive strategy in which the fitness cost of the ancient gene was ameliorated in part by upregulation of protein production. Our results suggest that an ancient-modern recombinant method may pave the way for the synthesis of organisms that exhibit ancient phenotypes, and that laboratory evolution of these organisms may prove useful in elucidating insights into historical adaptive processes.

  10. Identification of Diarrheagenic Escherichia coli Strains from Avian Organic Fertilizers

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    Juan Puño-Sarmiento

    2014-08-01

    Full Text Available The Brazilian poultry industry generates large amounts of organic waste, such as chicken litter, which is often used in agriculture. Among the bacteria present in organic fertilizer are members of the Enterobacteriaceae family. The objective of this study was to detect the presence of diarrheagenic Escherichia coli (DEC strains in avian organic fertilizer, and assess the potential damage they can cause in humans due to antimicrobial resistance. The presence of DEC pathotypes and phylogenetic groups were detected by multiplex-PCR. Phenotypic assays, such as tests for adhesion, cytotoxicity activity, biofilm formation and especially antimicrobial susceptibility, were performed. Fifteen DEC strains from 64 E. coli were isolated. Among these, four strains were classified as enteropathogenic (EPEC; 6.2%, three strains as Shiga toxin-producing (STEC; 4.7%, 10 strains as enteroaggregative (EAEC; 12.5%, but two of these harbored the eaeA gene too. The low number of isolated strains was most likely due to the composting process, which reduces the number of microorganisms. These strains were able to adhere to HEp-2 and HeLa cells and produce Shiga-toxins and biofilms; in addition, some of the strains showed antimicrobial resistance, which indicates a risk of the transfer of resistance genes to human E. coli. These results showed that DEC strains isolated from avian organic fertilizers can cause human infections.

  11. Identification of diarrheagenic Escherichia coli strains from avian organic fertilizers.

    Science.gov (United States)

    Puño-Sarmiento, Juan; Gazal, Luis Eduardo; Medeiros, Leonardo P; Nishio, Erick K; Kobayashi, Renata K T; Nakazato, Gerson

    2014-08-28

    The Brazilian poultry industry generates large amounts of organic waste, such as chicken litter, which is often used in agriculture. Among the bacteria present in organic fertilizer are members of the Enterobacteriaceae family. The objective of this study was to detect the presence of diarrheagenic Escherichia coli (DEC) strains in avian organic fertilizer, and assess the potential damage they can cause in humans due to antimicrobial resistance. The presence of DEC pathotypes and phylogenetic groups were detected by multiplex-PCR. Phenotypic assays, such as tests for adhesion, cytotoxicity activity, biofilm formation and especially antimicrobial susceptibility, were performed. Fifteen DEC strains from 64 E. coli were isolated. Among these, four strains were classified as enteropathogenic (EPEC; 6.2%), three strains as Shiga toxin-producing (STEC; 4.7%), 10 strains as enteroaggregative (EAEC; 12.5%), but two of these harbored the eaeA gene too. The low number of isolated strains was most likely due to the composting process, which reduces the number of microorganisms. These strains were able to adhere to HEp-2 and HeLa cells and produce Shiga-toxins and biofilms; in addition, some of the strains showed antimicrobial resistance, which indicates a risk of the transfer of resistance genes to human E. coli. These results showed that DEC strains isolated from avian organic fertilizers can cause human infections.

  12. Expression of fully functional tetrameric human hemoglobin in Escherichia coli

    International Nuclear Information System (INIS)

    Hoffman, S.J.; Looker, D.L.; Roehrich, J.M.; Cozart, P.E.; Durfee, S.L.; Tedesco, J.L.; Stetler, G.L.

    1990-01-01

    Synthesis genes encoding the human α- and β-globin polypeptides have been expressed from a single operon in Escherichia coli. The α- and β-globin polypeptides associate into soluble tetramers, incorporate heme, and accumulate to >5% of the total cellular protein. Purified recombinant hemoglobin has the correct stoichiometry of α- and β-globin chains and contains a full complement of heme. Each globin chain also contains an additional methionine as an extension to the amino terminus. The recombinant hemoglobin has a C 4 reversed-phase HPLC profile essentially identical to that of human hemoglobin A 0 and comigrates with hemoglobin A 0 on SDS/PAGE. The visible spectrum and oxygen affinity are similar to that of native human hemoglobin A 0 . The authors have also expressed the α- and β-globin genes separately and found that the expression of the α-globin gene alone results in a marked decrease in the accumulation of α-globin in the cell. Separate expression of the β-globin gene results in high levels of insoluble β-globin. These observations suggest that the presence of α- and β-globin in the same cell stabilizes α-globin and aids the correct folding of β-globin. This system provides a simple method for expressing large quantities of recombinant hemoglobin and allows facile manipulation of the genes encoding hemoglobin to produce functionally altered forms of this protein

  13. Distribution of Diverse Escherichia coli between Cattle and Pasture.

    Science.gov (United States)

    NandaKafle, Gitanjali; Seale, Tarren; Flint, Toby; Nepal, Madhav; Venter, Stephanus N; Brözel, Volker S

    2017-09-27

    Escherichia coli is widely considered to not survive for extended periods outside the intestines of warm-blooded animals; however, recent studies demonstrated that E. coli strains maintain populations in soil and water without any known fecal contamination. The objective of this study was to investigate whether the niche partitioning of E. coli occurs between cattle and their pasture. We attempted to clarify whether E. coli from bovine feces differs phenotypically and genotypically from isolates maintaining a population in pasture soil over winter. Soil, bovine fecal, and run-off samples were collected before and after the introduction of cattle to the pasture. Isolates (363) were genotyped by uidA and mutS sequences and phylogrouping, and evaluated for curli formation (Rough, Dry, And Red, or RDAR). Three types of clusters emerged, viz. bovine-associated, clusters devoid of cattle isolates and representing isolates endemic to the pasture environment, and clusters with both. All isolates clustered with strains of E. coli sensu stricto, distinct from the cryptic species Clades I, III, IV, and V. Pasture soil endemic and bovine fecal populations had very different phylogroup distributions, indicating niche partitioning. The soil endemic population was largely comprised of phylogroup B1 and had a higher average RDAR score than other isolates. These results indicate the existence of environmental E. coli strains that are phylogenetically distinct from bovine fecal isolates, and that have the ability to maintain populations in the soil environment.

  14. Genomic Comparative Study of Bovine Mastitis Escherichia coli.

    Science.gov (United States)

    Kempf, Florent; Slugocki, Cindy; Blum, Shlomo E; Leitner, Gabriel; Germon, Pierre

    2016-01-01

    Escherichia coli, one of the main causative agents of bovine mastitis, is responsible for significant losses on dairy farms. In order to better understand the pathogenicity of E. coli mastitis, an accurate characterization of E. coli strains isolated from mastitis cases is required. By using phylogenetic analyses and whole genome comparison of 5 currently available mastitis E. coli genome sequences, we searched for genotypic traits specific for mastitis isolates. Our data confirm that there is a bias in the distribution of mastitis isolates in the different phylogenetic groups of the E. coli species, with the majority of strains belonging to phylogenetic groups A and B1. An interesting feature is that clustering of strains based on their accessory genome is very similar to that obtained using the core genome. This finding illustrates the fact that phenotypic properties of strains from different phylogroups are likely to be different. As a consequence, it is possible that different strategies could be used by mastitis isolates of different phylogroups to trigger mastitis. Our results indicate that mastitis E. coli isolates analyzed in this study carry very few of the virulence genes described in other pathogenic E. coli strains. A more detailed analysis of the presence/absence of genes involved in LPS synthesis, iron acquisition and type 6 secretion systems did not uncover specific properties of mastitis isolates. Altogether, these results indicate that mastitis E. coli isolates are rather characterized by a lack of bona fide currently described virulence genes.

  15. Incidence of Escherichia coli  - Glucuronidase Positive on Goat Milk

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    Zorica Voşgan

    2016-11-01

    Full Text Available Papers on beta- glucuronidase sensitivity and specificity for identifying Escherichia coli in sources of environment, food, water, etc. have been published since 1976. In this study we conducted a review of the incidence of E. coli β- glucuronidase -positive in goat milk, obtained by hand milking throughout the lactation: spring, summer, autumn. The presence of E. coli in milk is considered both as a health indicator and a pathogenic factor capable of causing food poisoning. The determination of the E. coli β-glucuronidase-positive was carried using TBX medium by cultivating colonies typical blue at 440C. The absence of E. coli in milk yielded during the spring, when the animal milking is done three times a day, was found in the performed analyses; the same was observed during fall, when the milk production is lower and the milking is done once a day. The load of E. coli β-glucuronidase-positive was averaging 66.67 CFU/ml of goat milk, during the middle lactation period (July-August, in conditions of higher temperature. During this period, milking is done in the mountain zone, where the transhumance of animals takes place in summer. The presence of the species E. coli was also confirmed by microscopic examination. Attention should be paid to hygiene and milk should be immediately cooled, during hot weather, as E. coli can be a source of food poisoning.

  16. Interaction of Escherichia coli with growing salad spinach plants.

    Science.gov (United States)

    Warriner, Keith; Ibrahim, Faozia; Dickinson, Matthew; Wright, Charles; Waites, William M

    2003-10-01

    In this study, the interaction of a bioluminescence-labeled Escherichia coli strain with growing spinach plants was assessed. Through bioluminescence profiles, the direct visualization of E. coli growing around the roots of developing seedlings was accomplished. Subsequent in situ glucuronidase (GUS) staining of seedlings confirmed that E. coli had become internalized within root tissue and, to a limited extent, within hypocotyls. When inoculated seeds were sown in soil microcosms and cultivated for 42 days, E. coli was recovered from the external surfaces of spinach roots and leaves as well as from surface-sterilized roots. When 20-day-old spinach seedlings (from uninoculated seeds) were transferred to soil inoculated with E. coli, the bacterium became established on the plant surface, but internalization into the inner root tissue was restricted. However, for seedlings transferred to a hydroponic system containing 10(2) or 10(3) CFU of E. coli per ml of the circulating nutrient solution, the bacterium was recovered from surface-sterilized roots, indicating that it had been internalized. Differences between E. coli interactions in the soil and those in the hydroponic system may be attributed to greater accessibility of the roots in the latter model. Alternatively, the presence of a competitive microflora in soil may have restricted root colonization by E. coli. The implications of this study's findings with regard to the microbiological safety of minimally processed vegetables are discussed.

  17. Characterization of diarrhoeagenic Escherichia coli isolates in Jordanian children.

    Science.gov (United States)

    Shehabi, Asem A; Bulos, Najawa-Kuri; Hajjaj, Kamal G

    2003-01-01

    In a prospective study carried out among Jordanian children in Amman, a total of 73/250 (29.2%) stool specimens were positive for 1 or more diarrhoeagenic Escherichia coli strains using a multiplex polymerase chain reaction method. This study indicated that diarrhoeagenic E. coli isolates were found frequently more in stools of children with diarrhoea (34%) than without diarrhoea (23.1%), but without any significant difference (p > 0.05). The predominant diarrhoeagenic E. coli strains associated with diarrhoea were enteropathogenic E. coli (11.3%), followed by enterotoxigenic E. coli (9.8%) and enteroaggrative E. coli (9%), whereas in the control group these were 4.3%, 11.1% and 6%, respectively. Enteroinvasive E. coli strains (2.9%) were found only in stools of children with diarrhoea. This study revealed the absence of enterohaemorrhagic E. coli in both diarrhoeal and control stools, and found that diarrhoeagenic E. coli isolates were highly resistance to tetracycline (55%), co-trimoxazole (60%) and ampicillin (89%), which are commonly used antibiotics in Jordan.

  18. Chromosome Replication in Escherichia coli: Life on the Scales

    Science.gov (United States)

    Norris, Vic; Amar, Patrick

    2012-01-01

    At all levels of Life, systems evolve on the 'scales of equilibria'. At the level of bacteria, the individual cell must favor one of two opposing strategies and either take risks to grow or avoid risks to survive. It has been proposed in the Dualism hypothesis that the growth and survival strategies depend on non-equilibrium and equilibrium hyperstructures, respectively. It has been further proposed that the cell cycle itself is the way cells manage to balance the ratios of these types of hyperstructure so as to achieve the compromise solution of living on the two scales. Here, we attempt to re-interpret a major event, the initiation of chromosome replication in Escherichia coli, in the light of scales of equilibria. This entails thinking in terms of hyperstructures as responsible for intensity sensing and quantity sensing and how this sensing might help explain the role of the DnaA protein in initiation of replication. We outline experiments and an automaton approach to the cell cycle that should test and refine the scales concept. PMID:25371267

  19. Current pathogenic Escherichia coli foodborne outbreak cases and therapy development.

    Science.gov (United States)

    Yang, Shih-Chun; Lin, Chih-Hung; Aljuffali, Ibrahim A; Fang, Jia-You

    2017-08-01

    Food contamination by pathogenic microorganisms has been a serious public health problem and a cause of huge economic losses worldwide. Foodborne pathogenic Escherichia coli (E. coli) contamination, such as that with E. coli O157 and O104, is very common, even in developed countries. Bacterial contamination may occur during any of the steps in the farm-to-table continuum from environmental, animal, or human sources and cause foodborne illness. To understand the causes of the foodborne outbreaks by E. coli and food-contamination prevention measures, we collected and investigated the past 10 years' worldwide reports of foodborne E. coli contamination cases. In the first half of this review article, we introduce the infection and symptoms of five major foodborne diarrheagenic E. coli pathotypes: enteropathogenic E. coli (EPEC), Shiga toxin-producing E. coli/enterohemorrhagic E. coli (STEC/EHEC), Shigella/enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), and enterotoxigenic E. coli (ETEC). In the second half of this review article, we introduce the foodborne outbreak cases caused by E. coli in natural foods and food products. Finally, we discuss current developments that can be applied to control and prevent bacterial food contamination.

