Synthesis of P1-(11-phenoxyundecyl)-P2-(2-acetamido-2-deoxy-3-O-α-D-rhamnopyranosyl-α-D-glucopyranosyl) diphosphate and P1-(11-phenoxyundecyl)-P2-(2-acetamido-2-deoxy-3-O-β-D-galactopyranosyl-α-D-galactopyranosyl) diphosphate for the investigation of biosynthesis of O-antigenic polysaccharides in Pseudomonas aeruginosa and Escherichia coli O104.

Science.gov (United States)

Torgov, Vladimir; Danilov, Leonid; Utkina, Natalia; Veselovsky, Vladimir; Brockhausen, Inka

2017-12-01

Two new phenoxyundecyl diphosphate sugars were synthesized for the first time: P 1 -(11-phenoxyundecyl)-P 2 - (2-acetamido-2-deoxy-3-O-α-D-rhamnopyranosyl-α-D-glucopyranosyl) diphosphate and P 1 -(11-phenoxyundecyl)-P 2 -(2-acetamido-2-deoxy-3-O-β-D-galactopyranosyl-α-D-galactopyranosyl) diphosphate to study the third step of biosynthesis of the repeating units of O-antigenic polysaccharides in Pseudomonas aeruginosa and E.coli O104 respectively. Copyright © 2017 Elsevier Ltd. All rights reserved.

Geranylgeranyl diphosphate synthases from Scoparia dulcis and Croton sublyratus. cDNA cloning, functional expression, and conversion to a farnesyl diphosphate synthase.

Science.gov (United States)

Kojima, N; Sitthithaworn, W; Viroonchatapan, E; Suh, D Y; Iwanami, N; Hayashi, T; Sankaw, U

2000-07-01

cDNAs encoding geranylgeranyl diphosphate synthase (GGPPS) of two diterpene producing plants, Scoparia dulcis and Croton sublyratus, were isolated using the homology-based polymerase chain reaction method. Both cloned genes showed high amino acid sequence homology (60-70%) to other plant GGPPSs and contained highly conserved aspartate-rich motifs. The obtained clones were functionally expressed in Escherichia coli and showed sufficient GGPPS activity to catalyze the condensation of farnesyl diphosphate (FPP) and isopentenyl diphosphate to form geranylgeranyl diphosphate. To investigate the factor determining the product chain length of plant GGPPSs, S. dulcis GGPPS mutants in which either the small amino acids at the fourth and fifth positions before the first aspartate-rich motif (FARM) were replaced with aromatic amino acids or in which two additional amino acids in FARM were deleted were constructed. Both mutants behaved like FPPS-like enzymes and almost exclusively produced FPP when dimethylallyl diphosphate was used as a primer substrate, and failed to accept FPP as a primer substrate. These results indicate that both small amino acids at the fourth and fifth positions before FARM and the amino acid insertion in FARM play essential roles in product length determination in plant GGPPSs.

Geranylgeranyl diphosphate synthase from Scoparia dulcis and Croton sublyratus. Plastid localization and conversion to a farnesyl diphosphate synthase by mutagenesis.

Science.gov (United States)

Sitthithaworn, W; Kojima, N; Viroonchatapan, E; Suh, D Y; Iwanami, N; Hayashi, T; Noji, M; Saito, K; Niwa, Y; Sankawa, U

2001-02-01

cDNAs encoding geranylgeranyl diphosphate synthase (GGPPS) of two diterpene-producing plants, Scoparia dulcis and Croton sublyratus, have been isolated using the homology-based polymerase chain reaction (PCR) method. Both clones contained highly conserved aspartate-rich motifs (DDXX(XX)D) and their N-terminal residues exhibited the characteristics of chloroplast targeting sequence. When expressed in Escherichia coli, both the full-length and truncated proteins in which the putative targeting sequence was deleted catalyzed the condensation of farnesyl diphosphate and isopentenyl diphosphate to produce geranylgeranyl diphosphate (GGPP). The structural factors determining the product length in plant GGPPSs were investigated by constructing S. dulcis GGPPS mutants on the basis of sequence comparison with the first aspartate-rich motif (FARM) of plant farnesyl diphosphate synthase. The result indicated that in plant GGPPSs small amino acids, Met and Ser, at the fourth and fifth positions before FARM and Pro and Cys insertion in FARM play essential roles in determination of product length. Further, when a chimeric gene comprised of the putative transit peptide of the S. dulcis GGPPS gene and a green fluorescent protein was introduced into Arabidopsis leaves by particle gun bombardment, the chimeric protein was localized in chloroplasts, indicating that the cloned S. dulcis GGPPS is a chloroplast protein.

Isolation and characterization of farnesyl diphosphate synthase from the cotton boll weevil, Anthonomus grandis.

Science.gov (United States)

Taban, A Huma; Tittiger, Claus; Blomquist, Gary J; Welch, William H

2009-06-01

Farnesyl diphosphate synthase (FPPS) catalyzes the consecutive condensation of two molecules of isopentenyl diphosphate with dimethylallyl diphosphate to form farnesyl diphosphate (FPP). In insects, FPP is used for the synthesis of ubiquinones, dolicols, protein prenyl groups, and juvenile hormone. A full-length cDNA of FPPS was cloned from the cotton boll weevil, Anthonomus grandis (AgFPPS). AgFPPS cDNA consists of 1,835 nucleotides and encodes a protein of 438 amino acids. The deduced amino acid sequence has high similarity to previously isolated insect FPPSs and other known FPPSs. Recombinant AgFPPS expressed in E. coli converted labeled isopentenyl diphosphate in the presence of dimethylallyl diphosphate to FPP. Southern blot analysis indicated the presence of a single copy gene. Using molecular modeling, the three-dimensional structure of coleopteran FPPS was determined and compared to the X-ray crystal structure of avian FPPS. The alpha-helical fold is conserved in AgFPPS and the size of the active site cavity is consistent with the enzyme being a FPPS. (c) 2009 Wiley Periodicals, Inc.

The biosynthetic origin of irregular monoterpenes in Lavandula: isolation and biochemical characterization of a novel cis-prenyl diphosphate synthase gene, lavandulyl diphosphate synthase.

Science.gov (United States)

Demissie, Zerihun A; Erland, Lauren A E; Rheault, Mark R; Mahmoud, Soheil S

2013-03-01

Lavender essential oils are constituted predominantly of regular monoterpenes, for example linalool, 1,8-cineole, and camphor. However, they also contain irregular monoterpenes including lavandulol and lavandulyl acetate. Although the majority of genes responsible for the production of regular monoterpenes in lavenders are now known, enzymes (including lavandulyl diphosphate synthase (LPPS)) catalyzing the biosynthesis of irregular monoterpenes in these plants have not been described. Here, we report the isolation and functional characterization of a novel cis-prenyl diphosphate synthase cDNA, termed Lavandula x intermedia lavandulyl diphosphate synthase (LiLPPS), through a homology-based cloning strategy. The LiLPPS ORF, encoding for a 305-amino acid long protein, was expressed in Escherichia coli, and the recombinant protein was purified by nickel-nitrilotriacetic acid affinity chromatography. The approximately 34.5-kDa bacterially produced protein specifically catalyzed the head-to-middle condensation of two dimethylallyl diphosphate units to LPP in vitro with apparent Km and kcat values of 208 ± 12 μm and 0.1 s(-1), respectively. LiLPPS is a homodimeric enzyme with a sigmoidal saturation curve and Hill coefficient of 2.7, suggesting a positive co-operative interaction among its catalytic sites. LiLPPS could be used to modulate the production of lavandulol and its derivatives in plants through metabolic engineering.

Metabolic engineering of Escherichia coli to produce 2'-fucosyllactose via salvage pathway of guanosine 5'-diphosphate (GDP)-l-fucose.

Science.gov (United States)

Chin, Young-Wook; Seo, Nari; Kim, Jae-Han; Seo, Jin-Ho

2016-11-01

2'-Fucosyllactose (2-FL) is one of the key oligosaccharides in human milk. In the present study, the salvage guanosine 5'-diphosphate (GDP)-l-fucose biosynthetic pathway from fucose was employed in engineered Escherichia coli BL21star(DE3) for efficient production of 2-FL. Introduction of the fkp gene coding for fucokinase/GDP-l-fucose pyrophosphorylase (Fkp) from Bacteroides fragilis and the fucT2 gene encoding α-1,2-fucosyltransferase from Helicobacter pylori allows the engineered E. coli to produce 2-FL from fucose, lactose and glycerol. To enhance the lactose flux to 2-FL production, the attenuated, and deleted mutants of β-galactosidase were employed. Moreover, the 2-FL yield and productivity were further improved by deletion of the fucI-fucK gene cluster coding for fucose isomerase (FucI) and fuculose kinase (FucK). Finally, fed-batch fermentation of engineered E. coli BL21star(DE3) deleting lacZ and fucI-fucK, and expressing fkp and fucT2 resulted in 23.1 g/L of extracellular concentration of 2-FL and 0.39 g/L/h productivity. Biotechnol. Bioeng. 2016;113: 2443-2452. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

The 1.25 Å resolution structure of phosphoribosyl-ATP pyrophosphohydrolase from Mycobacterium tuberculosis

Energy Technology Data Exchange (ETDEWEB)

Javid-Majd, Farah; Yang, Dong [Department of Biochemistry and Biophysics, Texas A& M University, College Station, Texas 77843-2128 (United States); Ioerger, Thomas R. [Department of Computer Science, Texas A& M University, College Station, Texas 77843-2128 (United States); Sacchettini, James C., E-mail: sacchett@tamu.edu [Department of Biochemistry and Biophysics, Texas A& M University, College Station, Texas 77843-2128 (United States)

2008-06-01

The crystal structure of M. tuberculosis phosphoribosyl-ATP pyrophosphohydrolase, the second enzyme in the histidine-biosynthetic pathway, is presented. The structural and inferred functional relationships between M. tuberculosis phosphoribosyl-ATP pyrophosphohydrolase and other members of the nucleoside-triphosphate pyrophosphatase-fold family are described. Phosphoribosyl-ATP pyrophosphohydrolase is the second enzyme in the histidine-biosynthetic pathway, irreversibly hydrolyzing phosphoribosyl-ATP to phosphoribosyl-AMP and pyrophosphate. It is encoded by the hisE gene, which is present as a separate gene in many bacteria and archaea but is fused to hisI in other bacteria, fungi and plants. Because of its essentiality for growth in vitro, HisE is a potential drug target for tuberculosis. The crystal structures of two native (uncomplexed) forms of HisE from Mycobacterium tuberculosis have been determined to resolutions of 1.25 and 1.79 Å. The structure of the apoenzyme reveals that the protein is composed of five α-helices with connecting loops and is a member of the α-helical nucleoside-triphosphate pyrophosphatase superfamily. The biological unit of the protein is a homodimer, with an active site on each subunit composed of residues exclusively from that subunit. A comparison with the Campylobacter jejuni dUTPase active site allowed the identification of putative metal- and substrate-binding sites in HisE, including four conserved glutamate and glutamine residues in the sequence that are consistent with a motif for pyrophosphohydrolase activity. However, significant differences between family members are observed in the loop region between α-helices H1 and H3. The crystal structure of M. tuberculosis HisE provides insights into possible mechanisms of substrate binding and the diversity of the nucleoside-triphosphate pyrophosphatase superfamily.

Metabolic engineering of the Stevia rebaudiana ent-kaurene biosynthetic pathway in recombinant Escherichia coli.

Science.gov (United States)

Kong, Min Kyung; Kang, Hyun-Jun; Kim, Jin Ho; Oh, Soon Hwan; Lee, Pyung Cheon

2015-11-20

The ent-kaurene is a dedicated precursor pool and is responsible for synthesizing natural sweeteners such as steviol glycosides. In this study, to produce ent-kaurene in Escherichia coli, we modularly constructed and expressed two ent-kaurene genes encoding ent-copalyl diphosphate synthase (CPPS) and ent-kaurene synthase (KS) from Stevia rebaudiana known as a typical plant producing steviol glycoside. The CPPS and KS from S. rebaudiana were functionally expressed in a heterologous host E. coli. Furthermore, in order to enhance ent-kaurene production in E. coli, six geranylgeranyl diphosphate synthases (GGPPS) from various microorganisms and eight strains of E. coli as host were compared by measuring ent-kaurene production. The highest ent-kaurene production of approximately 41.1mg/L was demonstrated in E. coli strain MG1655 co-expressing synthetic CPPS-KS module and GGPPS from Rhodobacter sphaeroides. The ent-kaurene production was further increased up to 179.6 mg/L by overexpression of the three key enzymes for isoprenoid precursor, 1-deoxyxylulose-5-phosphate synthase (DXS), farnesyl diphosphate synthase (IspA) and isopentenyl diphosphate isomerase (IDI) from E. coli. Finally, the highest titer of ent-kaurene (578 mg/L) with a specific yield of ent-kaurene of 143.5mg/g dry cell weight was obtained by culturing E. coli strain MG1655 co-expressing the ent-kaurene module, DXS, IDI and IspA in 1L bioreactor containing 20 g/L glycerol. Copyright © 2015 Elsevier B.V. All rights reserved.

Structure of active IspH enzyme from escherichia coli provides mechanistic insights into substrate reduction

KAUST Repository

Grä wert, Tobias; Rohdich, Felix; Span, Lngrid; Backer, Adelbert; Eisenreich, Wolfgang; Eppinger, Jö rg; Groll, Michael

2009-01-01

The terminal step of the non-mevalonate pathway of terpene biosynthesis is catalyzed by IspH (see scheme). In the crystal structure of IspH from E. coli, a bound inorganic diphosphate ligand occupies the position of the diphosphate residue

Monoterpenes in the glandular trichomes of tomato are synthesized from a neryl diphosphate precursor rather than geranyl diphosphate.

Science.gov (United States)

Schilmiller, Anthony L; Schauvinhold, Ines; Larson, Matthew; Xu, Richard; Charbonneau, Amanda L; Schmidt, Adam; Wilkerson, Curtis; Last, Robert L; Pichersky, Eran

2009-06-30

We identified a cis-prenyltransferase gene, neryl diphosphate synthase 1 (NDPS1), that is expressed in cultivated tomato (Solanum lycopersicum) cultivar M82 type VI glandular trichomes and encodes an enzyme that catalyzes the formation of neryl diphosphate from isopentenyl diphosphate and dimethylallyl diphosphate. mRNA for a terpene synthase gene, phellandrene synthase 1 (PHS1), was also identified in these glands. It encodes an enzyme that uses neryl diphosphate to produce beta-phellandrene as the major product as well as a variety of other monoterpenes. The profile of monoterpenes produced by PHS1 is identical with the monoterpenes found in type VI glands. PHS1 and NDPS1 map to chromosome 8, and the presence of a segment of chromosome 8 derived from Solanum pennellii LA0716 causes conversion from the M82 gland monoterpene pattern to that characteristic of LA0716 plants. The data indicate that, contrary to the textbook view of geranyl diphosphate as the "universal" substrate of monoterpene synthases, in tomato glands neryl diphosphate serves as a precursor for the synthesis of monoterpenes.

Transcriptional activation of a geranylgeranyl diphosphate synthase gene, GGPPS2, isolated from Scoparia dulcis by treatment with methyl jasmonate and yeast extract.

Science.gov (United States)

Yamamura, Y; Mizuguchi, Y; Taura, F; Kurosaki, F

2014-10-01

A cDNA clone, designated SdGGPPS2, was isolated from young seedlings of Scoparia dulcis. The putative amino acid sequence of the translate of the gene showed high homology with geranylgeranyl diphosphate synthase (GGPPS) from various plant sources, and the N-terminal residues exhibited the characteristics of chloroplast targeting sequence. An appreciable increase in the transcriptional level of SdGGPPS2 was observed by exposure of the leaf tissues of S. dulcis to methyl jasmonate, yeast extract or Ca(2+) ionophore A23187. In contrast, SdGGPPS1, a homologous GGPPS gene of the plant, showed no or only negligible change in the expression level upon treatment with these stimuli. The truncated protein heterologously expressed in Escherichia coli in which the putative targeting domain was deleted catalyzed the condensation of farnesyl diphosphate and isopentenyl diphosphate to liberate geranylgeranyl diphosphate. These results suggested that SdGGPPS2 plays physiological roles in methyl jasmonate and yeast extract-induced metabolism in the chloroplast of S. dulcis cells.

Functional identification of a Lippia dulcis bornyl diphosphate synthase that contains a duplicated, inhibitory arginine-rich motif.

Science.gov (United States)

Hurd, Matthew C; Kwon, Moonhyuk; Ro, Dae-Kyun

2017-08-26

Lippia dulcis (Aztec sweet herb) contains the potent natural sweetener hernandulcin, a sesquiterpene ketone found in the leaves and flowers. Utilizing the leaves for agricultural application is challenging due to the presence of the bitter-tasting and toxic monoterpene, camphor. To unlock the commercial potential of L. dulcis leaves, the first step of camphor biosynthesis by a bornyl diphosphate synthase needs to be elucidated. Two putative monoterpene synthases (LdTPS3 and LdTPS9) were isolated from L. dulcis leaf cDNA. To elucidate their catalytic functions, E. coli-produced recombinant enzymes with truncations of their chloroplast transit peptides were assayed with geranyl diphosphate (GPP). In vitro enzyme assays showed that LdTPS3 encodes bornyl diphosphate synthase (thus named LdBPPS) while LdTPS9 encodes linalool synthase. Interestingly, the N-terminus of LdBPPS possesses two arginine-rich (RRX 8 W) motifs, and enzyme assays showed that the presence of both RRX 8 W motifs completely inhibits the catalytic activity of LdBPPS. Only after the removal of the putative chloroplast transit peptide and the first RRX 8 W, LdBPPS could react with GPP to produce bornyl diphosphate. LdBPPS is distantly related to the known bornyl diphosphate synthase from sage in a phylogenetic analysis, indicating a converged evolution of camphor biosynthesis in sage and L. dulcis. The discovery of LdBPPS opens up the possibility of engineering L. dulcis to remove the undesirable product, camphor. Copyright © 2017 Elsevier Inc. All rights reserved.