  20. Recombination Phenotypes of Escherichia coli greA Mutants

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    Poteete Anthony R

    2011-03-01

    Full Text Available Abstract Background The elongation factor GreA binds to RNA polymerase and modulates transcriptional pausing. Some recent research suggests that the primary role of GreA may not be to regulate gene expression, but rather, to promote the progression of replication forks which collide with RNA polymerase, and which might otherwise collapse. Replication fork collapse is known to generate dsDNA breaks, which can be recombinogenic. It follows that GreA malfunction could have consequences affecting homologous recombination. Results Escherichia coli mutants bearing substitutions of the active site acidic residues of the transcription elongation factor GreA, D41N and E44K, were isolated as suppressors of growth inhibition by a toxic variant of the bacteriophage lambda Red-beta recombination protein. These mutants, as well as a D41A greA mutant and a greA deletion, were tested for proficiency in recombination events. The mutations were found to increase the efficiency of RecA-RecBCD-mediated and RecA-Red-mediated recombination, which are replication-independent, and to decrease the efficiency of replication-dependent Red-mediated recombination. Conclusion These observations provide new evidence for a role of GreA in resolving conflicts between replication and transcription.

  1. Noncomplementing diploidy resulting from spontaneous zygogenesis in Escherichia coli.

    Science.gov (United States)

    Gratia, Jean-Pierre

    2005-09-01

    With the aim of understanding sexual reproduction and phenotypic expression, a novel type of mating recently discovered in Escherichia coli was investigated. Termed spontaneous zygogenesis (or Z-mating), it differs from F-mediated conjugation. Its products proved phenotypically unstable, losing part of the phenotype for which they were selected. Inactivation of a parental chromosome in the zygote is strongly suggested by fluctuation tests, respreading experiments, analysis of reisolates, and segregation of non-viable cells detected by epifluorescence staining. Some phenotypically haploid subclones were interpreted as stable noncomplementing diploids carrying an inactivated co-replicating chromosome. Pedigree analysis indicated that the genetic composition of such cells consisted of parental genomes or one parental plus a recombinant genome. Inactivation of a chromosome carrying a prophage resulted in the disappearance of both the ability to produce phage particles and the immunity to superinfection. Phage production signalled transient reactivation of such a chromosome and constituted a sensitive test for stable noncomplementing diploidy. Chromosome inactivation thus appears to be a spontaneous event in bacteria.

  2. Escherichia coli lipoprotein binds human plasminogen via an intramolecular domain

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    Tammy eGonzalez

    2015-10-01

    Full Text Available Escherichia coli lipoprotein (Lpp is a major cellular component that exists in two distinct states, bound-form and free-form. Bound-form Lpp is known to interact with the periplasmic bacterial cell wall, while free-form Lpp is localized to the bacterial cell surface. A function for surface-exposed Lpp has yet to be determined. We hypothesized that the presence of C-terminal lysines in the surface-exposed region of Lpp would facilitate binding to the host zymogen plasminogen, a protease commandeered by a number of clinically important bacteria. Recombinant Lpp was synthesized and the binding of Lpp to plasminogen, the effect of various inhibitors on this binding, and the effects of various mutations of Lpp on Lpp-plasminogen interactions were examined. Additionally, the ability of Lpp-bound plasminogen to be converted to active plasmin was analyzed. We determined that Lpp binds plasminogen via an atypical domain located near the center of mature Lpp that may not be exposed on the surface of intact E. coli according to the current localization model. Finally, we found that plasminogen bound by Lpp can be converted to active plasmin. While the consequences of Lpp binding plasminogen are unclear, these results prompt further investigation of the ability of surface exposed Lpp to interact with host molecules such as extracellular matrix components and complement regulators, and the role of these interactions in infections caused by E. coli and other bacteria.

  3. Adsorption kinetics of Escherichia Coli on different Carbon Nanoforms

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    Md. Shamimul Haque Choudhury

    2012-03-01

    Full Text Available Adsorption of Escherichia coli (E. Coli bacterial cells on different carbon nanoforms (i.e. Single walled carbon nanotube (SWCNT, Multiwalled Carbon nanotube (MWCNT, graphite and mixedFullerene aggregates is studied. The diffusivities of pure cultures of E. Coli cells in SWCNT aggregates, MWCN aggregates, Graphite aggregates and Mixed Fullerenes was observed to be 1.5×10-9 cm2/s, 0.55×10-9 cm2/s, 0.8×10-9 cm2/s, and 1.016×10-9 cm2/s, respectively. In addition to batch adsorption studies, optical microscopy studies were also performed. The results suggest that diffusion kinetics ofbacterial cells depends on the concentration and average diameter of the nano-carbon aggregates and also on the type of material used. Diffusivity of E. Coli. in SWCNT was observed to be highest and isabout three times greater than for MWCNT, about two times greater than for graphite and about 1.5 times greater than for Fullerene aggregates. SWCNT seems to be best candidates (amongst the othermaterials studied for adsorption of microorganisms – paying their way for application towards microorganisms filters and for biosensors (where it is desired to simultaneously detect and capture bio-threat agents.

  4. Chemotactic response and adaptation dynamics in Escherichia coli.

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    Diana Clausznitzer

    2010-05-01

    Full Text Available Adaptation of the chemotaxis sensory pathway of the bacterium Escherichia coli is integral for detecting chemicals over a wide range of background concentrations, ultimately allowing cells to swim towards sources of attractant and away from repellents. Its biochemical mechanism based on methylation and demethylation of chemoreceptors has long been known. Despite the importance of adaptation for cell memory and behavior, the dynamics of adaptation are difficult to reconcile with current models of precise adaptation. Here, we follow time courses of signaling in response to concentration step changes of attractant using in vivo fluorescence resonance energy transfer measurements. Specifically, we use a condensed representation of adaptation time courses for efficient evaluation of different adaptation models. To quantitatively explain the data, we finally develop a dynamic model for signaling and adaptation based on the attractant flow in the experiment, signaling by cooperative receptor complexes, and multiple layers of feedback regulation for adaptation. We experimentally confirm the predicted effects of changing the enzyme-expression level and bypassing the negative feedback for demethylation. Our data analysis suggests significant imprecision in adaptation for large additions. Furthermore, our model predicts highly regulated, ultrafast adaptation in response to removal of attractant, which may be useful for fast reorientation of the cell and noise reduction in adaptation.

  5. Serum Antibodies Protect against Intraperitoneal Challenge with Enterotoxigenic Escherichia coli

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    Xinghong Yang

    2011-01-01

    Full Text Available To assess whether anticolonization factor antigen I (CFA/I fimbriae antibodies (Abs from enterotoxigenic Escherichia coli (ETEC can protect against various routes of challenge, BALB/c mice were immunized with a live attenuated Salmonella vaccine vector expressing CFA/I fimbriae. Vaccinated mice elicited elevated systemic IgG and mucosal IgA Abs, unlike mice immunized with the empty Salmonella vector. Mice were challenged with wild-type ETEC by the oral, intranasal (i.n., and intraperitoneal (i.p. routes. Naïve mice did not succumb to oral challenge, but did to i.n. challenge, as did immunized mice; however, vaccinated mice were protected against i.p. ETEC challenge. Two intramuscular (i.m. immunizations with CFA/I fimbriae without adjuvant conferred 100% protection against i.p. ETEC challenge, while a single 30 μg dose conferred 88% protection. Bactericidal assays showed that ETEC is highly sensitive to anti-CFA/I sera. These results suggest that parenteral immunization with purified CFA/I fimbriae can induce protective Abs and may represent an alternative method to elicit protective Abs for passive immunity to ETEC.

  6. Voltage-gated calcium flux mediates Escherichia coli mechanosensation.

    Science.gov (United States)

    Bruni, Giancarlo N; Weekley, R Andrew; Dodd, Benjamin J T; Kralj, Joel M

    2017-08-29

    Electrically excitable cells harness voltage-coupled calcium influx to transmit intracellular signals, typically studied in neurons and cardiomyocytes. Despite intense study in higher organisms, investigations of voltage and calcium signaling in bacteria have lagged due to their small size and a lack of sensitive tools. Only recently were bacteria shown to modulate their membrane potential on the timescale of seconds, and little is known about the downstream effects from this modulation. In this paper, we report on the effects of electrophysiology in individual bacteria. A genetically encoded calcium sensor expressed in Escherichia coli revealed calcium transients in single cells. A fusion sensor that simultaneously reports voltage and calcium indicated that calcium influx is induced by voltage depolarizations, similar to metazoan action potentials. Cytoplasmic calcium levels and transients increased upon mechanical stimulation with a hydrogel, and single cells altered protein concentrations dependent on the mechanical environment. Blocking voltage and calcium flux altered mechanically induced changes in protein concentration, while inducing calcium flux reproduced these changes. Thus, voltage and calcium relay a bacterial sense of touch and alter cellular lifestyle. Although the calcium effectors remain unknown, these data open a host of new questions about E. coli , including the identity of the underlying molecular players, as well as other signals conveyed by voltage and calcium. These data also provide evidence that dynamic voltage and calcium exists as a signaling modality in the oldest domain of life, and therefore studying electrophysiology beyond canonical electrically excitable cells could yield exciting new findings.

  7. Engineering Escherichia coli for improved ethanol production from gluconate.

    Science.gov (United States)

    Hildebrand, Amanda; Schlacta, Theresa; Warmack, Rebeccah; Kasuga, Takao; Fan, Zhiliang

    2013-10-10

    We report on engineering Escherichia coli to produce ethanol at high yield from gluconic acid (gluconate). Knocking out genes encoding for the competing pathways (l-lactate dehydrogenase and pyruvate formate lyase A) in E. coli KO11 eliminated lactate production, lowered the carbon flow toward acetate production, and improved the ethanol yield from 87.5% to 97.5% of the theoretical maximum, while the growth rate of the mutant strain was about 70% of the wild type. The corresponding genetic modifications led to a small improvement of ethanol yield from 101.5% to 106.0% on glucose. Deletion of the pyruvate dehydrogenase gene (pdh) alone improved the ethanol yield from 87.5% to 90.4% when gluconate was a substrate. The growth rate of the mutant strain was identical to that of the wild type. The corresponding genetic modification led to no improvements on ethanol yield on glucose. Copyright © 2013 Elsevier B.V. All rights reserved.

  8. Antimutagenic effect of isocyanates and related compounds in escherichia coli

    International Nuclear Information System (INIS)

    Kawazoe, Yutaka; Kato, Masanari

    1982-01-01

    Isocyanates and isothiocyanates have been suggested to inactivate enzymes involved in the metabolic activation of chemical carcinogens and the repair of DNA damage. These compounds decrease the mutability of a tester strain of Escherichia coli B under UV irradiation. This paper deals with the antimutagenicity of acylating agents, including isocyanates and isothiocyanates, and some anti-oxidants which are suspected to be anticarcinogenic. The results can be summarized as follows. (1) The antimutagenic effect observed in the present study operates on UV-induced mutagenesis but not on X-ray-induced mutagenesis. (2) This effect operates only on the wild-type strain, H/r30R, but not on Hs30R deficient in the excision repair system. (3) This effect may function through giving the irradiated cells a greater chance to carry out excision repair by prolonging the lag-period before entry into the S-phase. (4) The carbamoylating ability of isocyanates and isothiocyanates may be responsible for the antimutagenicity, but other type of reactivities may also be involved. These antimutagens also participate in inactivating enzymes relevant to the metabolic activation of mutagens, resulting in a decrease in the frequency of chemically induced mutagenesis. (author)

  9. Molecular prophage typing of avian pathogenic Escherichia coli.

    Science.gov (United States)

    Kwon, Hyuk-Joon; Seong, Won-Jin; Kim, Jae-Hong

    2013-03-23

    Escherichia coli prophages confer virulence and resistance to physico-chemical, nutritional, and antibiotic stresses on their hosts, and they enhance the evolution of E. coli. Thus, studies on profiles of E. coli prophages are valuable to understand the population structure and evolution of E. coli pathogenicity. Large terminase genes participate in phage genome packaging and are one of the cornerstones for the identification of prophages. Thus, we designed primers to detect 16 types of large terminase genes and analyzed the genomes of 48 E. coli and Shigella reference strains for the prophage markers. We also investigated the distribution of the 16 prophage markers among 92 avian pathogenic E. coli (APEC) strains. APEC strains were classified into 61 prophage types (PPTs). Each strain was different from the reference strains as measured by the PPTs and from the frequency of each prophage marker. Investigation of the distribution of prophage-related serum resistance (bor), toxin (stx1 and cdtI), and T3SS effector (lom, espK, sopE, nleB, and ospG) genes revealed the presence of bor (44.1%), lom (95.5%) and cdtI (9.1%) in APEC strains with related prophages. Therefore, the molecular prophage typing method may be useful to understand population structure and evolution of E. coli pathogenicity, and further studies on the mobility of the prophages and the roles of virulence genes in APEC pathogenicity may be valuable. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Outbreaks of virulent diarrheagenic Escherichia coli - are we in control?