A unique deubiquitinase that deconjugates phosphoribosyl-linked protein ubiquitination

Energy Technology Data Exchange (ETDEWEB)

Qiu, Jiazhang; Yu, Kaiwen; Fei, Xiaowen; Liu, Yao; Nakayasu, Ernesto S.; Piehowski, Paul D.; Shaw, Jared B.; Puvar, Kedar; Das, Chittaranjan; Liu, Xiaoyun; Luo, Zhao-Qing

2017-05-12

Ubiquitination regulates many aspects of host immunity and thus is a common target for infectious agents. Recent studies revealed that members of the SidE effector family of the bacterial pathogen Legionella pneumophila attacked several small GTPases associated with the endoplasmic reticulum by a novel ubiquitination mechanism that does not require the E1 and E2 enzymes of the host ubiquitination machinery. Following ubiquitin activation by ADP- ribosylation via a mono-ADP-ribosylation motif, ADP-ribosylated ubiquitin is cleaved by a phosphodiesterasedomainwithinSdeA,whichisconcomitantwiththelinkof phosphoribosylated ubiquitin to serine residues in the substrate. Here we demonstrate that the activity of SidEs is regulated by SidJ, another effector encoded by a gene situated in the locus coding for three members of the SidE family (SdeC, SdeB and SdeA). SidJ functions to remove ubiquitin from SidEs-modified substrates by cleaving the phosphodiester bond that links phosphoribosylated ubiquitin to protein substrates. Further, the deubiquitinase activity of SidJ is essential for its role in L. pneumophila infection. Finally, the activity of SidJ is required for efficiently reducing the abundance of ubiquitinated Rab33b in infected cells within a few hours after bacterial uptake. Our results establish SidJ as a deubiquitinase that functions to impose temporal regulation of the activity of the SidE effectors. The identification of SidJ may shed light on future study of signaling cascades mediated by this unique ubiquitination that also potentially regulates cellular processes in eukaryotic cells.

Structural determinants of enzyme binding affinity: the E1 component of pyruvate dehydrogenase from Escherichia coli in complex with the inhibitor thiamin thiazolone diphosphate.

Science.gov (United States)

Arjunan, Palaniappa; Chandrasekhar, Krishnamoorthy; Sax, Martin; Brunskill, Andrew; Nemeria, Natalia; Jordan, Frank; Furey, William

2004-03-09

Thiamin thiazolone diphosphate (ThTDP), a potent inhibitor of the E1 component from the Escherichia coli pyruvate dehydrogenase multienzyme complex (PDHc), binds to the enzyme with greater affinity than does the cofactor thiamin diphosphate (ThDP). To identify what determines this difference, the crystal structure of the apo PDHc E1 component complex with ThTDP and Mg(2+) has been determined at 2.1 A and compared to the known structure of the native holoenzyme, PDHc E1-ThDP-Mg(2+) complex. When ThTDP replaces ThDP, reorganization occurs in the protein structure in the vicinity of the active site involving positional and conformational changes in some amino acid residues, a change in the V coenzyme conformation, addition of new hydration sites, and elimination of others. These changes culminate in an increase in the number of hydrogen bonds to the protein, explaining the greater affinity of the apoenzyme for ThTDP. The observed hydrogen bonding pattern is not an invariant feature of ThDP-dependent enzymes but rather specific to this enzyme since the extra hydrogen bonds are made with nonconserved residues. Accordingly, these sequence-related hydrogen bonding differences likewise explain the wide variation in the affinities of different thiamin-dependent enzymes for ThTDP and ThDP. The sequence of each enzyme determines its ability to form hydrogen bonds to the inhibitor or cofactor. Mechanistic roles are suggested for the aforementioned reorganization and its reversal in PDHc E1 catalysis: to promote substrate binding and product release. This study also provides additional insight into the role of water in enzyme inhibition and catalysis.

Reaction of uridine diphosphate galactose 4-epimerase with a suicide inactivator

International Nuclear Information System (INIS)

Flentke, G.R.; Frey, P.A.

1990-01-01

UDPgalactose 4-epimerase from Escherichia coli is rapidly inactivated by the compounds uridine 5'-diphosphate chloroacetol (UDC) and uridine 5'-diphosphate bromoacetol (UCB). Both UDC and UDB inactivate the enzyme in neutral solution concomitant with the appearance of chromophores absorbing maximally at 325 and 328 nm, respectively. The reaction of UDC with the enzyme follows saturation kinetics characterized by a K D of 0.110 mM and k inact of 0.84 min -1 at pH 8.5 and ionic strength 0.2 M. The inactivation by UDC is competitively inhibited by competitive inhibitors of UDPgalactose 4-epimerase, and it is accompanied by the tight but noncovalent binding of UDC to the enzyme in a stoichiometry of 1 mol of UDC/mol of enzyme dimer, corresponding to 1 mol of UDC/mol of enzyme-bound NAD + . The inactivation of epimerase by uridine 5'-diphosphate [ 2 H 2 ]chloroacetol proceeds with a primary kinetic isotope effect (k H /k D ) of 1.4. The inactivation mechanism is proposed to involve a minimum of three steps: (a) reversible binding of UDC to the active site of UDPgalactose 4-epimerase; (b) enolization of the chloroacetol moiety of enzyme-bound UDC, catalyzed by an enzymic general base at the active site; (c) alkylation of the nicotinamide ring of NAD + at the active site by the chloroacetol enolate. The resulting adduct between UDC and NAD + is proposed to be the chromophore with λ max at 325 nm. The enzymic general base required to facilitate proton transfer in redox catalysis by this enzyme may be the general base that facilitates enolization of the chloroacetol moiety of UDC in the inactivation reaction

Geranylgeranyl diphosphate synthase in fission yeast is a heteromer of farnesyl diphosphate synthase (FPS), Fps1, and an FPS-like protein, Spo9, essential for sporulation.

Science.gov (United States)

Ye, Yanfang; Fujii, Makoto; Hirata, Aiko; Kawamukai, Makoto; Shimoda, Chikashi; Nakamura, Taro

2007-09-01

Both farnesyl diphosphate synthase (FPS) and geranylgeranyl diphosphate synthase (GGPS) are key enzymes in the synthesis of various isoprenoid-containing compounds and proteins. Here, we describe two novel Schizosaccharomyces pombe genes, fps1(+) and spo9(+), whose products are similar to FPS in primary structure, but whose functions differ from one another. Fps1 is essential for vegetative growth, whereas, a spo9 null mutant exhibits temperature-sensitive growth. Expression of fps1(+), but not spo9(+), suppresses the lethality of a Saccharomyces cerevisiae FPS-deficient mutant and also restores ubiquinone synthesis in an Escherichia coli ispA mutant, which lacks FPS activity, indicating that S. pombe Fps1 in fact functions as an FPS. In contrast to a typical FPS gene, no apparent GGPS homologues have been found in the S. pombe genome. Interestingly, although neither fps1(+) nor spo9(+) expression alone in E. coli confers clear GGPS activity, coexpression of both genes induces such activity. Moreover, the GGPS activity is significantly reduced in the spo9 mutant. In addition, the spo9 mutation perturbs the membrane association of a geranylgeranylated protein, but not that of a farnesylated protein. Yeast two-hybrid and coimmunoprecipitation analyses indicate that Fps1 and Spo9 physically interact. Thus, neither Fps1 nor Spo9 alone functions as a GGPS, but the two proteins together form a complex with GGPS activity. Because spo9 was originally identified as a sporulation-deficient mutant, we show here that expansion of the forespore membrane is severely inhibited in spo9Delta cells. Electron microscopy revealed significant accumulation membrane vesicles in spo9Delta cells. We suggest that lack of GGPS activity in a spo9 mutant results in impaired protein prenylation in certain proteins responsible for secretory function, thereby inhibiting forespore membrane formation.

Catalytic site interactions in yeast OMP synthase

DEFF Research Database (Denmark)

Hansen, Michael Riis; Barr, Eric W.; Jensen, Kaj Frank

2014-01-01

45 (2006) 5330-5342]. This behavior was investigated in the yeast enzyme by mutations in the conserved catalytic loop and 5-phosphoribosyl-1-diphosphate (PRPP) binding motif. Although the reaction is mechanistically sequential, the wild-type (WT) enzyme shows parallel lines in double reciprocal...

Heteromeric and homomeric geranyl diphosphate synthases from Catharanthus roseus and their role in monoterpene indole alkaloid biosynthesis.

Science.gov (United States)

Rai, Avanish; Smita, Shachi S; Singh, Anup Kumar; Shanker, Karuna; Nagegowda, Dinesh A

2013-09-01

Catharanthus roseus is the sole source of two most important monoterpene indole alkaloid (MIA) anti-cancer agents: vinblastine and vincristine. MIAs possess a terpene and an indole moiety derived from terpenoid and shikimate pathways, respectively. Geranyl diphosphate (GPP), the entry point to the formation of terpene moiety, is a product of the condensation of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) by GPP synthase (GPPS). Here, we report three genes encoding proteins with sequence similarity to large subunit (CrGPPS.LSU) and small subunit (CrGPPS.SSU) of heteromeric GPPSs, and a homomeric GPPSs. CrGPPS.LSU is a bifunctional enzyme producing both GPP and geranyl geranyl diphosphate (GGPP), CrGPPS.SSU is inactive, whereas CrGPPS is a homomeric enzyme forming GPP. Co-expression of both subunits in Escherichia coli resulted in heteromeric enzyme with enhanced activity producing only GPP. While CrGPPS.LSU and CrGPPS showed higher expression in older and younger leaves, respectively, CrGPPS.SSU showed an increasing trend and decreased gradually. Methyl jasmonate (MeJA) treatment of leaves significantly induced the expression of only CrGPPS.SSU. GFP localization indicated that CrGPPS.SSU is plastidial whereas CrGPPS is mitochondrial. Transient overexpression of AmGPPS.SSU in C. roseus leaves resulted in increased vindoline, immediate monomeric precursor of vinblastine and vincristine. Although C. roseus has both heteromeric and homomeric GPPS enzymes, our results implicate the involvement of only heteromeric GPPS with CrGPPS.SSU regulating the GPP flux for MIA biosynthesis.

Structure of active IspH enzyme from escherichia coli provides mechanistic insights into substrate reduction

KAUST Repository

Gräwert, Tobias

2009-07-20

The terminal step of the non-mevalonate pathway of terpene biosynthesis is catalyzed by IspH (see scheme). In the crystal structure of IspH from E. coli, a bound inorganic diphosphate ligand occupies the position of the diphosphate residue of the substrate. Together with mutation studies and theoretical calculations, these data support a mechanism which is analogous to the Birch reduction of allylic alcohols. © 2009 Wiley-VCH Verlag GmbH & Co. KGaA.

Manganese dipyridoxyl diphosphate:

DEFF Research Database (Denmark)

H, Brurok; Ardenkjær-Larsen, Jan Henrik; G, Hansson

1999-01-01

Manganese dipyridoxyl diphosphate (MnDPDP) is a contrast agent for magnetic resonance imaging (MRI) of the liver. Aims of the study were to examine if MnDPDP possesses superoxide dismutase (SOD) mimetic activity in vitro, and if antioxidant protection can be demonstrated in an ex vivo rat heart...

Uranium (Vi) sorption onto zirconium diphosphate chemically modified

International Nuclear Information System (INIS)

Garcia G, N.; Ordonez R, E.

2010-10-01

This work deals with the uranium (Vi) speciation after sorption onto zirconium diphosphate (ZrP 2 O 7 ) surface, hydrated and in a surface modified with organic acids. Oxalic and citric acids were chosen to modify the ZrP 2 O 7 surface because they have poly carboxylic groups and they mimic the organic matter in nature. Thus the interest of this work is to evaluate the uranium (Vi) sorption edge at different s ph values in natural and modified surfaces. The luminescence technique (fluorescence and phosphorescence, respectively) was used for the quantification and speciation of uranyl sorbed at the zirconium diphosphate interface. The fluorescence experiment, showed that adsorption of uranyl on surface of zirconium diphosphate tends to 100%. The speciation shows that there are different complexes in surface which were formed between zirconium diphosphate and uranyl, since it is produced a displacement of wavelength in fluorescence spectra of each system. (Author)

Bacillus caldolyticus prs gene encoding phosphoribosyldiphosphate synthase

DEFF Research Database (Denmark)

Krath, Britta N.; Hove-Jensen, Bjarne

1996-01-01

The prs gene, encoding phosphoribosyl-diphosphate (PRPP) synthase, as well as the flanking DNA sequences were cloned and sequenced from the Gram-positive thermophile, Bacillus caldolyticus. Comparison with the homologous sequences from the mesophile, Bacillus subtilis, revealed a gene (gca......D) encoding N-acetylglucosamine-l-phosphate uridyltransferase upstream of prs, and a gene homologous to ctc downstream of prs. cDNA synthesis with a B. caldolyticus gcaD-prs-ctc-specified mRNA as template, followed by amplification utilising the polymerase chain reaction indicated that the three genes are co......-transcribed. Comparison of amino acid sequences revealed a high similarity among PRPP synthases across a wide phylogenetic range. An E. coli strain harbouring the B. caldolyticus prs gene in a multicopy plasmid produced PRPP synthase activity 33-fold over the activity of a haploid B. caldolyticus strain. B. caldolyticus...

Production, purification, crystallization and preliminary X-ray diffraction studies of the nucleoside diphosphate kinase b from Leishmania major

International Nuclear Information System (INIS)

Tonoli, Celisa Caldana Costa; Vieira, Plinio Salmazo; Ward, Richard John; Arni, Raghuvir Krishnaswamy; Oliveira, Arthur Henrique Cavalcante de; Murakami, Mario Tyago

2009-01-01

Overexpression, purification, crystallization and preliminary X-ray diffraction analysis of the nucleoside diphosphate kinase b from Leishmania major are reported. The crystals belonged to the trigonal space group P3 2 21 and diffracted to 2.18 Å resolution. Nucleoside diphosphate kinases (NDKs; EC 2.7.4.6) play an essential role in the synthesis of nucleotides from intermediates in the salvage pathway in all parasitic trypanosomatids and their structural studies will be instrumental in shedding light on the biochemical machinery involved in the parasite life cycle and host–parasite interactions. In this work, NDKb from Leishmania major was overexpressed in Escherichia coli, purified to homogeneity and crystallized using the sitting-drop vapour-diffusion method. The NDK crystal diffracted to 2.2 Å resolution and belonged to the trigonal crystal system, with unit-cell parameters a = 114.2, c = 93.9 Å. Translation-function calculations yielded an unambiguous solution in the enantiomorphic space group P3 2 21

Open reading frame 176 in the photosynthesis gene cluster of Rhodobacter capsulatus encodes idi, a gene for isopentenyl diphosphate isomerase.

OpenAIRE

Hahn, F M; Baker, J A; Poulter, C D

1996-01-01

Isopentenyl diphosphate (IPP) isomerase catalyzes an essential activation step in the isoprenoid biosynthetic pathway. A database search based on probes from the highly conserved regions in three eukaryotic IPP isomerases revealed substantial similarity with ORF176 in the photosynthesis gene cluster in Rhodobacter capsulatus. The open reading frame was cloned into an Escherichia coli expression vector. The encoded 20-kDa protein, which was purified in two steps by ion exchange and hydrophobic...

Functional characterization of ent-copalyl diphosphate synthase from Andrographis paniculata with putative involvement in andrographolides biosynthesis.

Science.gov (United States)

Shen, Qinqin; Li, Lixia; Jiang, Yu; Wang, Qiang

2016-01-01

To characterize the ent-copalyl diphosphate (ent-CPP) synthase involved in the biosynthetic pathway of andrographolides in a medicinal plant, Andrographis paniculata. The ent-CPP synthase (ent-CPS) gene was cloned from A. paniculata and its encoded ApCPS was demonstrated to react with (E,E,E)-geranylgeranyl diphosphate to form ent-CPP through recombinant expression in Escherichia coli. Site-directed mutagenesis of the Asp to Ala in the conserved DXDD motif of ApCPS resulted in loss of function. One Arg is located in the conserved position close to DXDD motif indicating the involvement of ApCPS in specialized metabolism. In addition, RT-PCR analysis revealed that ApCPS was expressed in all tissues of A. paniculata at all growth stages, which is consistent with andrographolides accumulating in these organs. Methyl jasmonate induced ApCPS gene expression, matching inducible accumulation of andrographolides in vivo. ApCPS is the first ent-CPS characterized in A. paniculata and is suggested to be involved in biosynthesis of andrographolides that have high pharmaceutical values.

Protein preparation, crystallization and preliminary X-ray analysis of Trypanosoma cruzi nucleoside diphosphate kinase 1

International Nuclear Information System (INIS)

Gómez Barroso, J. A.; Pereira, H.; Miranda, M.; Pereira, C.; Garratt, R. C.; Aguilar, C. F.

2010-01-01

T. cruzi TcNDPK1 was overexpressed in Escherichia coli as an N-terminally poly-His-tagged fusion protein and crystallized. The flagellated protozoan parasite Trypanosoma cruzi is the aetiological agent of Chagas disease. Nucleoside diphosphate kinases (NDPKs) are enzymes that are involved in energy management and nucleoside balance in the cell. T. cruzi TcNDPK1, a canonical isoform, was overexpressed in Escherichia coli as an N-terminally poly-His-tagged fusion protein and crystallized. Crystals grew after 72 h in 0.2 M MgCl 2 , 20% PEG 3350. Data were collected to 3.5 Å resolution using synchrotron X-ray radiation at the National Synchrotron Light Laboratory (Campinas, Brazil). The crystals belonged to the trigonal space group P3, with unit-cell parameters a = b = 127.84, c = 275.49 Å. Structure determination is under way and will provide relevant information that may lead to the first step in rational drug design for the treatment of Chagas disease

Geranylfarnesyl diphosphate synthase from Methanosarcina mazei: Different role, different evolution

International Nuclear Information System (INIS)

Ogawa, Takuya; Yoshimura, Tohru; Hemmi, Hisashi

2010-01-01

The gene of (all-E) geranylfarnesyl diphosphate synthase that is responsible for the biosynthesis of methanophenazine, an electron carrier utilized for methanogenesis, was cloned from a methanogenic archaeon Methanosarcina mazei Goe1. The properties of the recombinant enzyme and the results of phylogenetic analysis suggest that the enzyme is closely related to (all-E) prenyl diphosphate synthases that are responsible for the biosynthesis of respiratory quinones, rather than to the enzymes involved in the biosynthesis of archaeal membrane lipids, including (all-E) geranylfarnesyl diphosphate synthase from a thermophilic archaeon.