    Directory of Open Access Journals (Sweden)

    Werber Dirk

    2012-02-01

    Full Text Available Abstract Shiga toxin-producing Escherichia coli (STEC are the most virulent diarrheagenic E. coli known to date. They can be spread with alarming ease via food as exemplified by a large sprout-borne outbreak of STEC O104:H4 in 2011 that was centered in northern Germany and affected several countries. Effective control of such outbreaks is an important public health task and necessitates early outbreak detection, fast identification of the outbreak vehicle and immediate removal of the suspected food from the market, flanked by consumer advice and measures to prevent secondary spread. In our view, opportunities to improve control of STEC outbreaks lie in early clinical suspicion for STEC infection, timely diagnosis of all STEC at the serotype-level and integrating molecular subtyping information into surveillance systems. Furthermore, conducting analytical studies that supplement patients' imperfect food history recall and performing, as an investigative element, product tracebacks, are pivotal but underutilized tools for successful epidemiologic identification of the suspected vehicle in foodborne outbreaks. As a corollary, these tools are amenable to tailor microbiological testing of suspected food. Please see related article: http://www.biomedcentral.com/1741-7015/10/12

  11. PENGARUH PERASAN DAUN BELIMBING WULUH (Averrhoa bilimbi TERHADAP PERTUMBUHAN BAKTERI Escherichia coli PATOGEN

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    Fitrotin Azizah

    2017-12-01

    abstract  (Averrhoa bilimbi is one of the plants that can be used as an antibacterial, good flowers, stems, leaves and stems have benefits and efficacy. Chemical constituents of the leaves starfruit are tannins, flavonoids, saponins. The active ingredient in the leaves starfruit is tannin. Escherichia coli is a bacterium that causes diarrhea. From the above discussion, the authors raised the theme of Influence starfruit juice of the leaves on the growth of pathogenic E. coli bacteria. Formulation of the problem researchers is whether there is influence starfruit juice of the leaves on the growth of Escherichia coli pathogens. This study aims to determine the concentration that could inhibit and kill Esherichia coli Escherichia coli. This research is experimental. The sample used is leaf green starfruit not so young in a fresh state taken in the area around the boarding author Sutorejo 11B stay. In this study, the sample size for each treatment as much as 3 100%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10% and C (control. Independent variables are starfruit juice of the leaves, while the dependent variable growth of Escherichia coli. When the study carried out in January and July 2012. Data on the effect of starfruit juice of the leaves on the growth of Escherichia coli tested by laboratory examination and data collection techniques using Chi-Square 0:05. Based on the results it appears that at a concentration of 100% and 90% were able to kill the bacteria Escherichia coli, whereas the inhibitory power ranging from a concentration of 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%. From Chi-Square test was obtained λ2 count Escherichia coli pathogenic bacteria. Keyword : Leaves starfruit, E. Coli

  12. Longitudinal characterization of Escherichia coli in healthy captive nonhuman primates

    Directory of Open Access Journals (Sweden)

    Jonathan B Clayton

    2014-11-01

    Full Text Available The gastrointestinal (GI tracts of nonhuman primates are well known to harbor Escherichia coli, a known commensal of humans and animals. While E. coli is a normal inhabitant of the mammalian gut, it also exists in a number of pathogenic forms or pathotypes, including those with predisposition for the GI tract, as well the urogenital tract. Diarrhea in captive nonhuman primates (NHPs has long been a problem in both zoo settings and research colonies, including the Como Zoo. It is an animal welfare concern, as well as a public health concern. E. coli has not been extensively studied in correlation with diarrhea in captive primates; therefore, a study was performed during the summer of 2009 in collaboration with a zoo in Saint Paul, MN, which was experiencing an increased incidence and severity of diarrhea among their NHP collection. Fresh fecal samples were collected weekly from each member of the primate collection, between June and August of 2009, and E. coli were isolated. A total of 33 individuals were included in the study, representing eight species. E. coli isolates were examined for their genetic relatedness, phylogenetic relationships, plasmid replicon types, virulence gene profiles, and antimicrobial susceptibility profiles. A number of isolates were identified containing virulence genes commonly found in several different E. coli pathotypes, and there was evidence of clonal transmission of isolates between animals and over time. Overall, the manifestation of chronic diarrhea in the Como Zoo primate collection is a complex problem whose solution will require regular screening for microbial agents and consideration of environmental causes. This study provides some insight towards the sharing of enteric bacteria between such animals.

  13. Deactivation of Escherichia coli by the plasma needle

    International Nuclear Information System (INIS)

    Sladek, R E J; Stoffels, E

    2005-01-01

    In this paper we present a parameter study on deactivation of Escherichia coli (E. coli) by means of a non-thermal plasma (plasma needle). The plasma needle is a small-sized (1 mm) atmospheric glow sustained by radio-frequency excitation. This plasma will be used to disinfect heat-sensitive objects; one of the intended applications is in vivo deactivation of dental bacteria: destruction of plaque and treatment of caries. We use E. coli films plated on agar dishes as a model system to optimize the conditions for bacterial destruction. Plasma power, treatment time and needle-to-sample distance are varied. Plasma treatment of E. coli films results in formation of a bacteria-free void with a size up to 12 mm. 10 4 -10 5 colony forming units are already destroyed after 10 s of treatment. Prolongation of treatment time and usage of high powers do not significantly improve the destruction efficiency: short exposure at low plasma power is sufficient. Furthermore, we study the effects of temperature increase on the survival of E. coli and compare it with thermal effects of the plasma. The population of E. coli heated in a warm water bath starts to decrease at temperatures above 40 deg. C. Sample temperature during plasma treatment has been monitored. The temperature can reach up to 60 deg. C at high plasma powers and short needle-to-sample distances. However, thermal effects cannot account for bacterial destruction at low power conditions. For safe and efficient in vivo disinfection, the sample temperature should be kept low. Thus, plasma power and treatment time should not exceed 150 mW and 60 s, respectively

  14. Reduction of verotoxigenic Escherichia coli in production of fermented sausages.

    Science.gov (United States)

    Holck, Askild L; Axelsson, Lars; Rode, Tone Mari; Høy, Martin; Måge, Ingrid; Alvseike, Ole; L'abée-Lund, Trine M; Omer, Mohamed K; Granum, Per Einar; Heir, Even

    2011-11-01

    After a number of foodborne outbreaks of verotoxigenic Escherichia coli involving fermented sausages, some countries have imposed regulations on sausage production. For example, the US Food Safety and Inspection Service requires a 5 log(10) reduction of E. coli in fermented products. Such regulations have led to a number of studies on the inactivation of E. coli in fermented sausages by changing processing and post-processing conditions. Several factors influence the survival of E. coli such as pre-treatment of the meat, amount of NaCl, nitrite and lactic acid, water activity, pH, choice of starter cultures and addition of antimicrobial compounds. Also process variables like fermentation temperature and storage time play important roles. Though a large variety of different production processes of sausages exist, generally the reduction of E. coli caused by production is in the range 1-2 log(10). In many cases this may not be enough to ensure microbial food safety. By optimising ingredients and process parameters it is possible to increase E. coli reduction to some extent, but in some cases still other post process treatments may be required. Such treatments may be storage at ambient temperatures, specific heat treatments, high pressure processing or irradiation. HACCP analyses have identified the quality of the raw materials, low temperature in the batter when preparing the sausages and a rapid pH drop during fermentation as critical control points in sausage production. This review summarises the literature on the reduction verotoxigenic E. coli in production of fermented sausages. Copyright © 2011 Elsevier Ltd. All rights reserved.

  15. Starved Escherichia coli preserve reducing power under nitric oxide stress

    Energy Technology Data Exchange (ETDEWEB)

    Gowers, Glen-Oliver F. [Department of Molecular Biology, Princeton University, Princeton, NJ (United States); Robinson, Jonathan L. [Department of Chemical and Biological Engineering, Princeton University, Princeton, NJ (United States); Brynildsen, Mark P., E-mail: mbrynild@princeton.edu [Department of Molecular Biology, Princeton University, Princeton, NJ (United States); Department of Chemical and Biological Engineering, Princeton University, Princeton, NJ (United States)

    2016-07-15

    Nitric oxide (NO) detoxification enzymes, such as NO dioxygenase (NOD) and NO reductase (NOR), are important to the virulence of numerous bacteria. Pathogens use these defense systems to ward off immune-generated NO, and they do so in environments that contain additional stressors, such as reactive oxygen species, nutrient deprivation, and acid stress. NOD and NOR both use reducing equivalents to metabolically deactivate NO, which suggests that nutrient deprivation could negatively impact their functionality. To explore the relationship between NO detoxification and nutrient deprivation, we examined the ability of Escherichia coli to detoxify NO under different levels of carbon source availability in aerobic cultures. We observed failure of NO detoxification under both carbon source limitation and starvation, and those failures could have arisen from inabilities to synthesize Hmp (NOD of E. coli) and/or supply it with sufficient NADH (preferred electron donor). We found that when limited quantities of carbon source were provided, NO detoxification failed due to insufficient NADH, whereas starvation prevented Hmp synthesis, which enabled cells to maintain their NADH levels. This maintenance of NADH levels under starvation was confirmed to be dependent on the absence of Hmp. Intriguingly, these data show that under NO stress, carbon-starved E. coli are better positioned with regard to reducing power to cope with other stresses than cells that had consumed an exhaustible amount of carbon. -- Highlights: •Carbon source availability is critical to aerobic E. coli NO detoxification. •Carbon source starvation, under NO stress, preserves intracellular NADH levels. •Preservation of NADH depends on starvation-dependent inhibition of Hmp induction.

  16. Copper Homeostasis in Escherichia coli and Other Enterobacteriaceae.

    Science.gov (United States)

    Rensing, Christopher; Franke, Sylvia

    2007-04-01

    An interesting model for studying environmental influences shaping microbial evolution is provided by a multitude of copper resistance and copper homeostasis determinants in enteric bacteria. This review describes these determinants and tries to relate their presence to the habitat of the respective organism, as a current hypothesis predicts that the environment should determine an organism's genetic makeup. In Escherichia coli there are four regulons that are induced in the presence of copper. Two, the CueR and the CusR regulons, are described in detail. A central component regulating intracellular copper levels, present in all free-living enteric bacteria whose genomes have so far been sequenced, is a Cu(I)translocating P-type ATPase. The P-type ATPase superfamily is a ubiquitous group of proteins involved in the transport of charged substrates across biological membranes. Whereas some components involved in copper homeostasis can be found in both anaerobes and aerobes, multi-copper oxidases (MCOs) implicated in copper tolerance in E. coli, such as CueO and the plasmid-based PcoA, can be found only in aerobic organisms. Several features indicate that CueO, PcoA, and other related MCOs are specifically adapted to combat copper-mediated oxidative damage. In addition to these well-characterized resistance operons, there are numerous other genes that appear to be involved in copper binding and trafficking that have not been studied in great detail. SilE and its homologue PcoE, for example, are thought to effect the periplasmic binding and sequestration of silver and copper, respectively.

  17. Killing of Escherichia coli using the gas diffusion electrode system.

    Science.gov (United States)

    Xu, W Y; Li, P; Dong, B

    2010-01-01

    To be best of our knowledge, this study is one of the first investigations to be performed into the potential benefits of gas diffusion electrode (GDE) system in controlling inactivation of E. coli. This study mainly focused on the dual electrodes disinfection with gas diffusion cathode, using Escherichia coli as the indicator microorganisms. The effects of Pt load W(Pt) and the pore-forming agent content W(NH(4)HCO(3)) in GDE, operating conditions such as pH value, oxygen flow rate Q(O(2)), salt content and current density on the disinfection were investigated, respectively. The experimental results showed that the disinfection improved with increasing Pt load W(Pt), but its efficiency at Pt load of 3 per thousand was equivalent to that at Pt load of 4 per thousand. Addition of the pore-forming agent in the appropriate amount improved the disinfection while drop of pH value resulted in the rapid rise of the germicidal efficacy and the disinfection shortened with increasing oxygen flow rate Q(O(2)). The system is more suitable for highly salt water. The germicidal efficacy increased with current density. However, the accelerating rate was different: it first increased with the current density, then decreased, and reached a maximum at current density of 6.7-8.3 mA/cm(2). The germicidal efficacy in the cathode compartment was about the same as in the anode compartment indicating the contribution of direct oxidation and indirect treatment of E. coli by the hydroxyl radical was similar to the oxidative indirect effect of the generated H(2)O(2). This technology is expensive in operating cost, further research is required to advance the understanding and reduce the operating cost of this technology.

  18. The upper surface of an Escherichia coli swarm is stationary.

    Science.gov (United States)

    Zhang, Rongjing; Turner, Linda; Berg, Howard C

    2010-01-05

    When grown in a rich medium on agar, many bacteria elongate, produce more flagella, and swim in a thin film of fluid over the agar surface in swirling packs. Cells that spread in this way are said to swarm. The agar is a solid gel, with pores smaller than the bacteria, so the swarm/agar interface is fixed. Here we show, in experiments with Escherichia coli, that the swarm/air interface also is fixed. We deposited MgO smoke particles on the top surface of an E. coli swarm near its advancing edge, where cells move in a single layer, and then followed the motion of the particles by dark-field microscopy and the motion of the underlying cells by phase-contrast microscopy. Remarkably, the smoke particles remained fixed (diffusing only a few micrometers) while the swarming cells streamed past underneath. The diffusion coefficients of the smoke particles were smaller over the virgin agar ahead of the swarm than over the swarm itself. Changes between these two modes of behavior were evident within 10-20 microm of the swarm edge, indicating an increase in depth of the fluid in advance of the swarm. The only plausible way that the swarm/air interface can be fixed is that it is covered by a surfactant monolayer pinned at its edges. When a swarm is exposed to air, such a monolayer can markedly reduce water loss. When cells invade tissue, the ability to move rapidly between closely opposed fixed surfaces is a useful trait.

  19. Genomic and Phenomic Study of Mammary Pathogenic Escherichia coli

    Science.gov (United States)

    Blum, Shlomo E.; Heller, Elimelech D.; Sela, Shlomo; Elad, Daniel; Edery, Nir; Leitner, Gabriel

    2015-01-01

    Escherichia coli is a major etiological agent of intra-mammary infections (IMI) in cows, leading to acute mastitis and causing great economic losses in dairy production worldwide. Particular strains cause persistent IMI, leading to recurrent mastitis. Virulence factors of mammary pathogenic E. coli (MPEC) involved pathogenesis of mastitis as well as those differentiating strains causing acute or persistent mastitis are largely unknown. This study aimed to identify virulence markers in MPEC through whole genome and phenome comparative analysis. MPEC strains causing acute (VL2874 and P4) or persistent (VL2732) mastitis were compared to an environmental strain (K71) and to the genomes of strains representing different E. coli pathotypes. Intra-mammary challenge in mice confirmed experimentally that the strains studied here have different pathogenic potential, and that the environmental strain K71 is non-pathogenic in the mammary gland. Analysis of whole genome sequences and predicted proteomes revealed high similarity among MPEC, whereas MPEC significantly differed from the non-mammary pathogenic strain K71, and from E. coli genomes from other pathotypes. Functional features identified in MPEC genomes and lacking in the non-mammary pathogenic strain were associated with synthesis of lipopolysaccharide and other membrane antigens, ferric-dicitrate iron acquisition and sugars metabolism. Features associated with cytotoxicity or intra-cellular survival were found specifically in the genomes of strains from severe and acute (VL2874) or persistent (VL2732) mastitis, respectively. MPEC genomes were relatively similar to strain K-12, which was subsequently shown here to be possibly pathogenic in the mammary gland. Phenome analysis showed that the persistent MPEC was the most versatile in terms of nutrients metabolized and acute MPEC the least. Among phenotypes unique to MPEC compared to the non-mammary pathogenic strain were uric acid and D-serine metabolism. This study

  20. Toxicity mechanism of carbon nanotubes on Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Young, Yu-Fu [Department of Materials Science and Engineering, National Tsing-Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Lee, Hui-Ju [Department of Life Science, National Tsing-Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Shen, Yi-Shan; Tseng, Shih-Hao; Lee, Chi-Young [Department of Materials Science and Engineering, National Tsing-Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Tai, Nyan-Hwa, E-mail: nhtai@mx.nthu.edu.tw [Department of Materials Science and Engineering, National Tsing-Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Chang, Hwan-You, E-mail: hychang@mx.nthu.edu.tw [Department of Life Science, National Tsing-Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China)

    2012-05-15

    Highlights: Black-Right-Pointing-Pointer F-MWCNTs possess higher antibiotic performance than that of the F-SWCNTs. Black-Right-Pointing-Pointer E. coli cells were pierced when incubated with F-MWCNTs and trapped when incubated with F-SWCNTs. Black-Right-Pointing-Pointer The rigidity and moment of CNTs play important role on the antibiotic effect. - Abstract: The influences of carbon nanomaterials on bacteria were investigated using three types of dispersed and functionalized carbon nanomaterials (F-CNMs), viz. functionalized carbon nanopowder (F-CNP), functionalized single-walled carbon nanotubes (F-SWCNTs), and functionalized multi-walled carbon nanotubes (F-MWCNTs). F-CNMs with different aspect ratios were used to study the influence of material configuration on the viability of Escherichia coli (E. coli). Although these materials were functionalized to improve their dispersibility, the original morphologies and chemical properties of the materials were maintained. Traditional bacteria quantitative plating analysis was conducted, and the results of which revealed that the F-CNP and the F-SWCNTs showed a less significant effect on the viability of E. coli, while the F-MWCNTs obviously inhibited cell viability. A Fourier transform infrared spectroscopy and a scanning electron microscopy were used to verify the functionalization of the F-CNMs and to examine the interaction of F-CNMs with E. coli, respectively; in addition, we adopted chemiluminescence assays to measure the concentration of adenosine triphosphate (ATP) released from the damaged cells. The results showed that the ATP of the F-MWCNTs sample is two-fold higher than that of the control, indicating direct piercing of E. coli by F-MWCNTs leads to bacteria death. Furthermore, F-SWCNTs were concluded to have less influence on the viability of E. coli because ultra-long F-SWCNTs used in this study performed less rigidity to pierce the cells.

  1. Toxicity mechanism of carbon nanotubes on Escherichia coli

    International Nuclear Information System (INIS)

    Young, Yu-Fu; Lee, Hui-Ju; Shen, Yi-Shan; Tseng, Shih-Hao; Lee, Chi-Young; Tai, Nyan-Hwa; Chang, Hwan-You

    2012-01-01

    Highlights: ► F-MWCNTs possess higher antibiotic performance than that of the F-SWCNTs. ► E. coli cells were pierced when incubated with F-MWCNTs and trapped when incubated with F-SWCNTs. ► The rigidity and moment of CNTs play important role on the antibiotic effect. - Abstract: The influences of carbon nanomaterials on bacteria were investigated using three types of dispersed and functionalized carbon nanomaterials (F-CNMs), viz. functionalized carbon nanopowder (F-CNP), functionalized single-walled carbon nanotubes (F-SWCNTs), and functionalized multi-walled carbon nanotubes (F-MWCNTs). F-CNMs with different aspect ratios were used to study the influence of material configuration on the viability of Escherichia coli (E. coli). Although these materials were functionalized to improve their dispersibility, the original morphologies and chemical properties of the materials were maintained. Traditional bacteria quantitative plating analysis was conducted, and the results of which revealed that the F-CNP and the F-SWCNTs showed a less significant effect on the viability of E. coli, while the F-MWCNTs obviously inhibited cell viability. A Fourier transform infrared spectroscopy and a scanning electron microscopy were used to verify the functionalization of the F-CNMs and to examine the interaction of F-CNMs with E. coli, respectively; in addition, we adopted chemiluminescence assays to measure the concentration of adenosine triphosphate (ATP) released from the damaged cells. The results showed that the ATP of the F-MWCNTs sample is two-fold higher than that of the control, indicating direct piercing of E. coli by F-MWCNTs leads to bacteria death. Furthermore, F-SWCNTs were concluded to have less influence on the viability of E. coli because ultra-long F-SWCNTs used in this study performed less rigidity to pierce the cells.

  2. Production of recombinant proteins from Plasmodium falciparum in Escherichia coli.

    Science.gov (United States)

    Guerra, Ángela Patricia; Calvo, Eliana Patricia; Wasserman, Moisés; Chaparro-Olaya, Jacqueline

    2016-02-23

    The production of recombinant proteins is essential for the characterization and functional study of proteins from Plasmodium falciparum. However, the proteins of P. falciparum are among the most challenging to express, and when expression is achieved, the recombinant proteins usually fold incorrectly and lead to the formation of inclusion bodies.  To obtain and purify four recombinant proteins and to use them as antigens to produce polyclonal antibodies. The production efficiency and solubility were evaluated as the proteins were expressed in two genetically modified strains of Escherichia coli to favor the production of heterologous proteins (BL21-CodonPlus (DE3)-RIL and BL21-pG-KJE8).  The four recombinant P. falciparum proteins corresponding to partial sequences of PfMyoA (Myosin A) and PfGAP50 (gliding associated protein 50), and the complete sequences of PfMTIP (myosin tail interacting protein) and PfGAP45 (gliding associated protein 45), were produced as glutathione S-transferase-fusion proteins, purified and used for immunizing mice.  The protein expression was much more efficient in BL21-CodonPlus, the strain that contains tRNAs that are rare in wild-type E. coli, compared to the expression in BL21-pG-KJE8. In spite of the fact that BL21-pG-KJE8 overexpresses chaperones, this strain did not minimize the formation of inclusion bodies.  The use of genetically modified strains of E. coli was essential to achieve high expression levels of the four evaluated P. falciparum proteins and lead to improved solubility of two of them. The approach used here allowed us to obtain and purify four P. falciparum proteins in enough quantity to produce polyclonal antibodies in mice, and a fair amount of two pure and soluble recombinant proteins for future assays.

  3. Discovery of Escherichia coli CRISPR sequences in an undergraduate laboratory.

    Science.gov (United States)

    Militello, Kevin T; Lazatin, Justine C

    2017-05-01

    Clustered regularly interspaced short palindromic repeats (CRISPRs) represent a novel type of adaptive immune system found in eubacteria and archaebacteria. CRISPRs have recently generated a lot of attention due to their unique ability to catalog foreign nucleic acids, their ability to destroy foreign nucleic acids in a mechanism that shares some similarity to RNA interference, and the ability to utilize reconstituted CRISPR systems for genome editing in numerous organisms. In order to introduce CRISPR biology into an undergraduate upper-level laboratory, a five-week set of exercises was designed to allow students to examine the CRISPR status of uncharacterized Escherichia coli strains and to allow the discovery of new repeats and spacers. Students started the project by isolating genomic DNA from E. coli and amplifying the iap CRISPR locus using the polymerase chain reaction (PCR). The PCR products were analyzed by Sanger DNA sequencing, and the sequences were examined for the presence of CRISPR repeat sequences. The regions between the repeats, the spacers, were extracted and analyzed with BLASTN searches. Overall, CRISPR loci were sequenced from several previously uncharacterized E. coli strains and one E. coli K-12 strain. Sanger DNA sequencing resulted in the discovery of 36 spacer sequences and their corresponding surrounding repeat sequences. Five of the spacers were homologous to foreign (non-E. coli) DNA. Assessment of the laboratory indicates that improvements were made in the ability of students to answer questions relating to the structure and function of CRISPRs. Future directions of the laboratory are presented and discussed. © 2016 by The International Union of Biochemistry and Molecular Biology, 45(3):262-269, 2017. © 2016 The International Union of Biochemistry and Molecular Biology.

  4. papA gene of avian pathogenic Escherichia coli.

    Science.gov (United States)

    Kariyawasam, Subhashinie; Nolan, Lisa K

    2011-12-01

    P fimbrial adhesins may be associated with the virulence of avian pathogenic Escherichia coli (APEC). However, most APECs are unable to express P fimbriae even when they are grown under conditions that favor P fimbrial expression. This failure can be explained by the complete absence of the pap operon or the presence of an incomplete pap operon in Pap-negative APEC strains. In the present study, we analyzed the pap operon, specifically the papA gene that encodes the major fimbrial shaft, to better understand the pap gene cluster at the genetic level. First, by PCR, we examined a collection of 500 APEC strains for the presence of 11 genes comprising the pap operon. Except for papA, all the other genes of the operon were present in 38% to 41.2% of APEC, whereas the papA was present only in 10.4% of the APEC tested. Using multiplex PCR to probe for allelic variants of papA, we sought to determine if the low prevalence of papA among APEC was related to genetic heterogeneity of the gene itself. It was determined that the papA of APEC always belongs to the F11 allelic variant. Finally, we sequenced the 'papA region' from two papA-negative strains, both of which contain all the other genes of the pap operon. Interestingly, both strains had an 11,104-bp contig interruptingpapA at the 281-bp position. This contig harbored a streptomycin resistance gene and a classic Tn10 transposon containing the genes that confer tetracycline resistance. However, we noted that the papA gene of every papA-negative APEC strain was not interrupted by an 11,104-bp contig. It is likely that transposons bearing antibiotic resistance genes have inserted within pap gene cluster of some APEC strains, and such genetic events may have been selected for by antibiotic use.

  5. Synanthropic rodents as possible reservoirs of shigatoxigenic Escherichia coli strains

    Directory of Open Access Journals (Sweden)

    Ximena eBlanco Crivelli

    2012-11-01

    Full Text Available Shigatoxigenic Escherichia coli (STEC strains are worldwide zoonotic pathogen responsible for different cases of human disease including hemolytic uremic syndrome (HUS. Transmission of STEC to humans occurs through the consumption of food and water contaminated by faeces of carriers and by person-to-person contact.The objective of this study was twofold: (a to investigate whether synanthropic rodents are possible reservoirs of STEC in the urban area and (b whether a particular genus out of synanthropic rodent is the principal carrier of STEC.One hundred forty-five rodents were captured in Buenos Aires City. Screening for stx1/stx2 and rfbO157 was done by PCR from the confluence zone. STEC isolates were further characterized with biochemical tests by standard methods. Additional virulence factors (eae, ehxA and saa were also determined by PCR. Forty-one of the rodents were necropsied and sample of kidney and small and large intestine were taken for histopathological diagnosis. The samples sections were stained with hematoxylin-eosin, and observed by light microscopy to evaluate the systemic involvement of these species in natural infections. STEC was isolated from seven out of twenty seven suspect animals at screening. The following genotypes were found in the STEC strains: stx1/stx2/ehxA (1, stx2 (4, stx2/ehxA (1, stx2/ehxA/eae (1. Neither gross nor microscopic lesions compatible with those produced by Shiga toxin were observed in the studied organs of necropsied rodents.The bivariate analysis including the hundred forty-five rodents data showed that the isolation of STEC is associated positively to Rattus genus. This synanthropic species may play a role in the transmissibility of the agent thus being a risk to the susceptible population. Their control should be included specifically in actions to dismiss the contamination of food and water by STEC in the urban area, as additional strategies for epidemiological control.

  6. Intestinal Pathogenic Escherichia coli: Insights for Vaccine Development

    Directory of Open Access Journals (Sweden)

    Maricarmen Rojas-Lopez

    2018-03-01

    Full Text Available Diarrheal diseases are one of the major causes of mortality among children under five years old and intestinal pathogenic Escherichia coli (InPEC plays a role as one of the large causative groups of these infections worldwide. InPECs contribute significantly to the burden of intestinal diseases, which are a critical issue in low- and middle-income countries (Asia, Africa and Latin America. Intestinal pathotypes such as enteropathogenic E. coli (EPEC and enterotoxigenic E. coli (ETEC are mainly endemic in developing countries, while ETEC strains are the major cause of diarrhea in travelers to these countries. On the other hand, enterohemorrhagic E. coli (EHEC are the cause of large outbreaks around the world, mainly affecting developed countries and responsible for not only diarrheal disease but also severe clinical complications like hemorrhagic colitis and hemolytic uremic syndrome (HUS. Overall, the emergence of antibiotic resistant strains, the annual cost increase in the health care system, the high incidence of traveler diarrhea and the increased number of HUS episodes have raised the need for effective preventive treatments. Although the use of antibiotics is still important in treating such infections, non-antibiotic strategies are either a crucial option to limit the increase in antibiotic resistant strains or absolutely necessary for diseases such as those caused by EHEC infections, for which antibiotic therapies are not recommended. Among non-antibiotic therapies, vaccine development is a strategy of choice but, to date, there is no effective licensed vaccine against InPEC infections. For several years, there has been a sustained effort to identify efficacious vaccine candidates able to reduce the burden of diarrheal disease. The aim of this review is to summarize recent milestones and insights in vaccine development against InPECs.