Evaluation of the sorption of Eu(III) in titanium diphosphate

International Nuclear Information System (INIS)

Ortiz O, H.B.; Ordonez R, E.; Fernandez V, S.M.

2007-01-01

In this work its are presented: the synthesis, physicochemical characterization and the surface parameters estimation that can be related with the retention properties of the titanium diphosphate for the actinides of valence III (Pu, Am, Cm among others), using the Eu 3+ like a chemical analog. The surface area, hydration time, zero charge point, density of active sites and the surface species distribution in the titanium diphosphate are reported. This information was used to explain the retention of the Eu(lll) in the surface of the titanium diphosphate. (Author)

Allosteric regulation of the GTP activated and CTP inhibited uracil phosphoribosyltransferase from the thermophilic archaeon Sulfolobus solfataricus

DEFF Research Database (Denmark)

Jensen, Kaj Frank; Arent, Susan; Larsen, Sine

2005-01-01

The upp gene, encoding uracil phosphoribosyltransferase (UPRTase) from the thermoacidophilic archaeon Sulfolobus solfataricus, was cloned and expressed in Escherichia coli. The enzyme was purified to homogeneity. It behaved as a tetramer in solution and showed optimal activity at pH 5.5 when...... assayed at 60 °C. Enzyme activity was strongly stimulated by GTP and inhibited by CTP. GTP caused an approximately 20-fold increase in the turnover number kcat and raised the Km values for 5-phosphoribosyl-1-diphosphate (PRPP) and uracil by two- and >10-fold, respectively. The inhibition by CTP...... was complex as it depended on the presence of the reaction product UMP. Neither CTP nor UMP were strong inhibitors of the enzyme, but when present in combination their inhibition was extremely powerful. Ligand binding analyses showed that GTP and PRPP bind cooperatively to the enzyme and that the inhibitors...

Mevalonate 5-diphosphate mediates ATP binding to the mevalonate diphosphate decarboxylase from the bacterial pathogen Enterococcus faecalis

Energy Technology Data Exchange (ETDEWEB)

Chen, Chun-Liang; Mermoud, James C.; Paul, Lake N.; Steussy, Calvin Nicklaus; Stauffacher, Cynthia V. (Purdue)

2017-10-12

The mevalonate pathway produces isopentenyl diphosphate (IPP), a building block for polyisoprenoid synthesis, and is a crucial pathway for growth of the human bacterial pathogen Enterococcus faecalis. The final enzyme in this pathway, mevalonate diphosphate decarboxylase (MDD), acts on mevalonate diphosphate (MVAPP) to produce IPP while consuming ATP. This essential enzyme has been suggested as a therapeutic target for the treatment of drug-resistant bacterial infections. Here, we report functional and structural studies on the mevalonate diphosphate decarboxylase from E. faecalis (MDDEF). The MDDEF crystal structure in complex with ATP (MDDEF–ATP) revealed that the phosphate-binding loop (amino acids 97–105) is not involved in ATP binding and that the phosphate tail of ATP in this structure is in an outward-facing position pointing away from the active site. This suggested that binding of MDDEF to MVAPP is necessary to guide ATP into a catalytically favorable position. Enzymology experiments show that the MDDEF performs a sequential ordered bi-substrate reaction with MVAPP as the first substrate, consistent with the isothermal titration calorimetry (ITC) experiments. On the basis of ITC results, we propose that this initial prerequisite binding of MVAPP enhances ATP binding. In summary, our findings reveal a substrate-induced substrate-binding event that occurs during the MDDEF-catalyzed reaction. The disengagement of the phosphate-binding loop concomitant with the alternative ATP-binding configuration may provide the structural basis for antimicrobial design against these pathogenic enterococci.

Molecular analysis of formaldehyde-induced mutations in human lymphoblasts and E. coli

International Nuclear Information System (INIS)

Crosby, R.M.; Richardson, K.K.; Craft, T.R.; Benforado, K.B.; Liber, H.L.; Skopek, T.R.

1988-01-01

The molecular nature of formaldehyde (HCHO)-induced mutations was studied in both human lymphoblasts and E. coli. Thirty HPRT - human lymphoblast colonies induced by eight repetitive 150 μM HCHO treatments were characterized by Southern blot analysis. Fourteen of these mutants (47%) had visible deletions of some or all of the X-linked HPRT bands, indicating that HCHO can induce large losses of DNA in human lymphoblasts. In E. coli., DNA alterations induced by HCHO were characterized with use of the xanthine guanine phosphoribosyl transferase (gpt) gene as the genetic target. Exposure of E. coli to 4 mM HCHO for 1 hr induced large insertions (41%), large deletions (18%), and point mutations (41%). Dideoxy DNA sequencing revealed that most of the point mutations were transversions at GC base pairs. In contrast, exposure of E. coli to 40 mM HCHO for 1 hr produced 92% point mutations, 62% of which were transitions at a single AT base pair in the gene. Therefore, HCHO is capable of producing different genetic alterations in E. coli at different concentrations, suggesting fundamental differences in the mutagenic mechanisms operating at the two concentrations used. Naked pSV2gpt plasmid DNA was exposed to 3.3 or 10 mM HCHO and transformed into E. coli. Most of the resulting mutations were frameshifts, again suggesting a different mutagenic mechanism

Properties of ribulose diphosphate carboxylase immobilized on porous glass

Science.gov (United States)

Shapira, J.; Hanson, C. L.; Lyding, J. M.; Reilly, P. J.

1974-01-01

Ribulose-1,5-diphosphate carboxylase from spinach has been bound to arylamine porous glass with a diazo linkage and to alklamine porous glass with glutaraldehyde. Stability at elevated temperatures and responses to changes of pH and ribulose-1,5-diphosphate, Mg(2+), and dithiothreitol concentrations were not significantly different from the soluble enzyme, though stability at 4 C was somewhat improved.

Solid-Phase Synthesis of a New Diphosphate 5-Aminoimidazole-4-carboxamide Riboside (AICAR Derivative and Studies toward Cyclic AICAR Diphosphate Ribose

Directory of Open Access Journals (Sweden)

Gennaro Piccialli

2011-09-01

Full Text Available The solid-phase synthesis of the first example of a new diphosphate AICAR derivative is reported. The new substance is characterized by the presence of a 5'-phosphate group while a second phosphate moiety is installed on a 5-hydroxypentyl chain attached to the 4-N-position of AICAR. Cyclization of the diphosphate derivative by pyrophosphate bond formation allowed for the formation of a novel AICAR-based cyclic ADP-ribose (cADPR mimic.

Chrysanthemyl diphosphate synthase operates in planta as a bifunctional enzyme with chrysanthemol synthase activity

DEFF Research Database (Denmark)

Yang, Ting; Gao, Liping; Hu, Hao

2014-01-01

Chrysanthemyl diphosphate synthase (CDS) is the first path-way-specific enzyme in the biosynthesis of pyrethrins, the most widely used plant-derived pesticide. CDS catalyzes c1′-2-3 cyclopropanation reactions of two molecules of dimethylallyl diphosphate (DMAPP) to yield chrysanthemyl diphosphate...

Synthesis and behavior at heating of amorphous calcium diphosphate

International Nuclear Information System (INIS)

Levchenko, L.V.; Nurkina, Z.S.; Griggs, D.; Sinyayev, V.A.

2005-01-01

There is description of synthesis of amorphous calcium diphosphate in the article. The compound was received via exchange reaction between sodium diphosphate and calcium chloride in water solution. The results of investigation of behavior when heated of produced substance are presented here as well. Composition and structure of precipitated substances and products of its thermal convention were determined by methods of IR-, NMR 31 P and X-ray spectroscopy

In vivo photoinactivation of Escherichia coli ribonucleoside reductase by near-ultraviolet light

International Nuclear Information System (INIS)

Peters, J.

1977-01-01

Some experimental work is described showing that near-U.V. irradiation of E.coli cells selectively destroys RDP-reductase (ribonucleoside diphosphate reductase) activity in vivo are providing evidence relating the loss of RDP-reductase to loss of cellular visibility and the inactivity of irrdiated cells to support the replication of DNA phages. The data are consistent with the interpretation that the principal cause in the killing of exponentially growing E.coli cells by near-U.V., and the loss of ability of irradiated host cells to support the replication of DNA phages, is the photoinactivation of the RDP-reductase complex. (U.K.)

In vivo photoinactivation of Escherichia coli ribonucleoside reductase by near-ultraviolet light

Energy Technology Data Exchange (ETDEWEB)

Peters, J [California Univ., Irvine (USA)

1977-06-09

Some experimental work is described showing that near-uv irradiation of E.coli cells selectively destroys RDP-reductase (ribonucleoside diphosphate reductase) activity in vivo are providing evidence relating the loss of RDP-reductase to loss of cellular visibility and the inactivity of irrdiated cells to support the replication of DNA phages. The data are consistent with the interpretation that the principal cause in the killing of exponentially growing E.coli cells by near-uv, and the loss of ability of irradiated host cells to support the replication of DNA phages, is the photoinactivation of the RDP-reductase complex.

The ura5 gene of the ascomycete Sordaria macrospora: molecular cloning, characterization and expression in Escherichia coli.

Science.gov (United States)

Le Chevanton, L; Leblon, G

1989-04-15

We cloned the ura5 gene coding for the orotate phosphoribosyl transferase from the ascomycete Sordaria macrospora by heterologous probing of a Sordaria genomic DNA library with the corresponding Podospora anserina sequence. The Sordaria gene was expressed in an Escherichia coli pyrE mutant strain defective for the same enzyme, and expression was shown to be promoted by plasmid sequences. The nucleotide sequence of the 1246-bp DNA fragment encompassing the region of homology with the Podospora gene has been determined. This sequence contains an open reading frame of 699 nucleotides. The deduced amino acid sequence shows 72% similarity with the corresponding Podospora protein.

Overexpression of an isopentenyl diphosphate isomerase gene to enhance trans-polyisoprene production in Eucommia ulmoides Oliver

Directory of Open Access Journals (Sweden)

Chen Ren

2012-10-01

Full Text Available Abstract Background Natural rubber produced by plants, known as polyisoprene, is the most widely used isoprenoid polymer. Plant polyisoprenes can be classified into two types; cis-polyisoprene and trans-polyisoprene, depending on the type of polymerization of the isoprene unit. More than 2000 species of higher plants produce latex consisting of cis-polyisoprene. Hevea brasiliensis (rubber tree produces cis-polyisoprene, and is the key source of commercial rubber. In contrast, relatively few plant species produce trans-polyisoprene. Currently, trans-polyisoprene is mainly produced synthetically, and no plant species is used for its commercial production. Results To develop a plant-based system suitable for large-scale production of trans-polyisoprene, we selected a trans-polyisoprene-producing plant, Eucommia ulmoides Oliver, as the target for genetic transformation. A full-length cDNA (designated as EuIPI, Accession No. AB041629 encoding isopentenyl diphosphate isomerase (IPI was isolated from E. ulmoides. EuIPI consisted of 1028 bp with a 675-bp open reading frame encoding a protein with 224 amino acid residues. EuIPI shared high identity with other plant IPIs, and the recombinant protein expressed in Escherichia coli showed IPI enzymatic activity in vitro. EuIPI was introduced into E. ulmoides via Agrobacterium-mediated transformation. Transgenic lines of E. ulmoides overexpressing EuIPI showed increased EuIPI expression (up to 19-fold that of the wild-type and a 3- to 4-fold increase in the total content of trans-polyisoprenes, compared with the wild-type (non-transgenic root line control. Conclusions Increasing the expression level of EuIPI by overexpression increased accumulation of trans-polyisoprenes in transgenic E. ulmoides. IPI catalyzes the conversion of isopentenyl diphosphate to its highly electrophilic isomer, dimethylallyl diphosphate, which is the first step in the biosynthesis of all isoprenoids, including polyisoprene. Our

Structure and mechanism of the diterpene cyclase ent-copalyl diphosphate synthase

Energy Technology Data Exchange (ETDEWEB)

Köksal, Mustafa; Hu, Huayou; Coates, Robert M.; Peters, Reuben J.; Christianson, David W. (UIUC); (Iowa State); (Penn)

2011-09-20

The structure of ent-copalyl diphosphate synthase reveals three {alpha}-helical domains ({alpha}, {beta} and {gamma}), as also observed in the related diterpene cyclase taxadiene synthase. However, active sites are located at the interface of the {beta}{gamma} domains in ent-copalyl diphosphate synthase but exclusively in the {alpha} domain of taxadiene synthase. Modular domain architecture in plant diterpene cyclases enables the evolution of alternative active sites and chemical strategies for catalyzing isoprenoid cyclization reactions.

Motesanib diphosphate in progressive differentiated thyroid cancer

DEFF Research Database (Denmark)

Sherman, Steven I; Wirth, Lori J; Droz, Jean-Pierre

2008-01-01

BACKGROUND: The expression of vascular endothelial growth factor (VEGF) is characteristic of differentiated thyroid cancer and is associated with aggressive tumor behavior and a poor clinical outcome. Motesanib diphosphate (AMG 706) is a novel oral inhibitor of VEGF receptors, platelet-derived gr...

Asymmetric Stetter reactions catalyzed by thiamine diphosphate-dependent enzymes.

Science.gov (United States)

Kasparyan, Elena; Richter, Michael; Dresen, Carola; Walter, Lydia S; Fuchs, Georg; Leeper, Finian J; Wacker, Tobias; Andrade, Susana L A; Kolter, Geraldine; Pohl, Martina; Müller, Michael

2014-12-01

The intermolecular asymmetric Stetter reaction is an almost unexplored transformation for biocatalysts. Previously reported thiamine diphosphate (ThDP)-dependent PigD from Serratia marcescens is the first enzyme identified to catalyze the Stetter reaction of α,β-unsaturated ketones (Michael acceptor substrates) and α-keto acids. PigD is involved in the biosynthesis of the potent cytotoxic agent prodigiosin. Here, we describe the investigation of two new ThDP-dependent enzymes, SeAAS from Saccharopolyspora erythraea and HapD from Hahella chejuensis. Both show a high degree of homology to the amino acid sequence of PigD (39 and 51 %, respectively). The new enzymes were heterologously overproduced in Escherichia coli, and the yield of soluble protein was enhanced by co-expression of the chaperone genes groEL/ES. SeAAS and HapD catalyze intermolecular Stetter reactions in vitro with high enantioselectivity. The enzymes possess a characteristic substrate range with respect to Michael acceptor substrates. This provides support for a new type of ThDP-dependent enzymatic activity, which is abundant in various species and not restricted to prodigiosin biosynthesis in different strains. Moreover, PigD, SeAAS, and HapD are also able to catalyze asymmetric carbon-carbon bond formation reactions of aldehydes and α-keto acids, resulting in 2-hydroxy ketones.

Mutational, Phylogeny and Evolution Analyses of Salvia Copalyl Diphosphate Synthase

International Nuclear Information System (INIS)

Hao, D. C.; Thimmappa, R. B.; Xiao, P. G.

2016-01-01

The cyclization of geranylgeranyl diphosphate (GGPP) is catalyzed by copalyl diphosphate synthase (CPS), a class II diterpene synthase (diTPS), to form copalyl diphosphate (CPP), which is an essential substrate of a variety of diterpenes in secondary metabolism of angiosperm including Salvia medicinal plants. The protein environment of the N-terminal class II active site stabilizes the carbocation intermediates and maintains the catalytic activity of angiosperm class II diTPS. The virtual modeling and mutagenesis of the class II diTPS of Salvia miltiorrhiza (SmCPS) were accomplished to illuminate the catalytic activity of SmCPS. Terminal truncations and point mutations established the role of the Beta-Gamma domain and Alpha domain, i.e., they facilitate the flexible conformational change of the class II active site after substrate binding. E203 and K238 in the N-ter Gamma domain of SmCPS1 are functional in the substrate binding and conformational transition and might be essential in catalysis. Similar to other CPSs, the ensuing protonation of the GGPP substrate and coordination of the diphosphate group are governed by highly conserved residues in the DxDD motif of SmCPS, e.g., D372 of CPS1. Moreover, F256 and Y505 stabilize the carbocation and control the enzymatic activity during CPP formation. The amino acids of the predicted active sites, despite under purifying selection, vary greatly, corresponding to the functional flexibility of angiosperm CPSs. Molecular phylogeny and evolution analyses suggest early and ongoing evolution of labdane-related diterpenoid metabolism in angiosperm. (author)

Integrated analysis of isopentenyl pyrophosphate (IPP) toxicity in isoprenoid-producing Escherichia coli

DEFF Research Database (Denmark)

George, Kevin W; Thompson, Mitchell; Kim, Joonhoon

2018-01-01

evidence that IPP can be transported by E. coli, findings that might be broadly relevant for the study of isoprenoid biosynthesis. Finally, we discover that IPP accumulation leads to the formation of ApppI, a nucleotide analog of IPP that may contribute to observed toxicity phenotypes. This comprehensive...... assessment of IPP stress suggests potential strategies for the alleviation of prenyl diphosphate toxicity and highlights possible engineering targets for improved IPP flux and high titer isoprenoid production....

Escherichia coli class Ib ribonucleotide reductase contains a dimanganese(III)-tyrosyl radical cofactor in vivo†

Science.gov (United States)

Cotruvo, Joseph A.; Stubbe, JoAnne

2011-01-01

Escherichia coli class Ib ribonucleotide reductase (RNR) converts nucleoside 5′-diphosphates to deoxynucleoside 5′-diphosphates in iron-limited and oxidative stress conditions. We have recently demonstrated in vitro that this RNR is active with both diferric-tyrosyl radical (FeIII2-Y•) and dimanganese(III)-Y• (MnIII2-Y•) cofactors in the β2 subunit, NrdF [Cotruvo J.A., Jr. and Stubbe J., Biochemistry (2010) 49, 1297–1309]. Here we demonstrate, by purification of this protein from its endogenous levels in an E. coli strain deficient in its five known iron uptake pathways and grown under iron-limited conditions, that the MnIII2-Y• cofactor is assembled in vivo. This is the first definitive determination of the active cofactor of a class Ib RNR purified from its native organism without overexpression. From 88 g of cell paste, 150 μg of NrdF was isolated with ~95% purity, with 0.2 Y•/β2, 0.9 Mn/β2, and a specific activity of 720 nmol/min/mg. In these conditions, the class Ib RNR is the primary active RNR in the cell. Our results strongly suggest that E. coli NrdF is an obligate manganese protein in vivo and that the MnIII2-Y• cofactor assembly pathway we have identified in vitro involving the flavodoxin-like protein NrdI, present inside the cell at catalytic levels, is operative in vivo. PMID:21250660

Utilization of biodiesel by-product as substrate for high-production of β-farnesene via relatively balanced mevalonate pathway in Escherichia coli.