  7. Genetic determinants of heat resistance in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Ryan eMercer

    2015-09-01

    Full Text Available Escherichia coli AW1.7 is a heat resistant food isolate and the occurrence of pathogenic strains with comparable heat resistance may pose a risk to food safety. To identify the genetic determinants of heat resistance, 29 strains of E. coli that differed in their of heat resistance were analyzed by comparative genomics. Strains were classified as highly heat resistant strains, exhibiting a D60-value of more than 6 min; moderately heat resistant strains, exhibiting a D60-value of more than 1 min; or as heat sensitive. A ~14 kb genomic island containing 16 predicted open reading frames encoding putative heat shock proteins and proteases was identified only in highly heat resistant strains. The genomic island was termed the locus of heat resistance (LHR. This putative operon is flanked by mobile elements and possesses >99% sequence identity to genomic islands contributing to heat resistance in Cronobacter sakazakii and Klebsiella pneumoniae. An additional 41 LHR sequences with >87% sequence identity were identified in 11 different species of β- and γ-proteobacteria. Cloning of the full length LHR conferred high heat resistance to the heat sensitive E. coli AW1.7ΔpHR1 and DH5α. The presence of the LHR correlates perfectly to heat resistance in several species of Enterobacteriaceae and occurs at a frequency of 2% of all E. coli genomes, including pathogenic strains. This study suggests the LHR has been laterally exchanged among the β- and γ-proteobacteria and is a reliable indicator of high heat resistance in E. coli.

  8. Phosphotyrosine-Mediated Regulation of Enterohemorrhagic Escherichia coli Virulence

    Science.gov (United States)

    Robertson, Colin D.; Hazen, Tracy H.; Kaper, James B.

    2018-01-01

    ABSTRACT Enteric pathogens with low infectious doses rely on the ability to orchestrate the expression of virulence and metabolism-associated genes in response to environmental cues for successful infection. Accordingly, the human pathogen enterohemorrhagic Escherichia coli (EHEC) employs a complex multifaceted regulatory network to link the expression of type III secretion system (T3SS) components to nutrient availability. While phosphorylation of histidine and aspartate residues on two-component system response regulators is recognized as an integral part of bacterial signaling, the involvement of phosphotyrosine-mediated control is minimally explored in Gram-negative pathogens. Our recent phosphotyrosine profiling study of E. coli identified 342 phosphorylated proteins, indicating that phosphotyrosine modifications in bacteria are more prevalent than previously anticipated. The present study demonstrates that tyrosine phosphorylation of a metabolite-responsive LacI/GalR family regulator, Cra, negatively affects T3SS expression under glycolytic conditions that are typical for the colonic lumen environment where production of the T3SS is unnecessary. Our data suggest that Cra phosphorylation affects T3SS expression by modulating the expression of ler, which encodes the major activator of EHEC virulence gene expression. Phosphorylation of the Cra Y47 residue diminishes DNA binding to fine-tune the expression of virulence-associated genes, including those of the locus of enterocyte effacement pathogenicity island that encode the T3SS, and thereby negatively affects the formation of attaching and effacing lesions. Our data indicate that tyrosine phosphorylation provides an additional mechanism to control the DNA binding of Cra and other LacI/GalR family regulators, including LacI and PurR. This study describes an initial effort to unravel the role of global phosphotyrosine signaling in the control of EHEC virulence potential. PMID:29487233

  9. Catabolite and Oxygen Regulation of Enterohemorrhagic Escherichia coli Virulence

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    Kimberly M. Carlson-Banning

    2016-11-01

    Full Text Available The biogeography of the gut is diverse in its longitudinal axis, as well as within specific microenvironments. Differential oxygenation and nutrient composition drive the membership of microbial communities in these habitats. Moreover, enteric pathogens can orchestrate further modifications to gain a competitive advantage toward host colonization. These pathogens are versatile and adept when exploiting the human colon. They expertly navigate complex environmental cues and interkingdom signaling to colonize and infect their hosts. Here we demonstrate how enterohemorrhagic Escherichia coli (EHEC uses three sugar-sensing transcription factors, Cra, KdpE, and FusR, to exquisitely regulate the expression of virulence factors associated with its type III secretion system (T3SS when exposed to various oxygen concentrations. We also explored the effect of mucin-derived nonpreferred carbon sources on EHEC growth and expression of virulence genes. Taken together, the results show that EHEC represses the expression of its T3SS when oxygen is absent, mimicking the largely anaerobic lumen, and activates its T3SS when oxygen is available through Cra. In addition, when EHEC senses mucin-derived sugars heavily present in the O-linked and N-linked glycans of the large intestine, virulence gene expression is initiated. Sugars derived from pectin, a complex plant polysaccharide digested in the large intestine, also increased virulence gene expression. Not only does EHEC sense host- and microbiota-derived interkingdom signals, it also uses oxygen availability and mucin-derived sugars liberated by the microbiota to stimulate expression of the T3SS. This precision in gene regulation allows EHEC to be an efficient pathogen with an extremely low infectious dose.

  10. Phosphotyrosine-Mediated Regulation of Enterohemorrhagic Escherichia coli Virulence

    Directory of Open Access Journals (Sweden)

    Colin D. Robertson

    2018-02-01

    Full Text Available Enteric pathogens with low infectious doses rely on the ability to orchestrate the expression of virulence and metabolism-associated genes in response to environmental cues for successful infection. Accordingly, the human pathogen enterohemorrhagic Escherichia coli (EHEC employs a complex multifaceted regulatory network to link the expression of type III secretion system (T3SS components to nutrient availability. While phosphorylation of histidine and aspartate residues on two-component system response regulators is recognized as an integral part of bacterial signaling, the involvement of phosphotyrosine-mediated control is minimally explored in Gram-negative pathogens. Our recent phosphotyrosine profiling study of E. coli identified 342 phosphorylated proteins, indicating that phosphotyrosine modifications in bacteria are more prevalent than previously anticipated. The present study demonstrates that tyrosine phosphorylation of a metabolite-responsive LacI/GalR family regulator, Cra, negatively affects T3SS expression under glycolytic conditions that are typical for the colonic lumen environment where production of the T3SS is unnecessary. Our data suggest that Cra phosphorylation affects T3SS expression by modulating the expression of ler, which encodes the major activator of EHEC virulence gene expression. Phosphorylation of the Cra Y47 residue diminishes DNA binding to fine-tune the expression of virulence-associated genes, including those of the locus of enterocyte effacement pathogenicity island that encode the T3SS, and thereby negatively affects the formation of attaching and effacing lesions. Our data indicate that tyrosine phosphorylation provides an additional mechanism to control the DNA binding of Cra and other LacI/GalR family regulators, including LacI and PurR. This study describes an initial effort to unravel the role of global phosphotyrosine signaling in the control of EHEC virulence potential.

  11. High mutation rates limit evolutionary adaptation in Escherichia coli

    Science.gov (United States)

    Wagner, Andreas

    2018-01-01

    Mutation is fundamental to evolution, because it generates the genetic variation on which selection can act. In nature, genetic changes often increase the mutation rate in systems that range from viruses and bacteria to human tumors. Such an increase promotes the accumulation of frequent deleterious or neutral alleles, but it can also increase the chances that a population acquires rare beneficial alleles. Here, we study how up to 100-fold increases in Escherichia coli’s genomic mutation rate affect adaptive evolution. To do so, we evolved multiple replicate populations of asexual E. coli strains engineered to have four different mutation rates for 3000 generations in the laboratory. We measured the ability of evolved populations to grow in their original environment and in more than 90 novel chemical environments. In addition, we subjected the populations to whole genome population sequencing. Although populations with higher mutation rates accumulated greater genetic diversity, this diversity conveyed benefits only for modestly increased mutation rates, where populations adapted faster and also thrived better than their ancestors in some novel environments. In contrast, some populations at the highest mutation rates showed reduced adaptation during evolution, and failed to thrive in all of the 90 alternative environments. In addition, they experienced a dramatic decrease in mutation rate. Our work demonstrates that the mutation rate changes the global balance between deleterious and beneficial mutational effects on fitness. In contrast to most theoretical models, our experiments suggest that this tipping point already occurs at the modest mutation rates that are found in the wild. PMID:29702649

  12. Brote causado por Escherichia coli en Chalco, México

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    Cortés-Ortiz Iliana Alejandra

    2002-01-01

    Full Text Available Objetivo. Identificar el agente causal del brote de diarrea asociado con el desbordamiento del canal de aguas negras en Chalco. Material y métodos. Estudio retrospectivo y transversal, efectuado en el Instituto de Diagnóstico y Referencia Epidemiológicos (InDRE, de la Secretaría de Salud, con 1 550 hisopos rectales para el aislamiento e identificación bioquímica de V. cholerae y enterobacterias, obtenidos de la población del Valle de Chalco, que presentó diarrea y vómito durante el desastre natural acontecido el 31 de mayo de 2000. El análisis de los resultados se efectuó por la diferencia entre las proporciones de dos poblaciones (prueba de Ji cuadrada. Las cepas de E. coli se hibridaron por "colony blot" para los grupos ETEC, EIEC, EPEC y EHEC. Resultados. El 0.45% correspondió a Salmonella: S. agona, S. infantis, S. enteritidis, S. muenchen, S. typhimurium; 0.06% a Shigella flexneri 3a, y 76.6% a E. coli: 62.2% a ETEC (44.6 % con LT, 11.2% con ST, y 44.1% con ambas sondas, 0.84% a EIEC (sonda ial, 0.84% a EPEC (sonda bundle-forming pilus BFP, 0.08% a E. coli enterohemorrágica no-O157:H7 (sonda pCVD419, y 36.02% no hibridó. No se encontró asociación entre E. coli patógena con la edad y género. Conclusiones. Escherichia coli podría ser responsable del brote de diarrea. Es importante conocer el agente etiológico del brote para encaminar las estrategias en el estudio y control sanitario del mismo.

  13. Expression and purification of recombinant hemoglobin in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Chandrasekhar Natarajan

    Full Text Available Recombinant DNA technologies have played a pivotal role in the elucidation of structure-function relationships in hemoglobin (Hb and other globin proteins. Here we describe the development of a plasmid expression system to synthesize recombinant Hbs in Escherichia coli, and we describe a protocol for expressing Hbs with low intrinsic solubilities. Since the α- and β-chain Hbs of different species span a broad range of solubilities, experimental protocols that have been optimized for expressing recombinant human HbA may often prove unsuitable for the recombinant expression of wildtype and mutant Hbs of other species.As a test case for our expression system, we produced recombinant Hbs of the deer mouse (Peromyscus maniculatus, a species that has been the subject of research on mechanisms of Hb adaptation to hypoxia. By experimentally assessing the combined effects of induction temperature, induction time and E. coli expression strain on the solubility of recombinant deer mouse Hbs, we identified combinations of expression conditions that greatly enhanced the yield of recombinant protein and which also increased the efficiency of post-translational modifications.Our protocol should prove useful for the experimental study of recombinant Hbs in many non-human animals. One of the chief advantages of our protocol is that we can express soluble recombinant Hb without co-expressing molecular chaperones, and without the need for additional reconstitution or heme-incorporation steps. Moreover, our plasmid construct contains a combination of unique restriction sites that allows us to produce recombinant Hbs with different α- and β-chain subunit combinations by means of cassette mutagenesis.

  14. Expression and purification of recombinant hemoglobin in Escherichia coli.

    Science.gov (United States)

    Natarajan, Chandrasekhar; Jiang, Xiaoben; Fago, Angela; Weber, Roy E; Moriyama, Hideaki; Storz, Jay F

    2011-01-01

    Recombinant DNA technologies have played a pivotal role in the elucidation of structure-function relationships in hemoglobin (Hb) and other globin proteins. Here we describe the development of a plasmid expression system to synthesize recombinant Hbs in Escherichia coli, and we describe a protocol for expressing Hbs with low intrinsic solubilities. Since the α- and β-chain Hbs of different species span a broad range of solubilities, experimental protocols that have been optimized for expressing recombinant human HbA may often prove unsuitable for the recombinant expression of wildtype and mutant Hbs of other species. As a test case for our expression system, we produced recombinant Hbs of the deer mouse (Peromyscus maniculatus), a species that has been the subject of research on mechanisms of Hb adaptation to hypoxia. By experimentally assessing the combined effects of induction temperature, induction time and E. coli expression strain on the solubility of recombinant deer mouse Hbs, we identified combinations of expression conditions that greatly enhanced the yield of recombinant protein and which also increased the efficiency of post-translational modifications. Our protocol should prove useful for the experimental study of recombinant Hbs in many non-human animals. One of the chief advantages of our protocol is that we can express soluble recombinant Hb without co-expressing molecular chaperones, and without the need for additional reconstitution or heme-incorporation steps. Moreover, our plasmid construct contains a combination of unique restriction sites that allows us to produce recombinant Hbs with different α- and β-chain subunit combinations by means of cassette mutagenesis.

  15. Anaerobic respiration of Escherichia coli in the mouse intestine.

    Science.gov (United States)

    Jones, Shari A; Gibson, Terri; Maltby, Rosalie C; Chowdhury, Fatema Z; Stewart, Valley; Cohen, Paul S; Conway, Tyrrell

    2011-10-01

    The intestine is inhabited by a large microbial community consisting primarily of anaerobes and, to a lesser extent, facultative anaerobes, such as Escherichia coli, which we have shown requires aerobic respiration to compete successfully in the mouse intestine (S. A. Jones et al., Infect. Immun. 75:4891-4899, 2007). If facultative anaerobes efficiently lower oxygen availability in the intestine, then their sustained growth must also depend on anaerobic metabolism. In support of this idea, mutants lacking nitrate reductase or fumarate reductase have extreme colonization defects. Here, we further explore the role of anaerobic respiration in colonization using the streptomycin-treated mouse model. We found that respiratory electron flow is primarily via the naphthoquinones, which pass electrons to cytochrome bd oxidase and the anaerobic terminal reductases. We found that E. coli uses nitrate and fumarate in the intestine, but not nitrite, dimethyl sulfoxide, or trimethylamine N-oxide. Competitive colonizations revealed that cytochrome bd oxidase is more advantageous than nitrate reductase or fumarate reductase. Strains lacking nitrate reductase outcompeted fumarate reductase mutants once the nitrate concentration in cecal mucus reached submillimolar levels, indicating that fumarate is the more important anaerobic electron acceptor in the intestine because nitrate is limiting. Since nitrate is highest in the absence of E. coli, we conclude that E. coli is the only bacterium in the streptomycin-treated mouse large intestine that respires nitrate. Lastly, we demonstrated that a mutant lacking the NarXL regulator (activator of the NarG system), but not a mutant lacking the NarP-NarQ regulator, has a colonization defect, consistent with the advantage provided by NarG. The emerging picture is one in which gene regulation is tuned to balance expression of the terminal reductases that E. coli uses to maximize its competitiveness and achieve the highest possible population in

  16. No evidence for a bovine mastitis Escherichia coli pathotype.

    Science.gov (United States)

    Leimbach, Andreas; Poehlein, Anja; Vollmers, John; Görlich, Dennis; Daniel, Rolf; Dobrindt, Ulrich

    2017-05-08

    Escherichia coli bovine mastitis is a disease of significant economic importance in the dairy industry. Molecular characterization of mastitis-associated E. coli (MAEC) did not result in the identification of common traits. Nevertheless, a mammary pathogenic E. coli (MPEC) pathotype has been proposed suggesting virulence traits that differentiate MAEC from commensal E. coli. The present study was designed to investigate the MPEC pathotype hypothesis by comparing the genomes of MAEC and commensal bovine E. coli. We sequenced the genomes of eight E. coli isolated from bovine mastitis cases and six fecal commensal isolates from udder-healthy cows. We analyzed the phylogenetic history of bovine E. coli genomes by supplementing this strain panel with eleven bovine-associated E. coli from public databases. The majority of the isolates originate from phylogroups A and B1, but neither MAEC nor commensal strains could be unambiguously distinguished by phylogenetic lineage. The gene content of both MAEC and commensal strains is highly diverse and dominated by their phylogenetic background. Although individual strains carry some typical E. coli virulence-associated genes, no traits important for pathogenicity could be specifically attributed to MAEC. Instead, both commensal strains and MAEC have very few gene families enriched in either pathotype. Only the aerobactin siderophore gene cluster was enriched in commensal E. coli within our strain panel. This is the first characterization of a phylogenetically diverse strain panel including several MAEC and commensal isolates. With our comparative genomics approach we could not confirm previous studies that argue for a positive selection of specific traits enabling MAEC to elicit bovine mastitis. Instead, MAEC are facultative and opportunistic pathogens recruited from the highly diverse bovine gastrointestinal microbiota. Virulence-associated genes implicated in mastitis are a by-product of commensalism with the primary function

  17. Binding of collagens to an enterotoxigenic strain of Escherichia coli

    International Nuclear Information System (INIS)

    Visai, L.; Speziale, P.; Bozzini, S.

    1990-01-01

    An enterotoxigenic strain of Escherichia coli, B34289c, has been shown to bind the N-terminal region of fibronectin with high affinity. We now report that this strain also binds collagen. The binding of 125I-labeled type II collagen to bacteria was time dependent and reversible. Bacteria expressed a limited number of collagen receptors (2.2 x 10(4) per cell) and bound collagen with a Kd of 20 nM. All collagen types tested (I to V) as well as all tested cyanogen bromide-generated peptides [alpha 1(I)CB2, alpha 1(I)CB3, alpha 1(I)CB7, alpha 1(I)CB8, and alpha 2(I)CB4] were recognized by bacterial receptors, as demonstrated by the ability of these proteins to inhibit the binding of 125I-labeled collagen to bacteria. Of several unlabeled proteins tested in competition experiments, fibronectin and its N-terminal region strongly inhibited binding of the radiolabeled collagen to E. coli cells. Conversely, collagen competed with an 125I-labeled 28-kilodalton fibronectin fragment for bacterial binding. Collagen bound to bacteria could be displaced by excess amounts of either unlabeled fibronectin or its N-terminal fragment. Similarly, collagen could displace 125I-labeled N-terminal peptide of fibronectin bound to the bacterial cell surface. Bacteria grown at 41 degrees C or in the presence of glucose did not express collagen or fibronectin receptors. These results indicate the presence of specific binding sites for collagen on the surface of E. coli cells and furthermore that the collagen and fibronectin binding sites are located in close proximity, possibly on the same structure

  18. Response of Escherichia coli growth rate to osmotic shock.

    Science.gov (United States)

    Rojas, Enrique; Theriot, Julie A; Huang, Kerwyn Casey

    2014-05-27

    It has long been proposed that turgor pressure plays an essential role during bacterial growth by driving mechanical expansion of the cell wall. This hypothesis is based on analogy to plant cells, for which this mechanism has been established, and on experiments in which the growth rate of bacterial cultures was observed to decrease as the osmolarity of the growth medium was increased. To distinguish the effect of turgor pressure from pressure-independent effects that osmolarity might have on cell growth, we monitored the elongation of single Escherichia coli cells while rapidly changing the osmolarity of their media. By plasmolyzing cells, we found that cell-wall elastic strain did not scale with growth rate, suggesting that pressure does not drive cell-wall expansion. Furthermore, in response to hyper- and hypoosmotic shock, E. coli cells resumed their preshock growth rate and relaxed to their steady-state rate after several minutes, demonstrating that osmolarity modulates growth rate slowly, independently of pressure. Oscillatory hyperosmotic shock revealed that although plasmolysis slowed cell elongation, the cells nevertheless "stored" growth such that once turgor was reestablished the cells elongated to the length that they would have attained had they never been plasmolyzed. Finally, MreB dynamics were unaffected by osmotic shock. These results reveal the simple nature of E. coli cell-wall expansion: that the rate of expansion is determined by the rate of peptidoglycan insertion and insertion is not directly dependent on turgor pressure, but that pressure does play a basic role whereby it enables full extension of recently inserted peptidoglycan.

  19. Role of Maltose Enzymes in Glycogen Synthesis by Escherichia coli▿

    Science.gov (United States)

    Park, Jong-Tae; Shim, Jae-Hoon; Tran, Phuong Lan; Hong, In-Hee; Yong, Hwan-Ung; Oktavina, Ershita Fitria; Nguyen, Hai Dang; Kim, Jung-Wan; Lee, Tae Soo; Park, Sung-Hoon; Boos, Winfried; Park, Kwan-Hwa

    2011-01-01

    Mutants with deletion mutations in the glg and mal gene clusters of Escherichia coli MC4100 were used to gain insight into glycogen and maltodextrin metabolism. Glycogen content, molecular mass, and branch chain distribution were analyzed in the wild type and in ΔmalP (encoding maltodextrin phosphorylase), ΔmalQ (encoding amylomaltase), ΔglgA (encoding glycogen synthase), and ΔglgA ΔmalP derivatives. The wild type showed increasing amounts of glycogen when grown on glucose, maltose, or maltodextrin. When strains were grown on maltose, the glycogen content was 20 times higher in the ΔmalP strain (0.97 mg/mg protein) than in the wild type (0.05 mg/mg protein). When strains were grown on glucose, the ΔmalP strain and the wild type had similar glycogen contents (0.04 mg/mg and 0.03 mg/mg protein, respectively). The ΔmalQ mutant did not grow on maltose but showed wild-type amounts of glycogen when grown on glucose, demonstrating the exclusive function of GlgA for glycogen synthesis in the absence of maltose metabolism. No glycogen was found in the ΔglgA and ΔglgA ΔmalP strains grown on glucose, but substantial amounts (0.18 and 1.0 mg/mg protein, respectively) were found when they were grown on maltodextrin. This demonstrates that the action of MalQ on maltose or maltodextrin can lead to the formation of glycogen and that MalP controls (inhibits) this pathway. In vitro, MalQ in the presence of GlgB (a branching enzyme) was able to form glycogen from maltose or linear maltodextrins. We propose a model of maltodextrin utilization for the formation of glycogen in the absence of glycogen synthase. PMID:21421758

  20. Mutations in Escherichia coli that effect sensitivity to oxygen

    International Nuclear Information System (INIS)

    Jamison, C.S.; Adler, H.I.

    1987-01-01

    Fifteen oxygen-sensitive (Oxy/sup s/) mutants of Escherichia coli were isolated after exposure to UV light. The mutants did not form macroscopic colonies when plated aerobically. They did form macroscopic colonies anaerobically. Oxygen, introduced during log phase, inhibited the growth of liquid cultures. The degree of inhibition was used to separate the mutants into three classes. Class I mutants did not grow after exposure to oxygen. Class II mutants were able to grow, but at a reduced rate and to a reduced final titer, when compared with the wild-type parent. Class III mutants formed filaments in response to oxygen. Genetic experiments indicated that the mutations map to six different chromosomal regions. The results of enzymatic assays indicated that 7 of the 10 class I mutants have low levels of catalase, peroxidase, superoxide dismutase, and respiratory enzymes when compared with the wild-type parent. Mutations in five of the seven class I mutants which have the low enzyme activities mapped within the region 8 to 13.5 min. P1 transduction data indicated that mutations in three of these five mutants, Oxy/sup s/-6, Oxy/sup s/-14, and Oxy/sup s/-17, mapped to 8.4 min. The correlation of low enzyme levels and mapping data suggest that a single gene may regulate several enzymes in response to oxygen. The remaining three class I mutants had wild-type levels of catalase, peroxidase, and superoxide dismutase, but decreased respiratory activity. The class II and III mutants had enzyme activities similar to those of the wild-type parent

  1. Succinic acid production by escherichia coli under anaerobic fermentation

    International Nuclear Information System (INIS)

    El Shafey, H.M.; Meleigy, S.A.

    2009-01-01

    The effect of alteration of growth conditions, addition of different sodium salts, and irradiation by gamma rays on succinic acid production by E. coli was studied. Twenty one isolates were obtained from buffalo's rumen, and anaerobic screening of the isolated bacterial strains showed the abilities of seventeen strains to produce succinic acid. The two bacterial strains having highest succinic acid production were identified as escherichia coli SP9 and SP16, and were selected for further studies. Results showed that growth conditions yielded highest succinic acid production for the two isolates were: 72 hours incubation, 37 degree c incubation temperature, initial ph of the fermentation medium 6.0,and 3% (v/v)inoculum size. Addition of 5 mm of nine different sodium salts to the fermentation medium showed stimulating effect on succinic acid production of the nine tried sodium salts, sodium carbonate was found to have the highest enhancing effect, especially if used at 15 mm concentration. Gamma irradiation doses tried were in the range of (0.25-1.50 kGy). An enhancing effect on succinic acid production was shown in the range of 0.25-0.75 kGy with a maximal production at 0.75 kGy (giving 8.36% increase) for e.coli SP9, and in the range of 0.25-1.00 kGy with a maximal production at 1.0 kGy (7.60% increase) for e.coli SP16. higher gamma doses led to a decrease in the enhancing effect. An overall increase in the succinic acid yield of 79.45% and 94.26% for e. coli SP9 and SP16, respectively, was achieved in implicating all optimized factors for succinic acid production in one time

  2. Pyranopterin Coordination Controls Molybdenum Electrochemistry in Escherichia coli Nitrate Reductase.

    Science.gov (United States)

    Wu, Sheng-Yi; Rothery, Richard A; Weiner, Joel H

    2015-10-09

    We test the hypothesis that pyranopterin (PPT) coordination plays a critical role in defining molybdenum active site redox chemistry and reactivity in the mononuclear molybdoenzymes. The molybdenum atom of Escherichia coli nitrate reductase A (NarGHI) is coordinated by two PPT-dithiolene chelates that are defined as proximal and distal based on their proximity to a [4Fe-4S] cluster known as FS0. We examined variants of two sets of residues involved in PPT coordination: (i) those interacting directly or indirectly with the pyran oxygen of the bicyclic distal PPT (NarG-Ser(719), NarG-His(1163), and NarG-His(1184)); and (ii) those involved in bridging the two PPTs and stabilizing the oxidation state of the proximal PPT (NarG-His(1092) and NarG-His(1098)). A S719A variant has essentially no effect on the overall Mo(VI/IV) reduction potential, whereas the H1163A and H1184A variants elicit large effects (ΔEm values of -88 and -36 mV, respectively). Ala variants of His(1092) and His(1098) also elicit large ΔEm values of -143 and -101 mV, respectively. An Arg variant of His(1092) elicits a small ΔEm of +18 mV on the Mo(VI/IV) reduction potential. There is a linear correlation between the molybdenum Em value and both enzyme activity and the ability to support anaerobic respiratory growth on nitrate. These data support a non-innocent role for the PPT moieties in controlling active site metal redox chemistry and catalysis. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Pyranopterin Coordination Controls Molybdenum Electrochemistry in Escherichia coli Nitrate Reductase*

    Science.gov (United States)

    Wu, Sheng-Yi; Rothery, Richard A.; Weiner, Joel H.