Science.gov (United States)

You, Shengping; Yin, Qingdian; Zhang, Jianye; Zhang, Chengyu; Qi, Wei; Gao, Lan; Tao, Zhiping; Su, Rongxin; He, Zhimin

2017-11-01

Farnesene has been identified as suitable jet fuel substitutes and metabolic engineering for microbial production of farnesene is an alternative and attractive route. In this study, due to accumulation of toxic intermediate isopentenyl pyrophosphate (IPP), an engineered Escherichia coli strain harboring heterologous mevalonate pathway produced only 4.11mg/L β-farnesene. Through higher-level expression of isopentenyl diphosphate isomerase and farnesyl diphosphate synthase to minimize the accumulated IPP, another engineered strain with relatively balanced mevalonate pathway was constructed and had the highest production of β-farnesene to date (8.74g/L) by Escherichia coli in a lab bioreactor. Furthermore, this is the first report on utilization of biodiesel by-product (simple purification) as substrate for high-production of β-farnesene by the engineered strain optimized and β-farnesene concentration reached 2.83g/L in a lab bioreactor. Therefore, the engineered strain optimized could be used as a platform host for high-production of other terpenoids using biodiesel by-product as substrate. Copyright © 2017 Elsevier Ltd. All rights reserved.

Structure of trihydrated rare-earth acid diphosphates LnHP2O7·3H2O (Ln=La, Er)

International Nuclear Information System (INIS)

Ben Moussa, S.; Ventemillas, S.; Cabeza, A.; Gutierrez-Puebla, E.; Sanz, J.

2004-01-01

In trihydrated lanthanum acid-diphosphates LnHP 2 O 7 ·3H 2 O, prepared from acid LnCl 3 and Na 4 P 2 O 7 solutions (pH=1), two crystal forms were obtained. Layered structures of two representative members of this family have been determined by single-crystal X-ray diffraction (XRD) technique. In the case of orthorhombic LaHP 2 O 7 ·3H 2 O (type I), lanthanum cations are ninefold coordinated and diphosphate groups adopt a staggered (alternated) configuration. In the case of triclinic ErHP 2 O 7 ·3H 2 O (type II), erbium cations are eightfold coordinated and diphosphate groups adopt an eclipsed configuration. In agreement with Infrared (IR) spectroscopic data, a bended configuration for diphosphate groups has been deduced. In both structures, one-dimensional chains of edge-sharing rare-earth polyhedra are linked together by diphosphate groups to form the phosphate layers. In both diphosphates, PO 4 and HPO 4 environments have been identified by 31 P MAS-NMR technique. In the two compounds, OH groups of HPO 4 tetrahedra point out of diphosphate planes interacting with adjacent layers. In La-diphosphate, the interaction between HPO 4 groups and water molecules of adjacent layers is favored; however, in Er-diphosphate, the interaction between phosphate acid groups of contiguous layers is produced. Based on structural information deduced, differences detected in IR and NMR spectra of two disphosphates are discussed

Modification of zirconium diphosphate with salicylic acid and its effect on the uranium (Vi) sorption

International Nuclear Information System (INIS)

Almazan T, M. G.; Garcia G, N.; Simoni, E.

2014-10-01

The surface of zirconium diphosphate (ZrP 2 O 7 ) was modified with salicylic acid and its effect was evaluated on the uranium (Vi) sorption. The modified surface of the material was analyzed with different analytical techniques among which are included the atomic force microscopy, scanning electron microscopy and X-ray photoelectron spectroscopy. This analysis allowed showing that the salicylic acid is being held on the surface of the zirconium diphosphate. The reactivity of modified zirconium diphosphate compared with uranium (Vi) was investigated using the classical method of batch sorption. The analysis of sorption isotherms shows that the salicylic acid has an important effect in the uranium (Vi) sorption. According to the study conducted, the interaction among the uranium (Vi) and the surface of zirconium diphosphate modified with the salicylic acid most likely leads to the complexes formation of binary (U(Vi)/ZrP 2 O 7 ) and ternary (U(Vi)/salicylate/ZrP 2 O 7 ) surface. (Author)

Recent Advances in the Development of Mammalian Geranylgeranyl Diphosphate Synthase Inhibitors

Directory of Open Access Journals (Sweden)

Staci L. Haney

2017-05-01

Full Text Available The enzyme geranylgeranyl diphosphate synthase (GGDPS catalyzes the synthesis of the 20-carbon isoprenoid geranylgeranyl diphosphate (GGPP. GGPP is the isoprenoid donor for protein geranylgeranylation reactions catalyzed by the enzymes geranylgeranyl transferase (GGTase I and II. Inhibitors of GGDPS result in diminution of protein geranylgeranylation through depletion of cellular GGPP levels, and there has been interest in GGDPS inhibitors as potential anti-cancer agents. Here we discuss recent advances in the development of GGDPS inhibitors, including insights gained by structure-function relationships, and review the preclinical data that support the continued development of this novel class of drugs.

Silver vanadium diphosphate Ag2VP2O8: Electrochemistry and characterization of reduced material providing mechanistic insights

International Nuclear Information System (INIS)

Takeuchi, Esther S.; Lee, Chia-Ying; Cheng, Po-Jen; Menard, Melissa C.; Marschilok, Amy C.; Takeuchi, Kenneth J.

2013-01-01

Silver vanadium phosphorous oxides (Ag w V x P y O z ) are notable battery cathode materials due to their high energy density and demonstrated ability to form in-situ Ag metal nanostructured electrically conductive networks within the cathode. While analogous silver vanadium diphosphate materials have been prepared, electrochemical evaluations of these diphosphate based materials have been limited. We report here the first electrochemical study of a silver vanadium diphosphate, Ag 2 VP 2 O 8 , where the structural differences associated with phosphorous oxides versus diphosphates profoundly affect the associated electrochemistry. Reminiscent of Ag 2 VO 2 PO 4 reduction, in-situ formation of silver metal nanoparticles was observed with reduction of Ag 2 VP 2 O 8 . However, counter to Ag 2 VO 2 PO 4 reduction, Ag 2 VP 2 O 8 demonstrates a significant decrease in conductivity upon continued electrochemical reduction. Structural analysis contrasting the crystallography of the parent Ag 2 VP 2 O 8 with that of the proposed Li 2 VP 2 O 8 reduction product is employed to gain insight into the observed electrochemical reduction behavior, where the structural rigidity associated with the diphosphate anion may be associated with the observed particle fracturing upon deep electrochemical reduction. Further, the diphosphate anion structure may be associated with the high thermal stability of the partially reduced Ag 2 VP 2 O 8 materials, which bodes well for enhanced safety of batteries incorporating this material. - Graphical abstract: Structure and galvanostatic intermittent titration-type test data for silver vanadium diphosphate, Ag 2 VP 2 O 8 . Highlights: ► First electrochemical study of a silver vanadium diphosphate, Ag 2 VP 2 O 8 . ► In-situ formation of Ag 0 nanoparticles was observed upon electrochemical reduction. ► Structural analysis used to provide insight of the electrochemical behavior

Purification and properties of phosphoribosyl-diphosphate synthetase from Bacillus subtilis

DEFF Research Database (Denmark)

Arnvig, Kirsten; Hove-Jensen, Bjarne; Switzer, Robert L.

1990-01-01

enzyme required Mg2+ and inorganic phosphate for activity; Mn2+ supported only 30% the activity seen with Mg2+. Michaelis constants for ATP and ribose 5-phosphate (Rib5P) were 0.66 mM and 0.48 mM, respectively. Of several end products tested, only ADP was strongly inhibitory; GDP was a weak inhibitor....... ADP inhibition displayed homotropic cooperativity and was enhanced by increasing saturation of the enzyme with ATP. These observations strongly suggest a specific allosteric site for ADP binding. A comparison of physical and kinetic properties of bacterial and mammalian PPRibP synthetases is presented....

Dependence of the product chain-length on detergents for long-chain E-polyprenyl diphosphate synthases

Science.gov (United States)

Pan, Jian-Jung; Ramamoorthy, Gurusankar; Poulter, C. Dale

2013-01-01

Long-chain E-polyprenyl diphosphate synthases (E-PDS) catalyze repetitive addition of isopentenyl diphosphate (IPP) to the growing prenyl chain of an allylic diphosphate. The polyprenyl diphosphate products are required for the biosynthesis of ubiquinones and menaquinones required for electron transport during oxidative phosphorylation to generate ATP. In vitro, the long-chain PDSs require addition of phospholipids or detergents to the assay buffer to enhance product release and maintain efficient turnover. During preliminary assays of product chain-length with anionic, zwitterionic, and non-ionic detergents, we discovered considerable variability. Examination of a series of non-ionic PEG detergents with several long-chain E-PDSs from different organisms revealed that in vitro incubations with nonaethylene glycol monododecyl ether or Triton X-100 typically gave chain lengths that corresponded to those of the isoprenoid moieties in respiratory quinones synthesized in vivo. In contrast incubations in buffer with n-butanol, CHAPS, DMSO, n-octyl-β-glucopyranoside, or β-cyclodextrin or in buffer without detergent typically proceeded more slowly and gave a broad range of chain lengths. PMID:23802587

Functional evidence for the critical amino-terminal conserved domain and key amino acids of Arabidopsis 4-HYDROXY-3-METHYLBUT-2-ENYL DIPHOSPHATE REDUCTASE.

Science.gov (United States)

Hsieh, Wei-Yu; Sung, Tzu-Ying; Wang, Hsin-Tzu; Hsieh, Ming-Hsiun

2014-09-01

The plant 4-HYDROXY-3-METHYLBUT-2-ENYL DIPHOSPHATE REDUCTASE (HDR) catalyzes the last step of the methylerythritol phosphate pathway to synthesize isopentenyl diphosphate and its allyl isomer dimethylallyl diphosphate, which are common precursors for the synthesis of plastid isoprenoids. The Arabidopsis (Arabidopsis thaliana) genomic HDR transgene-induced gene-silencing lines are albino, variegated, or pale green, confirming that HDR is essential for plants. We used Escherichia coli isoprenoid synthesis H (Protein Data Bank code 3F7T) as a template for homology modeling to identify key amino acids of Arabidopsis HDR. The predicted model reveals that cysteine (Cys)-122, Cys-213, and Cys-350 are involved in iron-sulfur cluster formation and that histidine (His)-152, His-241, glutamate (Glu)-242, Glu-243, threonine (Thr)-244, Thr-312, serine-379, and asparagine-381 are related to substrate binding or catalysis. Glu-242 and Thr-244 are conserved only in cyanobacteria, green algae, and land plants, whereas the other key amino acids are absolutely conserved from bacteria to plants. We used site-directed mutagenesis and complementation assay to confirm that these amino acids, except His-152 and His-241, were critical for Arabidopsis HDR function. Furthermore, the Arabidopsis HDR contains an extra amino-terminal domain following the transit peptide that is highly conserved from cyanobacteria, and green algae to land plants but not existing in the other bacteria. We demonstrated that the amino-terminal conserved domain was essential for Arabidopsis and cyanobacterial HDR function. Further analysis of conserved amino acids in the amino-terminal conserved domain revealed that the tyrosine-72 residue was critical for Arabidopsis HDR. These results suggest that the structure and reaction mechanism of HDR evolution have become specific for oxygen-evolving photosynthesis organisms and that HDR probably evolved independently in cyanobacteria versus other prokaryotes. © 2014

Targeted cytosine deaminase-uracil phosphoribosyl transferase suicide gene therapy induces small cell lung cancer-specific cytotoxicity and tumor growth delay

DEFF Research Database (Denmark)

Christensen, Camilla L; Gjetting, Torben; Poulsen, Thomas Tuxen

2010-01-01

Small cell lung cancer (SCLC) is a highly malignant cancer for which there is no curable treatment. Novel therapies are therefore in great demand. In the present study we investigated the therapeutic effect of transcriptionally targeted suicide gene therapy for SCLC based on the yeast cytosine...... deaminase (YCD) gene alone or fused with the yeast uracil phosphoribosyl transferase (YUPRT) gene followed by administration of 5-fluorocytosine (5-FC) prodrug. Experimental design: The YCD gene or the YCD-YUPRT gene was placed under regulation of the SCLC-specific promoter insulinoma-associated 1 (INSM1...

Thiamin diphosphate in biological chemistry: new aspects of thiamin metabolism, especially triphosphate derivatives acting other than as cofactors.

Science.gov (United States)

Bettendorff, Lucien; Wins, Pierre

2009-06-01

Prokaryotes, yeasts and plants synthesize thiamin (vitamin B1) via complex pathways. Animal cells capture the vitamin through specific high-affinity transporters essential for internal thiamin homeostasis. Inside the cells, thiamin is phosphorylated to higher phosphate derivatives. Thiamin diphosphate (ThDP) is the best-known thiamin compound because of its role as an enzymatic cofactor. However, in addition to ThDP, at least three other thiamin phosphates occur naturally in most cells: thiamin monophosphate, thiamin triphosphate (ThTP) and the recently discovered adenosine thiamin triphosphate. It has been suggested that ThTP has a specific neurophysiological role, but recent data favor a much more basic metabolic function. During amino acid starvation, Escherichia coli accumulate ThTP, possibly acting as a signal involved in the adaptation of the bacteria to changing nutritional conditions. In animal cells, ThTP can phosphorylate some proteins, but the physiological significance of this mechanism remains unknown. Adenosine thiamin triphosphate, recently discovered in E. coli, accumulates during carbon starvation and might act as an alarmone. Among the proteins involved in thiamin metabolism, thiamin transporters, thiamin pyrophosphokinase and a soluble 25-kDa thiamin triphosphatase have been characterized at the molecular level, in contrast to thiamin mono- and diphosphatases whose specificities remain to be proven. A soluble enzyme catalyzing the synthesis of adenosine thiamin triphosphate from ThDP and ADP or ATP has been partially characterized in E. coli, but the mechanism of ThTP synthesis remains elusive. The data reviewed here illustrate the complexity of thiamin biochemistry, which is not restricted to the cofactor role of ThDP.

Characterisation of a thiamine diphosphate-dependent alpha-keto acid decarboxylase from Proteus mirabilis JN458.

Science.gov (United States)

Wang, Biying; Bai, Yajun; Fan, Taiping; Zheng, Xiaohui; Cai, Yujie

2017-10-01

Alpha-keto acid decarboxylases can convert keto acids to their corresponding aldehydes, which are often volatile aroma compounds. The gene encoding α-keto acid decarboxylase in Proteus mirabilis JN458 was cloned, and the enzyme overexpressed in Escherichia coli BL21 (DE3), purified in high yield, and characterised. The molecular weight is 62.291kDa by MALDI-TOF MS, and optimum activity at pH 6.0 and 40-50°C. The enzyme is a typical decarboxylase, dependent on thiamine diphosphate and Mg 2+ as cofactors. For the decarboxylation reaction, the enzyme displayed a broad substrate range. Kinetic parameters were determined using 4-methyl-2-oxopentanoic acid, phenyl pyruvate and 3-methyl-2-oxopentanoic acid as substrates. K m and k cat values for phenyl pyruvate were 0.62mM and 77.38s -1 , respectively, and the k cat /K m value was 124.81mM -1 s -1 . The enzyme properties suggest it may act effectively under cheese ripening conditions. Copyright © 2017. Published by Elsevier Ltd.

Co-expression of peppermint geranyl diphosphate synthase small subunit enhances monoterpene production in transgenic tobacco plants.

Science.gov (United States)

Yin, Jun-Lin; Wong, Woon-Seng; Jang, In-Cheol; Chua, Nam-Hai

2017-02-01

Monoterpenes are important for plant survival and useful to humans. In addition to their function in plant defense, monoterpenes are also used as flavors, fragrances and medicines. Several metabolic engineering strategies have been explored to produce monoterpene in tobacco but only trace amounts of monoterpenes have been detected. We investigated the effects of Solanum lycopersicum 1-deoxy-d-xylulose-5-phosphate synthase (SlDXS), Arabidopsis thaliana geranyl diphosphate synthase 1 (AtGPS) and Mentha × piperita geranyl diphosphate synthase small subunit (MpGPS.SSU) on production of monoterpene and geranylgeranyl diphosphate (GGPP) diversities, and plant morphology by transient expression in Nicotiana benthamiana and overexpression in transgenic Nicotiana tabacum. We showed that MpGPS.SSU could enhance the production of various monoterpenes such as (-)-limonene, (-)-linalool, (-)-α-pinene/β-pinene or myrcene, in transgenic tobacco by elevating geranyl diphosphate synthase (GPS) activity. In addition, overexpression of MpGPS.SSU in tobacco caused early flowering phenotype and increased shoot branching by elevating contents of GA 3 and cytokinins due to upregulated transcript levels of several plastidic 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway genes, geranylgeranyl diphosphate synthases 3 (GGPPS3) and GGPPS4. Our method would allow the identification of new monoterpene synthase genes using transient expression in N. benthamiana and the improvement of monoterpene production in transgenic tobacco plants. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

Poly(adenosine 5'-diphosphate) ribose polymerase activation as a cause of metabolic dysfunction in critical illness.