    2015-01-01

    We test the hypothesis that pyranopterin (PPT) coordination plays a critical role in defining molybdenum active site redox chemistry and reactivity in the mononuclear molybdoenzymes. The molybdenum atom of Escherichia coli nitrate reductase A (NarGHI) is coordinated by two PPT-dithiolene chelates that are defined as proximal and distal based on their proximity to a [4Fe-4S] cluster known as FS0. We examined variants of two sets of residues involved in PPT coordination: (i) those interacting directly or indirectly with the pyran oxygen of the bicyclic distal PPT (NarG-Ser719, NarG-His1163, and NarG-His1184); and (ii) those involved in bridging the two PPTs and stabilizing the oxidation state of the proximal PPT (NarG-His1092 and NarG-His1098). A S719A variant has essentially no effect on the overall Mo(VI/IV) reduction potential, whereas the H1163A and H1184A variants elicit large effects (ΔEm values of −88 and −36 mV, respectively). Ala variants of His1092 and His1098 also elicit large ΔEm values of −143 and −101 mV, respectively. An Arg variant of His1092 elicits a small ΔEm of +18 mV on the Mo(VI/IV) reduction potential. There is a linear correlation between the molybdenum Em value and both enzyme activity and the ability to support anaerobic respiratory growth on nitrate. These data support a non-innocent role for the PPT moieties in controlling active site metal redox chemistry and catalysis. PMID:26297003

  4. Deactivation of Escherichia coli by the plasma needle

    Energy Technology Data Exchange (ETDEWEB)

    Sladek, R E J; Stoffels, E [Department of Biomedical Engineering, Eindhoven University of Technology, PO Box 513, 5600 MB Eindhoven (Netherlands)

    2005-06-07

    In this paper we present a parameter study on deactivation of Escherichia coli (E. coli) by means of a non-thermal plasma (plasma needle). The plasma needle is a small-sized (1 mm) atmospheric glow sustained by radio-frequency excitation. This plasma will be used to disinfect heat-sensitive objects; one of the intended applications is in vivo deactivation of dental bacteria: destruction of plaque and treatment of caries. We use E. coli films plated on agar dishes as a model system to optimize the conditions for bacterial destruction. Plasma power, treatment time and needle-to-sample distance are varied. Plasma treatment of E. coli films results in formation of a bacteria-free void with a size up to 12 mm. 10{sup 4}-10{sup 5} colony forming units are already destroyed after 10 s of treatment. Prolongation of treatment time and usage of high powers do not significantly improve the destruction efficiency: short exposure at low plasma power is sufficient. Furthermore, we study the effects of temperature increase on the survival of E. coli and compare it with thermal effects of the plasma. The population of E. coli heated in a warm water bath starts to decrease at temperatures above 40 deg. C. Sample temperature during plasma treatment has been monitored. The temperature can reach up to 60 deg. C at high plasma powers and short needle-to-sample distances. However, thermal effects cannot account for bacterial destruction at low power conditions. For safe and efficient in vivo disinfection, the sample temperature should be kept low. Thus, plasma power and treatment time should not exceed 150 mW and 60 s, respectively.

  5. ANALISIS KANDUNGAN BORAKS DAN Escherichia coli PADA JAJANAN BAKSO SAPI YANG DIPERDAGANGKAN DI KOTA BANJARBARU

    Directory of Open Access Journals (Sweden)

    Nur Rahmi

    2016-09-01

    Full Text Available This study aims to determine how the content of borax and Escherichia coli on meatballs snacks and the factors that affect the food security of meatballs snacks by using Easy Method of Borax Test and Method of Most Probable Number (MPN for Escherichia coli bacteria contamination. This research was conducted in Banjarbaru on 5 villages, and sampling technique used is stratified sampling. The results of the study showed that from 32 samples taken from five village location, it was not identified any borax based on PERMENKES No. 033 of 2012, while for the examination of Escherichia coli, there are 14 samples of meatballs (43.75% which were eligible, and 18 samples of meatballs (56.25% which containEscherichia coli ranges from 3.6 to 62 CFU /g or not meeting the criteria of ISO 7388: 2009. The factor that might not trigger the addition of borax is that the traders have a good knowledge and attitude toward borax which regarded as a toxic substance and can be harmful to health. Factors that cause microbial contamination of Escherichia coli on meatballs snacks is the lack of food hygiene and sanitation in the food processing, cooked food storage, transport, serving, sanitation facilities, and personnel handlers compared with the good supply of foodstuffs and food ingredients storage.

  6. Elimination of Escherichia coli and Salmonella in Clam by Using Zeolite in a Station of Depuration.

    Science.gov (United States)

    Gdoura, Morsi; Sellami, Hanen; Khannous, Lamia; Ketata, Najib; Neila, Idriss Ben; Traore, Al Ibrahim; Chekir, Zouhair; Gdoura, Radhouane

    2017-09-01

      The application of natural zeolite for water and wastewater treatment has been carried out and is still a promising technique in environmental cleaning processes. Natural zeolite can be used to improve the purification process of clams (Ruditapes decussatus). Thus, our study aimed at improving the clam purification system in order to reduce Escherichia coli and eliminate Salmonella in samples artificially contaminated with this bacterium using a natural zeolite to replace the biological filter. The results showed that zeolite used in a depuration system improved the clam purification process. Moreover, natural zeolite exhibited high performance in the adsorption of bacteria and allowed to reduce the Escherichia coli abundance in 24 h, thus ensuring purified clams conformity with the ISO 16649-3 standard. These results indicate the beneficial effects of using zeolite in the adsorption of bacteria and the reduction in the abundance of Escherichia coli and set the Salmonella from marine organisms.

  7. D10 value determination for Escherichia coli O157:H7 in different cultivations

    International Nuclear Information System (INIS)

    Oliveira, Sergio Eduardo M. de; Pires, Luis Fernando G.; Vital, Helio de C.

    2002-01-01

    Escherichia coli serum type O157:H7 is a highly pathogenic bacterium. Inside the human body, that microorganism causes a disease that leads to bloody diarrhea, stoppage of kidney functions and clots in the brain. That type of infection has been related to the consumption of different varieties of foods, mainly meat and other products of animal origin. Irradiation is an efficient method for elimination of pathogenic and spoiling microorganisms in foods. Thus, this work investigates the use of gamma irradiation for elimination of Escherichia coli O157:H7. For that purpose, inoculated samples in trypticase soy broth and saline solution 0,85% media were exposed to several gamma radiation doses. Counting the number of surviving bacteria yielded the following D 10 values for Escherichia coli O157:H7: 98±7 Gy, in trypticase soy broth and 49±4 Gy in saline solution 0,85% medium. (author)

  8. The propagation of Escherichia Coli and of conservative tracers. A comparison

    International Nuclear Information System (INIS)

    Alexander, I.; Seiler, K.P.

    1982-01-01

    The propagation of Escherichia Coli (ATCC 11229, Gelsenkirchen) is compared with that of conservative tracers in groundwater. The experiments were performed with injection quantities of 10 7 , 10 8 , 10 10 and 10 11 of Escherichia Coli. Both, bacteria and conservative tracers pass their maximum at the same instant in the observation gauges. With injection quantities of more than 10 8 , the propagation of the Escherichia Coli sets in at the same time as it begins with the dyes. When the quantities range below 10 8 , the propagation begins after that of conservative tracers, because Coli bacteria were measured with a lower degree of detecting sensitivity than the tracers. With Coli injection quantities ranging above 10 10 , an increased filtering of these bacteria can be observed. Coli bacteria propagate more laterally than conservative tracers, however it could not be proved that this lateral propagation depends on the bacteria concentration. (orig.) [de

  9. The antibacterial effect of four mouthwashes against streptococcus mutans and escherichia coli.

    Science.gov (United States)

    Ghapanchi, Janan; Lavaee, Fatemeh; Moattari, Afagh; Shakib, Mahmood

    2015-04-01

    To evaluate the antimicrobial properties of several mouthwash concentrations on oral Streptococcus mutans and Escherichia coli. The study was conducted at Shiraz Medicine School in 2011. Serial dilutions of Chlorohexidin, Oral B and Persica and Irsha (2,4,8,16,64,128) were prepared in Muller-Hinton media. Minimum inhibitory concentration was visually determined and defined as the lowest concentration of each oral washing which inhibited > 95% growth reduction compared to the growth control well. Chlorhexidine, Oral B and Irsha mouthwash inhibited Streptococcus mutans even with diluted concentrations. Also, Chlorhexidine and Oral B prohibited Escherichia coli with different potencies. But Persica had no antimicrobial activity against either Escherichia coli or Streptococcus mutans. Chlorhexidine, Irsha, and Oral B mouthwashes can be used for antimicrobial effects, especially on Streptococcus mutans. This chemical activity of mouthwashes is an adjuvant for mechanical removing of plaque. However, the antimicrobial effect of Persicaremains controversial.

  10. Bacteraemia Caused by Escherichia Coli in Cancer Patients at a Specialist Center in Pakistan

    International Nuclear Information System (INIS)

    Parveen, A.; Sultan, F.; Saleem, S.; Nazeer, S. H.; Raza, A.; Zafar, W.; Nizamuddin, S.; Mahboob, A.

    2015-01-01

    Objective: To analyse the antimicrobial susceptibility patterns of Escherichia coli bacteraemia among cancer patients, and to assess the risk factors and outcomes of multidrug-resistant Escherichia coli bacteraemia. Methods: The retrospective study was conducted at Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore, and comprised medical records of patients with Escherichia coli bacteraemia presenting between December 2012 and November 2013. Multivariable logistic regression analyses were used to determine the factors associated with the development and 30-day mortality of multidrug-resistant Escherichia coli bacteraemia. Results: Out of 1603 episodes of bacteraemia, 227(35.6 percent) were caused by E.coli, of which 98(43.2 percent) were multidrug-resistant. In multivariable analysis, age less than 18 years (adjusted odds ratio 3.92; 95 percent confidence interval 1.43-10.68), presence of central venous catheter (adjusted odds ratio 2.12; 95 percent confidence interval 1.04-4.33) and exposure to piperacillin/tazobactam within 90 days prior to infection (adjusted odds ratio 2.37; 95 percent confidence interval 1.15-4.86) were identified as independent risk factors for acquisition of multidrug-resistant Escherichia coli bacteraemia. The overall 30 day mortality rate was 35.2 percent (80/227). Risk factors for mortality were intensive care unit admission (adjusted odds ratio 3.95; 95 percent confidence interval 1.79-8.71) and profound neutropenia (adjusted odds ratio 4.03; 95 percent confidence interval 1.55-10.49). Conclusion: Bloodstream infections with multidrug-resistant Escherichia coli were common in cancer patients. However it was not a predictor of mortality. (author)

  11. Antibiofilm Effects of Lactobacilli against Ciprofloxacin-Resistant Uropathogenic Escherichia coli strains in Pasteurized Milk

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    Mahsa Yeganeh

    2017-11-01

    Full Text Available  Background and Objective: Uropathogenic Escherichia coli-induced urinary tract infections are the most common uropathogenic Escherichia coli etiological agent. In addition, most of biofilms created by these bacteria can be regarded as a serious problem in the food industry. Foodborne diseases have always been considered an emerging public health concern throughout the world. Many outbreaks have been found to be associated with biofilms. Thus, the aim of the present study is to investigate the anti-adhesive effects of lactic acid bacteria against strains of Ciprofloxacin-Resistant Uropathogenic Escherichia coli using microbial techniques in pasteurized milk.Material and Methods: In this study, strains of Lactobacillus plantarum, Lactobacillus casei and Lactobacillus acidophilus were provided from Pasteur Institute of Iran. Twenty strains of Uropathogenic Escherichia coli-Induced Urinary Tract Infections were isolated from patients with urinary tract infection in Shahid Labbafinejad hospital of Iran. Eight strains with ability of biofilm formation were selected for microbial tests. All of these eight strains were resistant to ciprofloxacin. Disk diffusion method was used to assess the susceptibility of all isolates to the ten common antibiotics. Eight samples of Uropathogenic Escherichia coli were inoculated in pasteurized milk. The microtitre plate 100 method was used to detect anti-adhesive activity of lactobacilli supernatant.Results and Conclusion: Results showed that the eight human isolates were resistant to antibiotics. Isolate of number 4 was the most susceptible strains to antibiofilm effects of lactobacilli in the pasteurized milk. The anti-adhesive effects of lactobacilli on Uropathogenic were confirmed in all microbial tests. In this study, Lactobacillus plantarum revealed the highest inhibitory activity against Uropathogenic Escherichia coli 4 strain with inhibition zones of 42 mm. This strain was reported as a proper probiotic

  12. Hydrogenase-3 contributes to anaerobic acid resistance of Escherichia coli.

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    Ken Noguchi

    Full Text Available BACKGROUND: Hydrogen production by fermenting bacteria such as Escherichia coli offers a potential source of hydrogen biofuel. Because H(2 production involves consumption of 2H(+, hydrogenase expression is likely to involve pH response and regulation. Hydrogenase consumption of protons in E. coli has been implicated in acid resistance, the ability to survive exposure to acid levels (pH 2-2.5 that are three pH units lower than the pH limit of growth (pH 5-6. Enhanced survival in acid enables a larger infective inoculum to pass through the stomach and colonize the intestine. Most acid resistance mechanisms have been defined using aerobic cultures, but the use of anaerobic cultures will reveal novel acid resistance mechanisms. METHODS AND PRINCIPAL FINDINGS: We analyzed the pH regulation of bacterial hydrogenases in live cultures of E. coli K-12 W3110. During anaerobic growth in the range of pH 5 to 6.5, E. coli expresses three hydrogenase isoenzymes that reversibly oxidize H(2 to 2H(+. Anoxic conditions were used to determine which of the hydrogenase complexes contribute to acid resistance, measured as the survival of cultures grown at pH 5.5 without aeration and exposed for 2 hours at pH 2 or at pH 2.5. Survival of all strains in extreme acid was significantly lower in low oxygen than for aerated cultures. Deletion of hyc (Hyd-3 decreased anoxic acid survival 3-fold at pH 2.5, and 20-fold at pH 2, but had no effect on acid survival with aeration. Deletion of hyb (Hyd-2 did not significantly affect acid survival. The pH-dependence of H(2 production and consumption was tested using a H(2-specific Clark-type electrode. Hyd-3-dependent H(2 production was increased 70-fold from pH 6.5 to 5.5, whereas Hyd-2-dependent H(2 consumption was maximal at alkaline pH. H(2 production, was unaffected by a shift in external or internal pH. H(2 production was associated with hycE expression levels as a function of external pH. CONCLUSIONS: Anaerobic growing