Science.gov (United States)

Liaudet, Lucas

2002-03-01

Poly(adenosine 5'-diphosphate) ribose polymerase is a nuclear enzyme activated in response to genotoxic stress induced by a variety of DNA damaging agents. Several oxygen and nitrogen-centered free radicals, notably peroxynitrite, are strong inducers of DNA damage and poly(adenosine 5'-diphosphate) ribose polymerase activation in vitro and in vivo. Activation of this nuclear enzyme depletes the intracellular stores of its substrate nicotinamide adenine dinucleotide, slowing the rate of glycolysis, mitochondrial electron transport and adenosine triphosphate formation. This process triggers a severe energetic crisis within the cell, leading to acute cell dysfunction and cell necrosis. Poly(adenosine 5'-diphosphate) ribose polymerase also plays an important role in the regulation of inflammatory cascades, through a functional association with various transcription factors and transcription co-activators. Recent works identified this enzyme as a critical mediator of cellular metabolic dysfunction, inflammatory injury, and organ damage in conditions associated with overwhelming oxidative stress, including systemic inflammation, circulatory shock, and ischemia-reperfusion. Accordingly, pharmacological inhibitors of poly(adenosine 5'-diphosphate) ribose polymerase protect against cell death and tissue injury in such conditions, and may therefore represent novel therapeutic tools to limit multiple organ damage and dysfunction in critically ill patients.

Crystallization and preliminary X ray analysis of nucleoside diphosphate kinase 1 from T. cruzi

International Nuclear Information System (INIS)

Gomez Barroso, J.A.; Aguilar, C.F.; Miranda, M.R.; Pereira, C.A.

2009-01-01

Introduction: Trypanosoma cruzi is the etiologic agent of Chagas disease. The Nucleoside diphosphate kinases (NDPKs) are enzymes involved in energy management and nucleoside balance in the cell. T. cruzi TcNDPK1, a canonical isoform. The objective of this work is obtaining protein's crystals, diffract and process the data for tridimensional structure resolution. Materials and Methods: TcNDPK1 was expressed in E. coli as a fusion protein with Nterminal His-tag. TcNDPK1 was overexpressed and purified by FPLC. Crystallization was assayed by sitting drop and hanging drop vapor diffusion method. Crystals was frozen and diffracted on synchrotron x-ray radiation in Campinas (Brasil). The data set collected was reduced and merged using MOSFLM and SCALA programs. Results and Discussion: His-TcNDPK was overexpressed, purified and crystallized. The crystals are diffracted and collected the data to 3.5A. The crystals belong to the trigonal space group P3, with unit cell parameters a=127.94, b=127.84, c=275.49. Structure determination is under way. These results will provide relevant information that could be the first step in rational drug design for treating Chagas disease.(authors)

The 1-hydroxy-2-methyl-butenyl 4-diphosphate reductase gene from ...

African Journals Online (AJOL)

The 1-hydroxy-2-methyl-butenyl 4-diphosphate reductase gene from Taxus media: Cloning, characterization and functional identification. Y Sun, M Chen, J Tang, W Liu, C Yang, Y Yang, X Lan, M Hsieh, Z Liao ...

Density fluctuation in a screened Coulombic colloid dispersion: comparison of the liquid and cubic phases of lipid A-diphosphate

International Nuclear Information System (INIS)

Brown, Helen; Ross, D. Keith; Paradies, Henrich H.

2004-01-01

Light-, small-angle X-ray and neutron scattering measurements of the dynamic structure factor S(Q,t) of strong interacting dispersions of lipid A-diphosphate were recorded and analysed applying existing models of liquid state theory. Lipid A-diphosphate ordering was observed at low volume fractions (phi=2.2x10 -4 ) and at very low ionic strength (I=10 -5 M). Upon increasing the particle number density of lipid A-diphosphate a transformation of the lattices of the colloidal crystals from a BCC lattice (a=36.20 nm) to a FCC lattice (a=57.30 nm) occurred. This strongly suggests a similarity in the preformed liquid structure and the cubic colloidal phase. The fit of both S eff (Q) and the principle peak I p (Q) with the effective particle charge supports of the main conclusions drawn from the SANS experiments and the liquid state theory indicating the presence of long-range order for the dispersions of lipid A-diphosphate

Trypanosoma brucei solanesyl-diphosphate synthase localizes to the mitochondrion

Czech Academy of Sciences Publication Activity Database

Lai, D.-H.; Bontempi, E. J.; Lukeš, Julius

2012-01-01

Roč. 183, č. 2 (2012), s. 189-192 ISSN 0166-6851 R&D Projects: GA ČR(CZ) GAP305/11/2179 Institutional support: RVO:60077344 Keywords : Trypanosoma brucei * Sleeping sickness * Ubiquinone * Solanesyl-diphosphate synthase * Digitonin permeabilization * In situ tagging Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.734, year: 2012 http://www.sciencedirect.com/science/article/pii/S0166685112000539

Cytosolic monoterpene biosynthesis is supported by plastid-generated geranyl diphosphate substrate in transgenic tomato fruits.

Science.gov (United States)

Gutensohn, Michael; Orlova, Irina; Nguyen, Thuong T H; Davidovich-Rikanati, Rachel; Ferruzzi, Mario G; Sitrit, Yaron; Lewinsohn, Efraim; Pichersky, Eran; Dudareva, Natalia

2013-08-01

Geranyl diphosphate (GPP), the precursor of most monoterpenes, is synthesized in plastids from dimethylallyl diphosphate and isopentenyl diphosphate by GPP synthases (GPPSs). In heterodimeric GPPSs, a non-catalytic small subunit (GPPS-SSU) interacts with a catalytic large subunit, such as geranylgeranyl diphosphate synthase, and determines its product specificity. Here, snapdragon (Antirrhinum majus) GPPS-SSU was over-expressed in tomato fruits under the control of the fruit ripening-specific polygalacturonase promoter to divert the metabolic flux from carotenoid formation towards GPP and monoterpene biosynthesis. Transgenic tomato fruits produced monoterpenes, including geraniol, geranial, neral, citronellol and citronellal, while exhibiting reduced carotenoid content. Co-expression of the Ocimum basilicum geraniol synthase (GES) gene with snapdragon GPPS-SSU led to a more than threefold increase in monoterpene formation in tomato fruits relative to the parental GES line, indicating that the produced GPP can be used by plastidic monoterpene synthases. Co-expression of snapdragon GPPS-SSU with the O. basilicum α-zingiberene synthase (ZIS) gene encoding a cytosolic terpene synthase that has been shown to possess both sesqui- and monoterpene synthase activities resulted in increased levels of ZIS-derived monoterpene products compared to fruits expressing ZIS alone. These results suggest that re-direction of the metabolic flux towards GPP in plastids also increases the cytosolic pool of GPP available for monoterpene synthesis in this compartment via GPP export from plastids. © 2013 The Authors The Plant Journal © 2013 John Wiley & Sons Ltd.

Two solanesyl diphosphate synthases with different subcellular localizations and their respective physiological roles in Oryza sativa.

Science.gov (United States)

Ohara, Kazuaki; Sasaki, Kanako; Yazaki, Kazufumi

2010-06-01

Long chain prenyl diphosphates are crucial biosynthetic precursors of ubiquinone (UQ) in many organisms, ranging from bacteria to humans, as well as precursors of plastoquinone in photosynthetic organisms. The cloning and characterization of two solanesyl diphosphate synthase genes, OsSPS1 and OsSPS2, in Oryza sativa is reported here. OsSPS1 was highly expressed in root tissue whereas OsSPS2 was found to be high in both leaves and roots. Enzymatic characterization using recombinant proteins showed that both OsSPS1 and OsSPS2 could produce solanesyl diphosphates as their final product, while OsSPS1 showed stronger activity than OsSPS2. However, an important biological difference was observed between the two genes: OsSPS1 complemented the yeast coq1 disruptant, which does not form UQ, whereas OsSPS2 only very weakly complemented the growth defect of the coq1 mutant. HPLC analyses showed that both OsSPS1 and OsSPS2 yeast transformants produced UQ9 instead of UQ6, which is the native yeast UQ. According to the complementation study, the UQ9 levels in OsSPS2 transformants were much lower than that of OsSPS1. Green fluorescent protein fusion analyses showed that OsSPS1 localized to mitochondria, while OsSPS2 localized to plastids. This suggests that OsSPS1 is involved in the supply of solanesyl diphosphate for ubiquinone-9 biosynthesis in mitochondria, whereas OsSPS2 is involved in providing solanesyl diphosphate for plastoquinone-9 formation. These findings indicate that O. sativa has a different mechanism for the supply of isoprenoid precursors in UQ biosynthesis from Arabidopsis thaliana, in which SPS1 provides a prenyl moiety for UQ9 at the endoplasmic reticulum.

Spectroscopic and Computational Investigations of Ligand Binding to IspH: Discovery of Non-diphosphate Inhibitors

Energy Technology Data Exchange (ETDEWEB)

O' Dowd, Bing [Department of Chemistry, University of Illinois, 600 South Mathews Avenue Urbana IL 61801 USA; Williams, Sarah [Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla CA 92093 USA; Wang, Hongxin [Department of Chemistry, University of California, 1 Shields Avenue Davis CA 95616 USA; Lawrence Berkeley National Laboratory, 1 Cyclotron Road Berkeley CA 94720 USA; No, Joo Hwan [Center for Biophysics and Computational Biology, Urbana, IL (United States); Rao, Guodong [Department of Chemistry, University of Illinois, 600 South Mathews Avenue Urbana IL 61801 USA; Wang, Weixue [Center for Biophysics and Computational Biology, Urbana, IL (United States); McCammon, J. Andrew [Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla CA 92093 USA; Howard Hughes Medical Institute, University of California at San Diego, La Jolla CA 92093 USA; National Biomedical Computation Resource, University of California at San Diego, La Jolla CA 92093 USA; Cramer, Stephen P. [Department of Chemistry, University of California, 1 Shields Avenue Davis CA 95616 USA; Lawrence Berkeley National Laboratory, 1 Cyclotron Road Berkeley CA 94720 USA; Oldfield, Eric [Department of Chemistry, University of Illinois, 600 South Mathews Avenue Urbana IL 61801 USA

2017-04-07

Isoprenoid biosynthesis is an important area for anti-infective drug development. One isoprenoid target described is (E)-1-hydroxy-2-methyl-but-2-enyl 4-diphosphate (HMBPP) reductase (IspH), which forms isopentenyl diphosphate and dimethylallyl diphosphate from HMBPP in a 2H + /2e - reduction. IspH contains a 4 Fe-4 S cluster, and in this work, we first investigated how small molecules bound to the cluster by using HYSCORE and NRVS spectroscopies. The results of these, as well as other structural and spectroscopic investigations, led to the conclusion that, in most cases, ligands bound to IspH 4 Fe-4 S clusters by η 1 coordination, forming tetrahedral geometries at the unique fourth Fe, ligand side chains preventing further ligand (e.g., H 2 O, O 2 ) binding. Based on these ideas, we used in silico methods to find drug-like inhibitors that might occupy the HMBPP substrate binding pocket and bind to Fe, leading to the discovery of a barbituric acid analogue with a K i value of ≈500 nm against Pseudomonas aeruginosa IspH.

Evaluation of the sorption of Eu(III) in titanium diphosphate; Evaluacion de la sorcion de Eu(III) en difosfato de titanio

Energy Technology Data Exchange (ETDEWEB)

Ortiz O, H.B.; Ordonez R, E.; Fernandez V, S.M. [ININ, Carretera Mexico-Toluca Km 36.5, Salazar, Estado de Mexico (Mexico)]. e-mail: hortiz@nuclear.inin.mx

2007-07-01

In this work its are presented: the synthesis, physicochemical characterization and the surface parameters estimation that can be related with the retention properties of the titanium diphosphate for the actinides of valence III (Pu, Am, Cm among others), using the Eu{sup 3+} like a chemical analog. The surface area, hydration time, zero charge point, density of active sites and the surface species distribution in the titanium diphosphate are reported. This information was used to explain the retention of the Eu(lll) in the surface of the titanium diphosphate. (Author)

Engineering the productivity of recombinant Escherichia coli for limonene formation from glycerol in minimal media.

Science.gov (United States)

Willrodt, Christian; David, Christian; Cornelissen, Sjef; Bühler, Bruno; Julsing, Mattijs K; Schmid, Andreas

2014-08-01

The efficiency and productivity of cellular biocatalysts play a key role in the industrial synthesis of fine and bulk chemicals. This study focuses on optimizing the synthesis of (S)-limonene from glycerol and glucose as carbon sources using recombinant Escherichia coli. The cyclic monoterpene limonene is extensively used in the fragrance, food, and cosmetic industries. Recently, limonene also gained interest as alternative jet fuel of biological origin. Key parameters that limit the (S)-limonene yield, related to genetics, physiology, and reaction engineering, were identified. The growth-dependent production of (S)-limonene was shown for the first time in minimal media. E. coli BL21 (DE3) was chosen as the preferred host strain, as it showed low acetate formation, fast growth, and high productivity. A two-liquid phase fed-batch fermentation with glucose as the sole carbon and energy source resulted in the formation of 700 mg L(org) (-1) (S)-limonene. Specific activities of 75 mU g(cdw) (-1) were reached, but decreased relatively quickly. The use of glycerol as a carbon source resulted in a prolonged growth and production phase (specific activities of ≥50 mU g(cdw) (-1) ) leading to a final (S)-limonene concentration of 2,700 mg L(org) (-1) . Although geranyl diphosphate (GPP) synthase had a low solubility, its availability appeared not to limit (S)-limonene formation in vivo under the conditions investigated. GPP rerouting towards endogenous farnesyl diphosphate (FPP) formation also did not limit (S)-limonene production. The two-liquid phase fed-batch setup led to the highest monoterpene concentration obtained with a recombinant microbial biocatalyst to date. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Synthesis of isoprenoid bisphosphonate ethers through C–P bond formations: Potential inhibitors of geranylgeranyl diphosphate synthase

Directory of Open Access Journals (Sweden)

Xiang Zhou

2014-07-01

Full Text Available A set of bisphosphonate ethers has been prepared through sequential phosphonylation and alkylation of monophosphonate ethers. After formation of the corresponding phosphonic acid salts, these compounds were tested for their ability to inhibit the enzyme geranylgeranyl diphosphate synthase (GGDPS. Five of the new compounds show IC50 values of less than 1 μM against GGDPS with little to no activity against the related enzyme farnesyl diphosphate synthase (FDPS. The most active compound displayed an IC50 value of 82 nM when assayed with GGDPS, and no activity against FDPS even at a 10 μM concentration.

A new strategy for the cloning, overexpression and one step purification of three DHAP-dependent aldolases: rhamnulose-1-phosphate aldolase, fuculose-1-phosphate aldolase and tagatose-1,6-diphosphate aldolase.

Science.gov (United States)

Garcia-Junceda, E; Shen, G J; Sugai, T; Wong, C H

1995-07-01

Three DHAP-dependent aldolases, rhamnulose-1-phosphate aldolase (Rham-1PA), fuculose-1-phosphate aldolase (Fuc-1PA) and tagatose-1,6-diphosphate aldolase (TDPA) have been cloned and overexpressed in Escherichia coli using two different expression vectors: pTrcHis for the expression of Rham-1PA and Fuc-1PA and pRSET for the expression of TDPA. In each case the recombinant enzyme is synthesized as a fusion protein with a hexahistidine tag on the N-terminus. The three enzymes have been purified in only one step by chelation affinity chromatography. The effects of cultivation temperature and concentration of inducer have been studied in order to optimize the expression of the recombinant proteins and to avoid the formation of inclusion bodies.

Cloning and Characterization of Farnesyl Diphosphate Synthase Gene Involved in Triterpenoids Biosynthesis from Poria cocos

Directory of Open Access Journals (Sweden)

Jianrong Wang

2014-12-01

Full Text Available Poria cocos (P. cocos has long been used as traditional Chinese medicine and triterpenoids are the most important pharmacologically active constituents of this fungus. Farnesyl pyrophosphate synthase (FPS is a key enzyme of triterpenoids biosynthesis. The gene encoding FPS was cloned from P. cocos by degenerate PCR, inverse PCR and cassette PCR. The open reading frame of the gene is 1086 bp in length, corresponding to a predicted polypeptide of 361 amino acid residues with a molecular weight of 41.2 kDa. Comparison of the P. cocos FPS deduced amino acid sequence with other species showed the highest identity with Ganoderma lucidum (74%. The predicted P. cocos FPS shares at least four conserved regions involved in the enzymatic activity with the FPSs of varied species. The recombinant protein was expressed in Pichia pastoris and purified. Gas chromatography analysis showed that the recombinant FPS could catalyze the formation of farnesyl diphosphate (FPP from geranyl diphosphate (GPP and isopentenyl diphosphate (IPP. Furthermore, the expression profile of the FPS gene and content of total triterpenoids under different stages of development and methyl jasmonate treatments were determined. The results indicated that there is a positive correlation between the activity of FPS and the amount of total triterpenoids produced in P. cocos.

Optimization of primaquine diphosphate tablet formulation for controlled drug release using the mixture experimental design.

Science.gov (United States)

Duque, Marcelo Dutra; Kreidel, Rogério Nepomuceno; Taqueda, Maria Elena Santos; Baby, André Rolim; Kaneko, Telma Mary; Velasco, Maria Valéria Robles; Consiglieri, Vladi Olga

2013-01-01

A tablet formulation based on hydrophilic matrix with a controlled drug release was developed, and the effect of polymer concentrations on the release of primaquine diphosphate was evaluated. To achieve this purpose, a 20-run, four-factor with multiple constraints on the proportions of the components was employed to obtain tablet compositions. Drug release was determined by an in vitro dissolution study in phosphate buffer solution at pH 6.8. The polynomial fitted functions described the behavior of the mixture on simplex coordinate systems to study the effects of each factor (polymer) on tablet characteristics. Based on the response surface methodology, a tablet composition was optimized with the purpose of obtaining a primaquine diphosphate release closer to a zero order kinetic. This formulation released 85.22% of the drug for 8 h and its kinetic was studied regarding to Korsmeyer-Peppas model, (Adj-R(2) = 0.99295) which has confirmed that both diffusion and erosion were related to the mechanism of the drug release. The data from the optimized formulation were very close to the predictions from statistical analysis, demonstrating that mixture experimental design could be used to optimize primaquine diphosphate dissolution from hidroxypropylmethyl cellulose and polyethylene glycol matrix tablets.

ACTIVATION OF G-PROTEINS BY RECEPTOR-STIMULATED NUCLEOSIDE DIPHOSPHATE KINASE IN DICTYOSTELIUM

NARCIS (Netherlands)

Bominaar, Anthony A.; Molijn, Anco C.; Pestel, Martine; Veron, Michel; Haastert, Peter J.M. van

Recently, interest in the enzyme nucleoside diphosphate kinase (EC 2.7.4.6) has increased as a result of its possible involvement in cell proliferation and development. Since NDP kinase is one of the major sources of GTP in cells, it has been suggested that the effects of an altered NDP kinase

Crystal Structures of Staphylococcus epidermidis Mevalonate Diphosphate Decarboxylase Bound to Inhibitory Analogs Reveal New Insight into Substrate Binding and Catalysis

Energy Technology Data Exchange (ETDEWEB)

Barta, Michael L.; Skaff, D. Andrew; McWhorter, William J.; Herdendorf, Timothy J.; Miziorko, Henry M.; Geisbrecht, Brian V. (UMKC)

2011-10-28

The polyisoprenoid compound undecaprenyl phosphate is required for biosynthesis of cell wall peptidoglycans in Gram-positive bacteria, including pathogenic Enterococcus, Streptococcus, and Staphylococcus spp. In these organisms, the mevalonate pathway is used to produce the precursor isoprenoid, isopentenyl 5-diphosphate. Mevalonate diphosphate decarboxylase (MDD) catalyzes formation of isopentenyl 5-diphosphate in an ATP-dependent irreversible reaction and is therefore an attractive target for inhibitor development that could lead to new antimicrobial agents. To facilitate exploration of this possibility, we report the crystal structure of Staphylococcus epidermidis MDD (1.85 {angstrom} resolution) and, to the best of our knowledge, the first structures of liganded MDD. These structures include MDD bound to the mevalonate 5-diphosphate analogs diphosphoglycolyl proline (2.05 {angstrom} resolution) and 6-fluoromevalonate diphosphate (FMVAPP; 2.2 {angstrom} resolution). Comparison of these structures provides a physical basis for the significant differences in K{sub i} values observed for these inhibitors. Inspection of enzyme/inhibitor structures identified the side chain of invariant Ser{sup 192} as making potential contributions to catalysis. Significantly, Ser {yields} Ala substitution of this side chain decreases k{sub cat} by {approx}10{sup 3}-fold, even though binding interactions between FMVAPP and this mutant are similar to those observed with wild type MDD, as judged by the 2.1 {angstrom} cocrystal structure of S192A with FMVAPP. Comparison of microbial MDD structures with those of mammalian counterparts reveals potential targets at the active site periphery that may be exploited to selectively target the microbial enzymes. These studies provide a structural basis for previous observations regarding the MDD mechanism and inform future work toward rational inhibitor design.

Structural Basis for Nucleotide Binding and Reaction Catalysis in Mevalonate Diphosphate Decarboxylase

Energy Technology Data Exchange (ETDEWEB)

Barta, Michael L.; McWhorter, William J.; Miziorko, Henry M.; Geisbrecht, Brian V. (UMKC)

2012-09-17

Mevalonate diphosphate decarboxylase (MDD) catalyzes the final step of the mevalonate pathway, the Mg{sup 2+}-ATP dependent decarboxylation of mevalonate 5-diphosphate (MVAPP), producing isopentenyl diphosphate (IPP). Synthesis of IPP, an isoprenoid precursor molecule that is a critical intermediate in peptidoglycan and polyisoprenoid biosynthesis, is essential in Gram-positive bacteria (e.g., Staphylococcus, Streptococcus, and Enterococcus spp.), and thus the enzymes of the mevalonate pathway are ideal antimicrobial targets. MDD belongs to the GHMP superfamily of metabolite kinases that have been extensively studied for the past 50 years, yet the crystallization of GHMP kinase ternary complexes has proven to be difficult. To further our understanding of the catalytic mechanism of GHMP kinases with the purpose of developing broad spectrum antimicrobial agents that target the substrate and nucleotide binding sites, we report the crystal structures of wild-type and mutant (S192A and D283A) ternary complexes of Staphylococcus epidermidis MDD. Comparison of apo, MVAPP-bound, and ternary complex wild-type MDD provides structural information about the mode of substrate binding and the catalytic mechanism. Structural characterization of ternary complexes of catalytically deficient MDD S192A and D283A (k{sub cat} decreased 10{sup 3}- and 10{sup 5}-fold, respectively) provides insight into MDD function. The carboxylate side chain of invariant Asp{sup 283} functions as a catalytic base and is essential for the proper orientation of the MVAPP C3-hydroxyl group within the active site funnel. Several MDD amino acids within the conserved phosphate binding loop ('P-loop') provide key interactions, stabilizing the nucleotide triphosphoryl moiety. The crystal structures presented here provide a useful foundation for structure-based drug design.

A functional (E)-4-hydroxy-3-methylbut-2-enyl diphosphate reductase exhibits diurnal regulation of expression in Stevia rebaudiana (Bertoni).

Science.gov (United States)

Kumar, Hitesh; Kumar, Sanjay

2013-09-15

The leaves of stevia [Stevia rebaudiana (Bertoni)] are a rich source of steviol glycosides that are used as non-calorific sweetener in many countries around the world. Steviol moiety of steviol glycosides is synthesized via plastidial 2C-methyl-D-erythritol 4-phosphate pathway, where (E)-4-hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR) is the key enzyme. HDR catalyzes the simultaneous conversion of (E)-4-hydroxy-3-methylbut-2-enyl diphosphate into five carbon isoprenoid units, isopentenyl diphosphate and dimethylallyl diphosphate. Stevia HDR (SrHDR) successfully rescued HDR lethal mutant strain MG1655 araispH upon genetic complementation, suggesting SrHDR to encode a functional protein. The gene exhibited diurnal variation in expression. To identify the possible regulatory elements, upstream region of the gene was cloned and putative cis-acting elements were detected by in silico analysis. Electrophoretic mobility shift assay, using a putative light responsive element GATA showed the binding of nuclear proteins (NP) isolated from leaves during light period of the day, but not with the NP from leaves during the dark period. Data suggested the involvement of GATA box in light mediated gene regulation of SrHDR in stevia. Copyright © 2013 Elsevier B.V. All rights reserved.

Physiology of inactivation of microbial cells by near-ultraviolet light: mode of action and application for the enrichment of mutants of Escherichia coli and saccharomyces cerevisiae

International Nuclear Information System (INIS)

Peters, J.

1976-01-01

The mode of action of near-ultraviolet (NUV) light was studied in Escherichia coli. NUV light (maximum emission at 365 nm) caused the photodestruction of ribonucleoside diphosphate (RDP) reductase activity in vivo. Evidence was presented for a model suggesting that the loss of RDP-reductase resulted in a metabolic state analogous to that produced during starvation for thymine. Some important properties of cells irradiated by NUV light, cell death, loss of the ability to support the replication of DNA phages and a delay in the onset of cell division in sublethally irradiated cells, were accounted for in terms of photoinactivation of RDP-reductase. Conditions were described under which NUV light was an effective counterselective agent for the enrichment of mutants of Escherichia coli and Saccharomyces cerevisiae

Phosphorylation of nm23/nucleoside diphosphate kinase by casein kinase 2 in vitro

DEFF Research Database (Denmark)

Engel, M; Issinger, O G; Lascu, I

1994-01-01

We have investigated phosphorylation of human nucleoside diphosphate kinase (NDPK) and of homologous NDPK from different species by human casein kinase 2 (CK-2). The human NDPK isotypes A and B were phosphorylated by CK-2 in vitro both when the purified proteins and total lysate of HL-60 leukemia...

Modulation of guanosine nucleotides biosynthetic pathways enhanced GDP-L-fucose production in recombinant Escherichia coli.

Science.gov (United States)

Lee, Won-Heong; Shin, So-Yeon; Kim, Myoung-Dong; Han, Nam Soo; Seo, Jin-Ho

2012-03-01

Guanosine 5'-triphosphate (GTP) is the key substrate for biosynthesis of guanosine 5'-diphosphate (GDP)-L-fucose. In this study, improvement of GDP-L-fucose production was attempted by manipulating the biosynthetic pathway for guanosine nucleotides in recombinant Escherichia coli-producing GDP-L-fucose. The effects of overexpression of inosine 5'-monophosphate (IMP) dehydrogenase, guanosine 5'-monophosphate (GMP) synthetase (GuaB and GuaA), GMP reductase (GuaC) and guanosine-inosine kinase (Gsk) on GDP-L-fucose production were investigated in a series of fed-batch fermentations. Among the enzymes tested, overexpression of Gsk led to a significant improvement of GDP-L-fucose production. Maximum GDP-L-fucose concentration of 305.5 ± 5.3 mg l(-1) was obtained in the pH-stat fed-batch fermentation of recombinant E. coli-overexpressing Gsk, which corresponds to a 58% enhancement in the GDP-L-fucose production compared with the control strain overexpressing GDP-L-fucose biosynthetic enzymes. Such an enhancement of GDP-L-fucose production could be due to the increase in the intracellular level of GMP.

Metabolic engineering of monoterpene biosynthesis in tomato fruits via introduction of the non-canonical substrate neryl diphosphate.

Science.gov (United States)

Gutensohn, Michael; Nguyen, Thuong T H; McMahon, Richard D; Kaplan, Ian; Pichersky, Eran; Dudareva, Natalia

2014-07-01

Recently it was shown that monoterpenes in tomato trichomes (Solanum lycopersicum) are synthesized by phellandrene synthase 1 (PHS1) from the non-canonical substrate neryl diphosphate (NPP), the cis-isomer of geranyl diphosphate (GPP). As PHS1 accepts both NPP and GPP substrates forming different monoterpenes, it was overexpressed in tomato fruits to test if NPP is also available in a tissue highly active in carotenoid production. However, transgenic fruits overexpressing PHS1 produced only small amounts of GPP-derived PHS1 monoterpene products, indicating the absence of endogenous NPP. Therefore, NPP formation was achieved by diverting the metabolic flux from carotenoids via expression of tomato neryl diphosphate synthase 1 (NDPS1). NDPS1 transgenic fruits produced NPP-derived monoterpenes, including nerol, neral and geranial, while displaying reduced lycopene content. NDPS1 co-expression with PHS1 resulted in a monoterpene blend, including β-phellandrene, similar to that produced from NPP by PHS1 in vitro and in trichomes. Unexpectedly, PHS1×NDPS1 fruits showed recovery of lycopene levels compared to NDPS1 fruits, suggesting that redirection of metabolic flux is only partially responsible for the reduction in carotenoids. In vitro assays demonstrated that NPP serves as an inhibitor of geranylgeranyl diphosphate synthase, thus its consumption by PHS1 leads to recovery of lycopene levels. Monoterpenes produced in PHS1×NDPS1 fruits contributed to direct plant defense negatively affecting feeding behavior of the herbivore Helicoverpa zea and displaying antifungal activity against Botrytis cinerea. These results show that NPP-derived terpenoids can be produced in plant tissues; however, NPP has to be consumed to avoid negative impacts on plant metabolism. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Engineering of a plasmid-free Escherichia coli strain for improved in vivo biosynthesis of astaxanthin

Directory of Open Access Journals (Sweden)

Steuer Kristin

2011-04-01

Full Text Available Abstract Background The xanthophyll astaxanthin is a high-value compound with applications in the nutraceutical, cosmetic, food, and animal feed industries. Besides chemical synthesis and extraction from naturally producing organisms like Haematococcus pluvialis, heterologous biosynthesis in non-carotenogenic microorganisms like Escherichia coli, is a promising alternative for sustainable production of natural astaxanthin. Recent achievements in the metabolic engineering of E. coli strains have led to a significant increase in the productivity of carotenoids like lycopene or β-carotene by increasing the metabolic flux towards the isoprenoid precursors. For the heterologous biosynthesis of astaxanthin in E. coli, however, the conversion of β-carotene to astaxanthin is obviously the most critical step towards an efficient biosynthesis of astaxanthin. Results Here we report the construction of the first plasmid-free E. coli strain that produces astaxanthin as the sole carotenoid compound with a yield of 1.4 mg/g cdw (E. coli BW-ASTA. This engineered E. coli strain harbors xanthophyll biosynthetic genes from Pantoea ananatis and Nostoc punctiforme as individual expression cassettes on the chromosome and is based on a β-carotene-producing strain (E. coli BW-CARO recently developed in our lab. E. coli BW-CARO has an enhanced biosynthesis of the isoprenoid precursor isopentenyl diphosphate (IPP and produces β-carotene in a concentration of 6.2 mg/g cdw. The expression of crtEBIY along with the β-carotene-ketolase gene crtW148 (NpF4798 and the β-carotene-hydroxylase gene (crtZ under controlled expression conditions in E. coli BW-ASTA directed the pathway exclusively towards the desired product astaxanthin (1.4 mg/g cdw. Conclusions By using the λ-Red recombineering technique, genes encoding for the astaxanthin biosynthesis pathway were stably integrated into the chromosome of E. coli. The expression levels of chromosomal integrated recombinant

Triclinic-cubic phase transition and negative expansion in the actinide IV (Th, U, Np, Pu) diphosphates

International Nuclear Information System (INIS)

Wallez, Gilles; Bregiroux, Damien; Raison, Philippe E.; Bykov, Denis; Konings, Rudy J.M.; Dacheux, Nicolas; Clavier, Nicolas; Delevoye, Laurent; Popa, Karin; Fitch, Andrew N.

2012-01-01

The AnP 2 O 7 diphosphates (An = Th, U, Np, Pu) have been synthesized by various routes depending on the stability of the An(IV) cation and its suitability for the unusual octahedral environment. Synchrotron and X-ray diffraction, thermal analysis, Raman spectroscopy, and 31 P nuclear magnetic resonance reveal them as a new family of diphosphates which probably includes the recently studied CeP 2 O 7 . Although they adopt at high temperature the same cubic archetypal cell as the other known MP 2 O 7 diphosphates, they differ by a very faint triclinic distortion at room temperature that results from an ordering of the P 2 O 7 units, as shown using high-resolution synchrotron diffraction for UP 2 O 7 . The uncommon triclinic-cubic phase transition is first order, and its temperature is very sensitive to the ionic radius of An(IV). The conflicting effects which control the thermal variations of the P-O-P angle are responsible for a strong expansion of the cell followed by a contraction at higher temperature. This inversion of expansion occurs at a temperature significantly higher than the phase transition, at variance with the parent compounds with smaller Mn(IV) cations in which the two phenomena coincide. As shown by various approaches, the P-O-b-P linkage remains bent in the cubic form. (authors)

X-ray characteristics and thermal transformations of double diphosphates MLnP2O7 (M-K, Rb, Cs; Ln-REE of yttrium subgroup)

International Nuclear Information System (INIS)

Anisimova, N.Yu.; Chudinova, N.N.; Trunov, V.K.; AN SSSR, Moscow

1993-01-01

Results on studying double diphosphates of rare earth metals with K, Rb and Cs are studied by methods of X-ray analysis, high-temperature roentgenography and DTA. Double diphosphates of rare earth and alkali metals are crystallized in three structural types: monoclinic, rhombic and hexagonal, whereby rhombic form by heating inversely transforms into hexagonal one. MLnP 2 O 7 thermal transformations are studied

Suppressing Farnesyl Diphosphate Synthase Alters Chloroplast Development and Triggers Sterol-Dependent Induction of Jasmonate- and Fe-Related Responses.

Science.gov (United States)

Manzano, David; Andrade, Paola; Caudepón, Daniel; Altabella, Teresa; Arró, Montserrat; Ferrer, Albert

2016-09-01

Farnesyl diphosphate synthase (FPS) catalyzes the synthesis of farnesyl diphosphate from isopentenyl diphosphate and dimethylallyl diphosphate. Arabidopsis (Arabidopsis thaliana) contains two genes (FPS1 and FPS2) encoding FPS. Single fps1 and fps2 knockout mutants are phenotypically indistinguishable from wild-type plants, while fps1/fps2 double mutants are embryo lethal. To assess the effect of FPS down-regulation at postembryonic developmental stages, we generated Arabidopsis conditional knockdown mutants expressing artificial microRNAs devised to simultaneously silence both FPS genes. Induction of silencing from germination rapidly caused chlorosis and a strong developmental phenotype that led to seedling lethality. However, silencing of FPS after seed germination resulted in a slight developmental delay only, although leaves and cotyledons continued to show chlorosis and altered chloroplasts. Metabolomic analyses also revealed drastic changes in the profile of sterols, ubiquinones, and plastidial isoprenoids. RNA sequencing and reverse transcription-quantitative polymerase chain reaction transcriptomic analysis showed that a reduction in FPS activity levels triggers the misregulation of genes involved in biotic and abiotic stress responses, the most prominent one being the rapid induction of a set of genes related to the jasmonic acid pathway. Down-regulation of FPS also triggered an iron-deficiency transcriptional response that is consistent with the iron-deficient phenotype observed in FPS-silenced plants. The specific inhibition of the sterol biosynthesis pathway by chemical and genetic blockage mimicked these transcriptional responses, indicating that sterol depletion is the primary cause of the observed alterations. Our results highlight the importance of sterol homeostasis for normal chloroplast development and function and reveal important clues about how isoprenoid and sterol metabolism is integrated within plant physiology and development. © 2016

Retinoid production using metabolically engineered Escherichia coli with a two-phase culture system.

Science.gov (United States)

Jang, Hui-Jeong; Yoon, Sang-Hwal; Ryu, Hee-Kyung; Kim, Jung-Hun; Wang, Chong-Long; Kim, Jae-Yean; Oh, Deok-Kun; Kim, Seon-Won

2011-07-29

Retinoids are lipophilic isoprenoids composed of a cyclic group and a linear chain with a hydrophilic end group. These compounds include retinol, retinal, retinoic acid, retinyl esters, and various derivatives of these structures. Retinoids are used as cosmetic agents and effective pharmaceuticals for skin diseases. Retinal, an immediate precursor of retinoids, is derived by β-carotene 15,15'-mono(di)oxygenase (BCM(D)O) from β-carotene, which is synthesized from the isoprenoid building blocks isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). Retinoids are chemically unstable and biologically degraded via retinoic acid. Although extensive studies have been performed on the microbial production of carotenoids, retinoid production using microbial metabolic engineering has not been reported. Here, we report retinoid production using engineered Escherichia coli that express exogenous BCM(D)O and the mevalonate (MVA) pathway for the building blocks synthesis in combination with a two-phase culture system using a dodecane overlay. Among the BCM(D)O tested in E. coli, the synthetic retinoid synthesis protein (SR), based on bacteriorhodopsin-related protein-like homolog (Blh) of the uncultured marine bacteria 66A03, showed the highest β-carotene cleavage activity with no residual intracellular β-carotene. By introducing the exogenous MVA pathway, 8.7 mg/L of retinal was produced, which is 4-fold higher production than that of augmenting the MEP pathway (dxs overexpression). There was a large gap between retinal production and β-carotene consumption using the exogenous MVA pathway; therefore, the retinal derivatives were analyzed. The derivatives, except for retinoic acid, that formed were identified, and the levels of retinal, retinol, and retinyl acetate were measured. Amounts as high as 95 mg/L retinoids were obtained from engineered E. coli DH5α harboring the synthetic SR gene and the exogenous MVA pathway in addition to dxs overexpression, which

Functional specialization of one copy of glutamine phosphoribosyl pyrophosphate amidotransferase in ureide production from symbiotically fixed nitrogen in Phaseolus vulgaris.