  13. Hydrogenase-3 contributes to anaerobic acid resistance of Escherichia coli.

    Science.gov (United States)

    Noguchi, Ken; Riggins, Daniel P; Eldahan, Khalid C; Kitko, Ryan D; Slonczewski, Joan L

    2010-04-12

    Hydrogen production by fermenting bacteria such as Escherichia coli offers a potential source of hydrogen biofuel. Because H(2) production involves consumption of 2H(+), hydrogenase expression is likely to involve pH response and regulation. Hydrogenase consumption of protons in E. coli has been implicated in acid resistance, the ability to survive exposure to acid levels (pH 2-2.5) that are three pH units lower than the pH limit of growth (pH 5-6). Enhanced survival in acid enables a larger infective inoculum to pass through the stomach and colonize the intestine. Most acid resistance mechanisms have been defined using aerobic cultures, but the use of anaerobic cultures will reveal novel acid resistance mechanisms. We analyzed the pH regulation of bacterial hydrogenases in live cultures of E. coli K-12 W3110. During anaerobic growth in the range of pH 5 to 6.5, E. coli expresses three hydrogenase isoenzymes that reversibly oxidize H(2) to 2H(+). Anoxic conditions were used to determine which of the hydrogenase complexes contribute to acid resistance, measured as the survival of cultures grown at pH 5.5 without aeration and exposed for 2 hours at pH 2 or at pH 2.5. Survival of all strains in extreme acid was significantly lower in low oxygen than for aerated cultures. Deletion of hyc (Hyd-3) decreased anoxic acid survival 3-fold at pH 2.5, and 20-fold at pH 2, but had no effect on acid survival with aeration. Deletion of hyb (Hyd-2) did not significantly affect acid survival. The pH-dependence of H(2) production and consumption was tested using a H(2)-specific Clark-type electrode. Hyd-3-dependent H(2) production was increased 70-fold from pH 6.5 to 5.5, whereas Hyd-2-dependent H(2) consumption was maximal at alkaline pH. H(2) production, was unaffected by a shift in external or internal pH. H(2) production was associated with hycE expression levels as a function of external pH. Anaerobic growing cultures of E. coli generate H(2) via Hyd-3 at low external pH, and

  14. Fluorometric determination of ethidium bromide efflux kinetics in Escherichia coli

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    Monteiro Gabriel A

    2009-10-01

    Full Text Available Abstract Background Efflux pump activity has been associated with multidrug resistance phenotypes in bacteria, compromising the effectiveness of antimicrobial therapy. The development of methods for the early detection and quantification of drug transport across the bacterial cell wall is a tool essential to understand and overcome this type of drug resistance mechanism. This approach was developed to study the transport of the efflux pump substrate ethidium bromide (EtBr across the cell envelope of Escherichia coli K-12 and derivatives, differing in the expression of their efflux systems. Results EtBr transport across the cell envelope of E. coli K-12 and derivatives was analysed by a semi-automated fluorometric method. Accumulation and efflux of EtBr was studied under limiting energy supply (absence of glucose and low temperature and in the presence and absence of the efflux pump inhibitor, chlorpromazine. The bulk fluorescence variations were also observed by single-cell flow cytometry analysis, revealing that once inside the cells, leakage of EtBr does not occur and that efflux is mediated by active transport. The importance of AcrAB-TolC, the main efflux system of E. coli, in the extrusion of EtBr was evidenced by comparing strains with different levels of AcrAB expression. An experimental model was developed to describe the transport kinetics in the three strains. The model integrates passive entry (influx and active efflux of EtBr, and discriminates different degrees of efflux between the studied strains that vary in the activity of their efflux systems, as evident from the calculated efflux rates: = 0.0173 ± 0.0057 min-1; = 0.0106 ± 0.0033 min-1; and = 0.0230 ± 0.0075 min-1. Conclusion The combined use of a semi-automated fluorometric method and an experimental model allowed quantifying EtBr transport in E. coli strains that differ in their overall efflux activity. This methodology can be used for the early detection of differences in

  15. Rapid Identification of Different Escherichia coli Sequence Type 131 Clades.

    Science.gov (United States)

    Matsumura, Yasufumi; Pitout, Johann D D; Peirano, Gisele; DeVinney, Rebekah; Noguchi, Taro; Yamamoto, Masaki; Gomi, Ryota; Matsuda, Tomonari; Nakano, Satoshi; Nagao, Miki; Tanaka, Michio; Ichiyama, Satoshi

    2017-08-01

    Escherichia coli sequence type 131 (ST131) is a pandemic clonal lineage that is responsible for the global increase in fluoroquinolone resistance and extended-spectrum-β-lactamase (ESBL) producers. The members of ST131 clade C, especially subclades C2 and C1-M27, are associated with ESBLs. We developed a multiplex conventional PCR assay with the ability to detect all ST131 clades (A, B, and C), as well as C subclades (C1-M27, C1-nM27 [C1-non-M27], and C2). To validate the assay, we used 80 ST131 global isolates that had been fully sequenced. We then used the assay to define the prevalence of each clade in two Japanese collections consisting of 460 ESBL-producing E. coli ST131 (2001-12) and 329 E. coli isolates from extraintestinal sites (ExPEC) (2014). The assay correctly identified the different clades in all 80 global isolates: clades A ( n = 12), B ( n = 12), and C, including subclades C1-M27 ( n = 16), C1-nM27 ( n = 20), C2 ( n = 17), and other C ( n = 3). The assay also detected all 565 ST131 isolates in both collections without any false positives. Isolates from clades A ( n = 54), B ( n = 23), and C ( n = 483) corresponded to the O serotypes and the fimH types of O16-H41, O25b-H22, and O25b-H30, respectively. Of the 483 clade C isolates, C1-M27 was the most common subclade (36%), followed by C1-nM27 (32%) and C2 (15%). The C1-M27 subclade with bla CTX-M-27 became especially prominent after 2009. Our novel multiplex PCR assay revealed the predominance of the C1-M27 subclade in recent Japanese ESBL-producing E. coli isolates and is a promising tool for epidemiological studies of ST131. Copyright © 2017 American Society for Microbiology.

  16. Secretion of d-alanine by Escherichia coli.

    Science.gov (United States)

    Katsube, Satoshi; Sato, Kazuki; Ando, Tasuke; Isogai, Emiko; Yoneyama, Hiroshi

    2016-07-01

    Escherichia coli has an l-alanine export system that protects the cells from toxic accumulation of intracellular l-alanine in the presence of l-alanyl-l-alanine (l-Ala-l-Ala). When a DadA-deficient strain was incubated with 6.0 mM l-Ala-l-Ala, we detected l-alanine and d-alanine using high-performance liquid chromatography (HPLC) analysis at a level of 7.0 mM and 3.0 mM, respectively, after 48 h incubation. Treatment of the culture supernatant with d-amino acid oxidase resulted in the disappearance of a signal corresponding to d-alanine. Additionally, the culture supernatant enabled a d-alanine auxotroph to grow without d-alanine supplementation, confirming that the signal detected by HPLC was authentic d-alanine. Upon introduction of an expression vector harbouring the alanine racemase genes, alr or dadX, the extracellular level of d-alanine increased to 11.5 mM and 8.5 mM, respectively, under similar conditions, suggesting that increased metabolic flow from l-alanine to d-alanine enhanced d-alanine secretion. When high-density DadA-deficient cells preloaded with l-Ala-l-Ala were treated with 20 µM carbonyl cyanide m-chlorophenyl hydrazone (CCCP), secretion of both l-alanine and d-alanine was enhanced ~twofold compared with that in cells without CCCP treatment. In contrast, the ATPase inhibitor dicyclohexylcarbodiimide did not exert such an effect on the l-alanine and d-alanine secretion. Furthermore, inverted membrane vesicles prepared from DadA-deficient cells lacking the l-alanine exporter AlaE accumulated [3H]D-alanine in an energy-dependent manner. This energy-dependent accumulation of [3H]D-alanine was strongly inhibited by CCCP. These results indicate that E. coli has a transport system(s) that exports d-alanine and that this function is most likely modulated by proton electrochemical potential.

  17. Structure and mechanisms of Escherichia coli aspartate transcarbamoylase.

    Science.gov (United States)

    Lipscomb, William N; Kantrowitz, Evan R

    2012-03-20

    Enzymes catalyze a particular reaction in cells, but only a few control the rate of this reaction and the metabolic pathway that follows. One specific mechanism for such enzymatic control of a metabolic pathway involves molecular feedback, whereby a metabolite further down the pathway acts at a unique site on the control enzyme to alter its activity allosterically. This regulation may be positive or negative (or both), depending upon the particular system. Another method of enzymatic control involves the cooperative binding of the substrate, which allows a large change in enzyme activity to emanate from only a small change in substrate concentration. Allosteric regulation and homotropic cooperativity are often known to involve significant conformational changes in the structure of the protein. Escherichia coli aspartate transcarbamoylase (ATCase) is the textbook example of an enzyme that regulates a metabolic pathway, namely, pyrimidine nucleotide biosynthesis, by feedback control and by the cooperative binding of the substrate, L-aspartate. The catalytic and regulatory mechanisms of this enzyme have been extensively studied. A series of X-ray crystal structures of the enzyme in the presence and absence of substrates, products, and analogues have provided details, at the molecular level, of the conformational changes that the enzyme undergoes as it shifts between its low-activity, low-affinity form (T state) to its high-activity, high-affinity form (R state). These structural data provide insights into not only how this enzyme catalyzes the reaction between l-aspartate and carbamoyl phosphate to form N-carbamoyl-L-aspartate and inorganic phosphate, but also how the allosteric effectors modulate this activity. In this Account, we summarize studies on the structure of the enzyme and describe how these structural data provide insights into the catalytic and regulatory mechanisms of the enzyme. The ATCase-catalyzed reaction is regulated by nucleotide binding some 60

  18. Development of a Vaccine against Escherichia coli Urinary Tract Infections

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    Harry L. T. Mobley

    2015-12-01

    Full Text Available Urinary tract infection (UTI is the second most common infection in humans after those involving the respiratory tract. This results not only in huge annual economic costs, but in decreased workforce productivity and high patient morbidity. Most infections are caused by uropathogenic Escherichia coli (UPEC. Antibiotic treatment is generally effective for eradication of the infecting strain; however, documentation of increasing antibiotic resistance, allergic reaction to certain pharmaceuticals, alteration of normal gut flora, and failure to prevent recurrent infections represent significant barriers to treatment. As a result, approaches to prevent UTI such as vaccination represent a gap that must be addressed. Our laboratory has made progress toward development of a preventive vaccine against UPEC. The long-term research goal is to prevent UTIs in women with recurrent UTIs. Our objective has been to identify the optimal combination of protective antigens for inclusion in an effective UTI vaccine, optimal adjuvant, optimal dose, and optimal route of delivery. We hypothesized that a multi-subunit vaccine elicits antibody that protects against experimental challenge with UPEC strains. We have systematically identified four antigens that can individually protect experimentally infected mice from colonization of the bladder and/or kidneys by UPEC when administered intranasally with cholera toxin (CT as an adjuvant. To advance the vaccine for utility in humans, we will group the individual antigens, all associated with iron acquisition (IreA, Hma, IutA, FyuA, into an effective combination to establish a multi-subunit vaccine. We demonstrated for all four vaccine antigens that antigen-specific serum IgG represents a strong correlate of protection in vaccinated mice. High antibody titers correlate with low colony forming units (CFUs of UPEC following transurethral challenge of vaccinated mice. However, the contribution of cell-mediated immunity cannot

  19. Production of extracellular fatty acid using engineered Escherichia coli

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    Liu Hui

    2012-04-01

    Full Text Available Abstract Background As an alternative for economic biodiesel production, the microbial production of extracellular fatty acid from renewable resources is receiving more concerns recently, since the separation of fatty acid from microorganism cells is normally involved in a series of energy-intensive steps. Many attempts have been made to construct fatty acid producing strains by targeting genes in the fatty acid biosynthetic pathway, while few studies focused on the cultivation process and the mass transfer kinetics. Results In this study, both strain improvements and cultivation process strategies were applied to increase extracellular fatty acid production by engineered Escherichia coli. Our results showed overexpressing ‘TesA and the deletion of fadL in E. coli BL21 (DE3 improved extracellular fatty acid production, while deletion of fadD didn’t strengthen the extracellular fatty acid production for an undetermined mechanism. Moreover, the cultivation process controls contributed greatly to extracellular fatty acid production with respect to titer, cell growth and productivity by adjusting the temperature, adding ampicillin and employing on-line extraction. Under optimal conditions, the E. coli strain (pACY-‘tesA-ΔfadL produced 4.8 g L−1 extracellular fatty acid, with the specific productivity of 0.02 g h−1 g−1dry cell mass, and the yield of 4.4% on glucose, while the ratios of cell-associated fatty acid versus extracellular fatty acid were kept below 0.5 after 15 h of cultivation. The fatty acids included C12:1, C12:0, C14:1, C14:0, C16:1, C16:0, C18:1, C18:0. The composition was dominated by C14 and C16 saturated and unsaturated fatty acids. Using the strain pACY-‘tesA, similar results appeared under the same culture conditions and the titer was also much higher than that ever reported previously, which suggested that the supposedly superior strain did not necessarily perform best for the efficient production of desired

  20. Postreplication repair gap filling in an Escherichia coli strain deficient in dnaB gene product

    International Nuclear Information System (INIS)

    Johnson, R.C.

    1975-01-01

    Gaps in daughter-strand DNA synthesized after exposure of Escherichia coli E279 to ultraviolet light are filled during reincubation at 30 0 C for 20 min. Escherichia coli E279 is phenotypically DnaB - when incubated at 43 0 C. Cells incubated at 43 0 C were tested for their ability to complete postreplication repair gap filling. It is concluded that the dnaB gene product is essential for postreplication repair gap filling and that the inhibition seen is not initially the result of degradation