Science.gov (United States)

Coleto, Inmaculada; Trenas, Almudena T; Erban, Alexander; Kopka, Joachim; Pineda, Manuel; Alamillo, Josefa M

2016-08-01

Purines are essential molecules formed in a highly regulated pathway in all organisms. In tropical legumes, the nitrogen fixed in the nodules is used to generate ureides through the oxidation of de novo synthesized purines. Glutamine phosphoribosyl pyrophosphate amidotransferase (PRAT) catalyses the first committed step of de novo purine synthesis. In Phaseolus vulgaris there are three genes coding for PRAT. The three full-length sequences, which are intron-less genes, were cloned, and their expression levels were determined under conditions that affect the synthesis of purines. One of the three genes, PvPRAT3, is highly expressed in nodules and protein amount and enzymatic activity in these tissues correlate with nitrogen fixation activity. Inhibition of PvPRAT3 gene expression by RNAi-silencing and subsequent metabolomic analysis of the transformed roots shows that PvPRAT3 is essential for the synthesis of ureides in P. vulgaris nodules. © 2016 John Wiley & Sons Ltd.

Insights into the Thiamine Diphosphate Enzyme Activation Mechanism: Computational Model for Transketolase Using a Quantum Mechanical/Molecular Mechanical Method.

Science.gov (United States)

Nauton, Lionel; Hélaine, Virgil; Théry, Vincent; Hecquet, Laurence

2016-04-12

We propose the first computational model for transketolase (TK), a thiamine diphosphate (ThDP)-dependent enzyme, using a quantum mechanical/molecular mechanical method on the basis of crystallographic TK structures from yeast and Escherichia coli, together with experimental kinetic data reported in the literature with wild-type and mutant TK. This model allowed us to define a new route for ThDP activation in the enzyme environment. We evidenced a strong interaction between ThDP and Glu418B of the TK active site, itself stabilized by Glu162A. The crucial point highlighted here is that deprotonation of ThDP C2 is not performed by ThDP N4' as reported in the literature, but by His481B, involving a HOH688A molecule bridge. Thus, ThDP N4' is converted from an amino form to an iminium form, ensuring the stabilization of the C2 carbanion or carbene. Finally, ThDP activation proceeds via an intermolecular process and not by an intramolecular one as reported in the literature. More generally, this proposed ThDP activation mechanism can be applied to some other ThDP-dependent enzymes and used to define the entire TK mechanism with donor and acceptor substrates more accurately.

Suppressing Farnesyl Diphosphate Synthase Alters Chloroplast Development and Triggers Sterol-Dependent Induction of Jasmonate- and Fe-Related Responses1[OPEN

Science.gov (United States)

Andrade, Paola; Caudepón, Daniel; Arró, Montserrat

2016-01-01

Farnesyl diphosphate synthase (FPS) catalyzes the synthesis of farnesyl diphosphate from isopentenyl diphosphate and dimethylallyl diphosphate. Arabidopsis (Arabidopsis thaliana) contains two genes (FPS1 and FPS2) encoding FPS. Single fps1 and fps2 knockout mutants are phenotypically indistinguishable from wild-type plants, while fps1/fps2 double mutants are embryo lethal. To assess the effect of FPS down-regulation at postembryonic developmental stages, we generated Arabidopsis conditional knockdown mutants expressing artificial microRNAs devised to simultaneously silence both FPS genes. Induction of silencing from germination rapidly caused chlorosis and a strong developmental phenotype that led to seedling lethality. However, silencing of FPS after seed germination resulted in a slight developmental delay only, although leaves and cotyledons continued to show chlorosis and altered chloroplasts. Metabolomic analyses also revealed drastic changes in the profile of sterols, ubiquinones, and plastidial isoprenoids. RNA sequencing and reverse transcription-quantitative polymerase chain reaction transcriptomic analysis showed that a reduction in FPS activity levels triggers the misregulation of genes involved in biotic and abiotic stress responses, the most prominent one being the rapid induction of a set of genes related to the jasmonic acid pathway. Down-regulation of FPS also triggered an iron-deficiency transcriptional response that is consistent with the iron-deficient phenotype observed in FPS-silenced plants. The specific inhibition of the sterol biosynthesis pathway by chemical and genetic blockage mimicked these transcriptional responses, indicating that sterol depletion is the primary cause of the observed alterations. Our results highlight the importance of sterol homeostasis for normal chloroplast development and function and reveal important clues about how isoprenoid and sterol metabolism is integrated within plant physiology and development. PMID

Structure of the ent-Copalyl Diphosphate Synthase PtmT2 from Streptomyces platensis CB00739, a Bacterial Type II Diterpene Synthase.

Science.gov (United States)

Rudolf, Jeffrey D; Dong, Liao-Bin; Cao, Hongnan; Hatzos-Skintges, Catherine; Osipiuk, Jerzy; Endres, Michael; Chang, Chin-Yuan; Ma, Ming; Babnigg, Gyorgy; Joachimiak, Andrzej; Phillips, George N; Shen, Ben

2016-08-31

Terpenoids are the largest and most structurally diverse family of natural products found in nature, yet their presence in bacteria is underappreciated. The carbon skeletons of terpenoids are generated through carbocation-dependent cyclization cascades catalyzed by terpene synthases (TSs). Type I and type II TSs initiate cyclization via diphosphate ionization and protonation, respectively, and protein structures of both types are known. Most plant diterpene synthases (DTSs) possess three α-helical domains (αβγ), which are thought to have arisen from the fusion of discrete, ancestral bacterial type I TSs (α) and type II TSs (βγ). Type II DTSs of bacterial origin, of which there are no structurally characterized members, are a missing piece in the structural evolution of TSs. Here, we report the first crystal structure of a type II DTS from bacteria. PtmT2 from Streptomyces platensis CB00739 was verified as an ent-copalyl diphosphate synthase involved in the biosynthesis of platensimycin and platencin. The crystal structure of PtmT2 was solved at a resolution of 1.80 Å, and docking studies suggest the catalytically active conformation of geranylgeranyl diphosphate (GGPP). Site-directed mutagenesis confirmed residues involved in binding the diphosphate moiety of GGPP and identified DxxxxE as a potential Mg(2+)-binding motif for type II DTSs of bacterial origin. Finally, both the shape and physicochemical properties of the active sites are responsible for determining specific catalytic outcomes of TSs. The structure of PtmT2 fundamentally advances the knowledge of bacterial TSs, their mechanisms, and their role in the evolution of TSs.

Crystal structure of heterodimeric hexaprenyl diphosphate synthase from Micrococcus luteus B-P 26 reveals that the small subunit is directly involved in the product chain length regulation.

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Sasaki, Daisuke; Fujihashi, Masahiro; Okuyama, Naomi; Kobayashi, Yukiko; Noike, Motoyoshi; Koyama, Tanetoshi; Miki, Kunio

2011-02-04

Hexaprenyl diphosphate synthase from Micrococcus luteus B-P 26 (Ml-HexPPs) is a heterooligomeric type trans-prenyltransferase catalyzing consecutive head-to-tail condensations of three molecules of isopentenyl diphosphates (C(5)) on a farnesyl diphosphate (FPP; C(15)) to form an (all-E) hexaprenyl diphosphate (HexPP; C(30)). Ml-HexPPs is known to function as a heterodimer of two different subunits, small and large subunits called HexA and HexB, respectively. Compared with homooligomeric trans-prenyltransferases, the molecular mechanism of heterooligomeric trans-prenyltransferases is not yet clearly understood, particularly with respect to the role of the small subunits lacking the catalytic motifs conserved in most known trans-prenyltransferases. We have determined the crystal structure of Ml-HexPPs both in the substrate-free form and in complex with 7,11-dimethyl-2,6,10-dodecatrien-1-yl diphosphate ammonium salt (3-DesMe-FPP), an analog of FPP. The structure of HexB is composed of mostly antiparallel α-helices joined by connecting loops. Two aspartate-rich motifs (designated the first and second aspartate-rich motifs) and the other characteristic motifs in HexB are located around the diphosphate part of 3-DesMe-FPP. Despite the very low amino acid sequence identity and the distinct polypeptide chain lengths between HexA and HexB, the structure of HexA is quite similar to that of HexB. The aliphatic tail of 3-DesMe-FPP is accommodated in a large hydrophobic cleft starting from HexB and penetrating to the inside of HexA. These structural features suggest that HexB catalyzes the condensation reactions and that HexA is directly involved in the product chain length control in cooperation with HexB.

Early Cessation of Adenosine Diphosphate Receptor Inhibitors Among Acute Myocardial Infarction Patients Treated With Percutaneous Coronary Intervention

DEFF Research Database (Denmark)

Fosbøl, Emil L; Ju, Christine; Anstrom, Kevin J

2016-01-01

BACKGROUND: Guidelines recommend the use of adenosine diphosphate receptor inhibitor (ADPri) therapy for 1 year postacute myocardial infarction; yet, early cessation of therapy occurs frequently in clinical practice. METHODS AND RESULTS: We examined 11 858 acute myocardial infarction patients tre...

Influence of the temperature in the uranium (Vi) sorption in zirconium diphosphate

International Nuclear Information System (INIS)

Garcia G, N.; Solis, D.; Ordonez R, E.

2012-10-01

In the present work was evaluated the uranium (Vi) sorption at 10, 20, 30, 40 and 60 C on the zirconium diphosphate (ZrP 2 O 7 ). They were carried out kinetic and isotherms using the method by lots, these will allow to fix the sorption time (kinetic) and to explain the behavior of this sorption in different ph conditions and temperature (isotherm). The quantity of retained uranium in the surface was quantified by means of the fluorescence technique. (Author)

Isolation and characterization of a copalyl diphosphate synthase gene promoter from Salvia miltiorrhiza

Directory of Open Access Journals (Sweden)

Piotr Szymczyk

2016-09-01

Full Text Available The promoter, 5' UTR, and 34-nt 5' fragments of protein encoding region of the Salvia miltiorrhiza copalyl diphosphate synthase gene were cloned and characterized. No tandem repeats, miRNA binding sites, or CpNpG islands were observed in the promoter, 5' UTR, or protein encoding fragments. The entire isolated promoter and 5' UTR is 2235 bp long and contains repetitions of many cis-active elements, recognized by homologous transcription factors, found in Arabidopsis thaliana and other plant species. A pyrimidine-rich fragment with only 6 non-pyrimidine bases was localized in the 33-nt stretch from nt 2185 to 2217 in the 5' UTR. The observed cis-active sequences are potential binding sites for trans-factors that could regulate spatio-temporal CPS gene expression in response to biotic and abiotic stress conditions. Obtained results are initially verified by in silico and co-expression studies based on A. thaliana microarray data. The quantitative RT-PCR analysis confirmed that the entire 2269-bp copalyl diphosphate synthase gene fragment has the promoter activity. Quantitative RT-PCR analysis was used to study changes in CPS promoter activity occurring in response to the application of four selected biotic and abiotic regulatory factors; auxin, gibberellin, salicylic acid, and high-salt concentration.

Design and optimization of N-acylhydrazone pyrimidine derivatives as E. coli PDHc E1 inhibitors: Structure-activity relationship analysis, biological evaluation and molecular docking study.

Science.gov (United States)

He, Haifeng; Xia, Hongying; Xia, Qin; Ren, Yanliang; He, Hongwu

2017-10-15

By targeting the thiamin diphosphate (ThDP) binding site of Escherichia coli (E. coli) pyruvate dehydrogenase multienzyme complex E1 (PDHc E1), a series of novel 'open-chain' classes of ThDP analogs A, B, and C with N-acylhydrazone moieties was designed and synthesized to explore their activities against E. coli PHDc E1 in vitro and their inhibitory activity against microbial diseases were further evaluated in vivo. As a result, A1-23 exhibited moderate to potent inhibitory activities against E. coli PDHc E1 (IC 50 =0.15-23.55μM). The potent inhibitors A13, A14, A15, C2, had strong inhibitory activities with IC 50 values of 0.60, 0.15, 0.39 and 0.34μM against E. coli PDHc E1 and with good enzyme-selective inhibition between microorganisms and mammals. Especially, the most powerful inhibitor A14 could 99.37% control Xanthimonas oryzae pv. Oryzae. Furthermore, the binding features of compound A14 within E. coli PDHc E1 were investigated to provide useful insights for the further construction of new inhibitor by molecular docking, site-directed mutagenesis, and enzymatic assays. The results indicated that A14 had most powerful inhibition against E. coli PDHc E1 due to the establishment of stronger interaction with Glu571, Met194, Glu522, Leu264 and Phe602 at active site of E.coli PDHc E1. It could be used as a lead compound for further optimization, and may have potential as a new microbicide. Copyright © 2017 Elsevier Ltd. All rights reserved.

Enhanced production of GDP-L-fucose by overexpression of NADPH regenerator in recombinant Escherichia coli.

Science.gov (United States)

Lee, Won-Heong; Chin, Young-Wook; Han, Nam Soo; Kim, Myoung-Dong; Seo, Jin-Ho

2011-08-01

Biosynthesis of guanosine 5'-diphosphate-L-fucose (GDP-L-fucose) requires NADPH as a reducing cofactor. In this study, endogenous NADPH regenerating enzymes such as glucose-6-phosphate dehydrogenase (G6PDH), isocitrate dehydrogenase (Icd), and NADP(+)-dependent malate dehydrogenase (MaeB) were overexpressed to increase GDP-L-fucose production in recombinant Escherichia coli. The effects of overexpression of each NADPH regenerating enzyme on GDP-L-fucose production were investigated in a series of batch and fed-batch fermentations. Batch fermentations showed that overexpression of G6PDH was the most effective for GDP-L-fucose production. However, GDP-L-fucose production was not enhanced by overexpression of G6PDH in the glucose-limited fed-batch fermentation. Hence, a glucose feeding strategy was optimized to enhance GDP-L-fucose production. Fed-batch fermentation with a pH-stat feeding mode for sufficient supply of glucose significantly enhanced GDP-L-fucose production compared with glucose-limited fed-batch fermentation. A maximum GDP-L-fucose concentration of 235.2 ± 3.3 mg l(-1), corresponding to a 21% enhancement in the GDP-L-fucose production compared with the control strain overexpressing GDP-L-fucose biosynthetic enzymes only, was achieved in the pH-stat fed-batch fermentation of the recombinant E. coli overexpressing G6PDH. It was concluded that sufficient glucose supply and efficient NADPH regeneration are crucial for NADPH-dependent GDP-L-fucose production in recombinant E. coli.

Chronic alcoholism in rats induces a compensatory response, preserving brain thiamine diphosphate, but the brain 2-oxo acid dehydrogenases are inactivated despite unchanged coenzyme levels.

Science.gov (United States)

Parkhomenko, Yulia M; Kudryavtsev, Pavel A; Pylypchuk, Svetlana Yu; Chekhivska, Lilia I; Stepanenko, Svetlana P; Sergiichuk, Andrej A; Bunik, Victoria I

2011-06-01

Thiamine-dependent changes in alcoholic brain were studied using a rat model. Brain thiamine and its mono- and diphosphates were not reduced after 20 weeks of alcohol exposure. However, alcoholism increased both synaptosomal thiamine uptake and thiamine diphosphate synthesis in brain, pointing to mechanisms preserving thiamine diphosphate in the alcoholic brain. In spite of the unchanged level of the coenzyme thiamine diphosphate, activities of the mitochondrial 2-oxoglutarate and pyruvate dehydrogenase complexes decreased in alcoholic brain. The inactivation of pyruvate dehydrogenase complex was caused by its increased phosphorylation. The inactivation of 2-oxoglutarate dehydrogenase complex (OGDHC) correlated with a decrease in free thiols resulting from an elevation of reactive oxygen species. Abstinence from alcohol following exposure to alcohol reactivated OGDHC along with restoration of the free thiol content. However, restoration of enzyme activity occurred before normalization of reactive oxygen species levels. Hence, the redox status of cellular thiols mediates the action of oxidative stress on OGDHC in alcoholic brain. As a result, upon chronic alcohol consumption, physiological mechanisms to counteract the thiamine deficiency and silence pyruvate dehydrogenase are activated in rat brain, whereas OGDHC is inactivated due to impaired antioxidant ability. © 2011 The Authors. Journal of Neurochemistry © 2011 International Society for Neurochemistry.

Overexpression of Farnesyl Diphosphate Synthase in Arabidopsis Mitochondria Triggers Light-dependent Lesion Formation and Alters Cytokinin Homeostasis

Czech Academy of Sciences Publication Activity Database

Manzano, D.; Busquets, A.; Closa, M.; Hoyerová, Klára; Schaller, H.; Kamínek, Miroslav; Arró, M.; Ferrer, A.

2006-01-01

Roč. 61, 1-2 (2006), s. 195-213 ISSN 0167-4412 R&D Projects: GA AV ČR(CZ) IAA600380507 Institutional research plan: CEZ:AV0Z50380511 Keywords : Arabidopsis thaliana * cytokinin * farnesyl diphosphate synthase * isoprenoid Subject RIV: EF - Botanics Impact factor: 3.577, year: 2006

  Adenosine-diphosphate (ADP) reduces infarct size and improves porcine heart function after myocardial infarction

DEFF Research Database (Denmark)

Bune, Laurids Touborg; Larsen, Jens Kjærgaard Rolighed; Thaning, Pia

2013-01-01

Acute myocardial infarction continues to be a major cause of morbidity and mortality. Timely reperfusion can substantially improve outcomes and the administration of cardioprotective substances during reperfusion is therefore highly attractive. Adenosine diphosphate (ADP) and uridine-5-triphoshat...... infusion during reperfusion reduces IS by ~20% independently from systemic release of t-PA. ADP-induced reduction in both preload and afterload could account for the beneficial myocardial effect....

Modification of zirconium diphosphate with salicylic acid and its effect on the uranium (Vi) sorption; Modificacion del difosfato de circonio con acido salicilico y su efecto sobre la sorcion de uranio (VI)

Energy Technology Data Exchange (ETDEWEB)

Almazan T, M. G.; Garcia G, N. [ININ, Carretera Mexico-Toluca s/n, 52750 Ocoyoacac, Estado de Mexico (Mexico); Simoni, E., E-mail: guadalupe.almazan@inin.gob.mx [Universidad Paris Sud, Instituto de Fisica Nuclear, Georges Clemenceau No. 15, Orsay (France)

2014-10-15

The surface of zirconium diphosphate (ZrP{sub 2}O{sub 7}) was modified with salicylic acid and its effect was evaluated on the uranium (Vi) sorption. The modified surface of the material was analyzed with different analytical techniques among which are included the atomic force microscopy, scanning electron microscopy and X-ray photoelectron spectroscopy. This analysis allowed showing that the salicylic acid is being held on the surface of the zirconium diphosphate. The reactivity of modified zirconium diphosphate compared with uranium (Vi) was investigated using the classical method of batch sorption. The analysis of sorption isotherms shows that the salicylic acid has an important effect in the uranium (Vi) sorption. According to the study conducted, the interaction among the uranium (Vi) and the surface of zirconium diphosphate modified with the salicylic acid most likely leads to the complexes formation of binary (U(Vi)/ZrP{sub 2}O{sub 7}) and ternary (U(Vi)/salicylate/ZrP{sub 2}O{sub 7}) surface. (Author)

Synthesis of a tritium labeled photolabile analogue of farnesyl diphosphate: (E,E)-[1-3H]-(2-diazo-3-trifluoropropionyloxy)geranyl diphosphate (DATFP-GDP)

International Nuclear Information System (INIS)

Liu, J.; Benedict, C.R.

1996-01-01

Tritiated (E,E)-(2-diazo-3-trifluoropropionyloxy)geranyl disphosphate (DATFP-GDP) has been used as a photolabile analogue of (E,E)-farnesyl diphosphate (E,E-FDP) for an aid in isolating enzymes utilizing E,E-FDP as a substrate. We now report an alternative method of synthesizing this probe in which the tritium label is introduced in the step just before the introduction of the diphospate group. Thus, DATFP-geranial is oxidized to DATFP-geranial with activated manganese dioxide. The tritium label is introduced by reduction of the aldehyde with NaBT 4 . The DATFP-group successfully withstands both of these steps. The overall yield for these two steps is 69%. Diphosphorylation of the resulting alcohol afforded DATFP-[1- 3 H]-GDP in 8% yield with a specific activity of 48.6 μCi/μmol and radiochemical purity of 94%. (Author)

7-methylguanosine diphosphate (m(7)GDP) is not hydrolyzed but strongly bound by decapping scavenger (DcpS) enzymes and potently inhibits their activity.

Science.gov (United States)

Wypijewska, Anna; Bojarska, Elzbieta; Lukaszewicz, Maciej; Stepinski, Janusz; Jemielity, Jacek; Davis, Richard E; Darzynkiewicz, Edward

2012-10-09

Decapping scavenger (DcpS) enzymes catalyze the cleavage of a residual cap structure following 3' → 5' mRNA decay. Some previous studies suggested that both m(7)GpppG and m(7)GDP were substrates for DcpS hydrolysis. Herein, we show that mononucleoside diphosphates, m(7)GDP (7-methylguanosine diphosphate) and m(3)(2,2,7)GDP (2,2,7-trimethylguanosine diphosphate), resulting from mRNA decapping by the Dcp1/2 complex in the 5' → 3' mRNA decay, are not degraded by recombinant DcpS proteins (human, nematode, and yeast). Furthermore, whereas mononucleoside diphosphates (m(7)GDP and m(3)(2,2,7)GDP) are not hydrolyzed by DcpS, mononucleoside triphosphates (m(7)GTP and m(3)(2,2,7)GTP) are, demonstrating the importance of a triphosphate chain for DcpS hydrolytic activity. m(7)GTP and m(3)(2,2,7)GTP are cleaved at a slower rate than their corresponding dinucleotides (m(7)GpppG and m(3)(2,2,7)GpppG, respectively), indicating an involvement of the second nucleoside for efficient DcpS-mediated digestion. Although DcpS enzymes cannot hydrolyze m(7)GDP, they have a high binding affinity for m(7)GDP and m(7)GDP potently inhibits DcpS hydrolysis of m(7)GpppG, suggesting that m(7)GDP may function as an efficient DcpS inhibitor. Our data have important implications for the regulatory role of m(7)GDP in mRNA metabolic pathways due to its possible interactions with different cap-binding proteins, such as DcpS or eIF4E.

Structural and thermodynamic basis of the inhibition of Leishmania major farnesyl diphosphate synthase by nitrogen-containing bisphosphonates

Energy Technology Data Exchange (ETDEWEB)

Aripirala, Srinivas [Johns Hopkins University, 725 North Wolfe Street WBSB 605, Baltimore, MD 21210 (United States); Gonzalez-Pacanowska, Dolores [López-Neyra Institute of Parasitology and Biomedicine, 18001 Granada (Spain); Oldfield, Eric [University of Illinois at Urbana-Champaign, Urbana, IL 61801 (United States); Kaiser, Marcel [University of Basel, Petersplatz 1, CH-4003 Basel (Switzerland); Amzel, L. Mario, E-mail: mamzel@jhmi.edu [Johns Hopkins University School of Medicine, 725 N. Wolfe Street WBSB 604, Baltimore, MD 21205 (United States); Gabelli, Sandra B., E-mail: mamzel@jhmi.edu [Johns Hopkins University School of Medicine, 725 N. Wolfe Street WBSB 604, Baltimore, MD 21205 (United States); Johns Hopkins University School of Medicine, Baltimore, MD 21205 (United States); Johns Hopkins University, 725 North Wolfe Street WBSB 605, Baltimore, MD 21210 (United States)

2014-03-01

Structural insights into L. major farnesyl diphosphate synthase, a key enzyme in the mevalonate pathway, are described. Farnesyl diphosphate synthase (FPPS) is an essential enzyme involved in the biosynthesis of sterols (cholesterol in humans and ergosterol in yeasts, fungi and trypanosomatid parasites) as well as in protein prenylation. It is inhibited by bisphosphonates, a class of drugs used in humans to treat diverse bone-related diseases. The development of bisphosphonates as antiparasitic compounds targeting ergosterol biosynthesis has become an important route for therapeutic intervention. Here, the X-ray crystallographic structures of complexes of FPPS from Leishmania major (the causative agent of cutaneous leishmaniasis) with three bisphosphonates determined at resolutions of 1.8, 1.9 and 2.3 Å are reported. Two of the inhibitors, 1-(2-hydroxy-2,2-diphosphonoethyl)-3-phenylpyridinium (300B) and 3-butyl-1-(2,2-diphosphonoethyl)pyridinium (476A), co-crystallize with the homoallylic substrate isopentenyl diphosphate (IPP) and three Ca{sup 2+} ions. A third inhibitor, 3-fluoro-1-(2-hydroxy-2,2-diphosphonoethyl)pyridinium (46I), was found to bind two Mg{sup 2+} ions but not IPP. Calorimetric studies showed that binding of the inhibitors is entropically driven. Comparison of the structures of L. major FPPS (LmFPPS) and human FPPS provides new information for the design of bisphosphonates that will be more specific for inhibition of LmFPPS. The asymmetric structure of the LmFPPS–46I homodimer indicates that binding of the allylic substrate to both monomers of the dimer results in an asymmetric dimer with one open and one closed homoallylic site. It is proposed that IPP first binds to the open site, which then closes, opening the site on the other monomer, which closes after binding the second IPP, leading to the symmetric fully occupied FPPS dimer observed in other structures.

Involvement of an ent-copalyl diphosphate synthase in tissue-specific accumulation of specialized diterpenes in Andrographis paniculata.

Science.gov (United States)

Misra, Rajesh Chandra; Garg, Anchal; Roy, Sudeep; Chanotiya, Chandan Singh; Vasudev, Prema G; Ghosh, Sumit

2015-11-01

Ent-labdane-related diterpene (ent-LRD) specialized (i.e. secondary) metabolites of the medicinal plant kalmegh (Andrographis paniculata) have long been known for several pharmacological activities. However, our understanding of the ent-LRD biosynthetic pathway has remained largely incomplete. Since ent-LRDs accumulate in leaves, we carried out a comparative transcriptional analysis using leaf and root tissues, and identified 389 differentially expressed transcripts, including 223 transcripts that were preferentially expressed in leaf tissue. Analysis of the transcripts revealed various specialized metabolic pathways, including transcripts of the ent-LRD biosynthetic pathway. Two class II diterpene synthases (ApCPS1 and ApCPS2) along with one (ApCPS1') and two (ApCPS2' and ApCPS2″) transcriptional variants that were the outcomes of alternative splicing of the precursor mRNA and alternative transcriptional termination, respectively, were identified. ApCPS1 and ApCPS2 encode for 832- and 817-amino acids proteins, respectively, and are phylogenetically related to the dicotyledons ent-copalyl diphosphate synthases (ent-CPSs). The spatio-temporal patterns of ent-LRD metabolites accumulation and gene expression suggested a likely role for ApCPS1 in general (i.e. primary) metabolism, perhaps by providing precursor for the biosynthesis of phytohormone gibberellin (GA). However, ApCPS2 is potentially involved in tissue-specific accumulation of ent-LRD specialized metabolites. Bacterially expressed recombinant ApCPS2 catalyzed the conversion of (E,E,E)-geranylgeranyl diphosphate (GGPP), the general precursor of diterpenes to ent-copalyl diphosphate (ent-CPP), the precursor of ent-LRDs. Taken together, these results advance our understanding of the tissue-specific accumulation of specialized ent-LRDs of medicinal importance. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Thymidylate Synthase, Thymidine Phosphorylase and Orotate Phosphoribosyl Transferase Levels as Predictive Factors of Chemotherapy in Oral Squamous Cell Carcinoma

International Nuclear Information System (INIS)

Ogiuchi, Yosuke; Maruoka, Yasubumi; Ando, Tomohiro; Kobayashi, Makio; Ogiuchi, Hideki

2008-01-01

We conducted a clinicopathologic study on protein and mRNA levels of thymidylate synthase (TS), thymidine phosphorylase (TP) and orotate phosphoribosyl transferase (OPRT) using biopsy tissue specimens before treatment. The mRNA levels have been measured in tumor cells microdissected from paraffin-embedded specimens (Danenberg Tumor Profile method: DTP method). We studied the mRNA and protein expression as effect predictive factors in chemotherapy. The subjects consisted of 20 cases of untreated oral squamous cell carcinoma who had undergone chemotherapy with TS-1 (16 males and 4 females, tongue in 8 cases, upper gingiva in 3 cases, lower gingiva in 3 cases, buccal mucosa in 5 cases and floor of the mouth in 1 case). TS gene expressions of the responders were lower than those for the nonresponders. Furthermore, regarding males who were less than 70 years of age, stage I and II, well differentiated type and tongue, TS mRNA expression of the responders were lower than that for the nonresponders. The mRNA expression of OPRT for the male responders was lower than that for the nonresponders. No remarkable difference was observed by immunohistochemistry. In this study, the measurement of the TS levels using the DTP method may potentially act as a predictive factor of antitumor effectiveness

Hepatic conversion of bilirubin monoglucuronide to diglucuronide in uridine diphosphate-glucuronyl transferase-deficient man and rat by bilirubin glucuronoside glucuronosyltransferase

NARCIS (Netherlands)

Chowdhury, J. R.; Jansen, P. L.; Fischberg, E. B.; Daniller, A.; Arias, I. M.

1978-01-01

The microsomal enzyme uridine diphosphate (UDP) glucuronate glucuronyltransferase (E.C. 2.4.1.17) catalyzes formation of bilirubin mono-glucuronide from bilirubin and UDPglucuronic acid. Bilirubin glucuronoside glucuronosyltransferase (E.C. 2.4.1.95), an enzyme concentrated in plasma

Structure of the ent -Copalyl Diphosphate Synthase PtmT2 from Streptomyces platensis CB00739, a Bacterial Type II Diterpene Synthase

Energy Technology Data Exchange (ETDEWEB)

Rudolf, Jeffrey D.; Dong, Liao-Bin; Cao, Hongnan; Hatzos-Skintges, Catherine; Osipiuk, Jerzy; Endres, Michael; Chang, Chin-Yuan; Ma, Ming; Babnigg, Gyorgy; Joachimiak, Andrzej; Phillips, George N.; Shen, Ben

2016-08-31

Terpenoids are the largest and most structurally diverse family of natural products found in nature, yet their presence in bacteria is underappreciated. The carbon skeletons of terpenoids are generated through carbocation-dependent cyclization cascades catalyzed by terpene synthases (TSs). Type I and type II TSs initiate cyclization via diphosphate ionization and protonation, respectively, and protein structures of both types are known. Most plant diterpene synthases (DTSs) possess three alpha-helical domains (alpha beta gamma), which are thought to have arisen from the fusion of discrete, ancestral bacterial type I TSs (alpha) and type II TSs (beta gamma). Type II DTSs of bacterial origin, of which there are no structurally characterized members, are a missing piece in the structural evolution of TSs. Here, we report the first crystal structure of a type II DTS from bacteria. PtnaT2 from Streptomyces platensis CB00739 was verified as an ent-copalyl diphosphate synthase involved in the biosynthesis of platensimycin and platencin. The crystal structure of PtmT2 was solved at a resolution of 1.80 angstrom, and docking studies suggest the catalytically active conformation of geranylgeranyl diphosphate (GGPP). Site-directed mutagenesis confirmed residues involved in binding the diphosphate moiety of GGPP and identified DxxxxE as a potential Mg2+-binding motif for type II DTSs of bacterial origin. Finally, both the shape and physicochemical properties of the active sites are responsible for determining specific catalytic outcomes of TSs. The structure of PtmT2 fundamentally advances the knowledge of bacterial TSs, their mechanisms, and their role in the evolution of TSs.

Location of the redox-active thiols of ribonucleotide reductase: sequences similarity between the Escherichia coli and Lactobacillus leichmannii enzymes

International Nuclear Information System (INIS)

Lin, A.N.I.; Ashley, G.W.; Stubbe, J.

1987-01-01

The redox-active thiols of Escherichia coli ribonucleoside diphosphate reductase and of Lactobacillus leichmannii ribonucleoside triphosphate reductase have been located by a procedure involving (1) prereduction of enzyme with dithiothreitol, (2) specific oxidation of the redox-active thiols by treatment with substrate in the absence of exogenous reductant, (3) alkylation of other thiols with iodoacetamide, and (4) reduction of the disulfides with dithiothreitol and alkylation with [1- 14 C]iodoacetamide. The dithiothreitol-reduce E. coli B1 subunit is able to convert 3 equiv of CDP to dCDP and is labeled with 5.4 equiv of 14 C. Sequencing of tryptic peptides shows that 2.8 equiv of 14 C is on cysteines-752 and -757 at the C-terminus of B1, while 1.0-1.5 equiv of 14 C is on cysteines-222 and -227. It thus appears that two sets of redox-active dithiols are involved in substrate reduction. The L. leichmannii reductase is able to convert 1.1 equiv of CTP to dCTP and is labeled with 2.1 equiv of 14 C. Sequencing of tryptic peptides shows that 1.4 equiv of 14 C is located on the two cysteines of C-E-G-G-A-C-P-I-K. This peptide shows remarkable and unexpected similarity to the thiol-containing region of the C-terminal peptide of E. coli B1, C-E-S-G-A-C-K-I

Kinetic Mechanism of Uracil Phosphoribosyltransferase from Escherichia coli and Catalytic Importance of the Conserved Proline in the PRPP Binding Site

DEFF Research Database (Denmark)

Lundegaard, Claus; Jensen, Kaj Frank

1999-01-01

Phosphoribosyltransferases catalyze the formation of nucleotides from a nitrogenous base and 5-phosphoribosyl-a-1-pyrophosphate (PRPP). These enzymes and the PRPP synthases resemble each other in a short homologous sequence of 13 amino acid residues which has been termed the PRPP binding site and...

Adenine phosphoribosyltransferase from Sulfolobus solfataricus is an enzyme with unusual kinetic properties and a crystal structure that suggests it evolved from a 6-oxopurine phosphoribosyltransferase.

Science.gov (United States)

Jensen, Kaj Frank; Hansen, Michael Riis; Jensen, Kristine Steen; Christoffersen, Stig; Poulsen, Jens-Christian Navarro; Mølgaard, Anne; Kadziola, Anders

2015-04-14

The adenine phosphoribosyltransferase (APRTase) encoded by the open reading frame SSO2342 of Sulfolobus solfataricus P2 was subjected to crystallographic, kinetic, and ligand binding analyses. The enzyme forms dimers in solution and in the crystals, and binds one molecule of the reactants 5-phosphoribosyl-α-1-pyrophosphate (PRPP) and adenine or the product adenosine monophosphate (AMP) or the inhibitor adenosine diphosphate (ADP) in each active site. The individual subunit adopts an overall structure that resembles a 6-oxopurine phosphoribosyltransferase (PRTase) more than known APRTases implying that APRT functionality in Crenarchaeotae has its evolutionary origin in this family of PRTases. Only the N-terminal two-thirds of the polypeptide chain folds as a traditional type I PRTase with a five-stranded β-sheet surrounded by helices. The C-terminal third adopts an unusual three-helix bundle structure that together with the nucleobase-binding loop undergoes a conformational change upon binding of adenine and phosphate resulting in a slight contraction of the active site. The inhibitor ADP binds like the product AMP with both the α- and β-phosphates occupying the 5'-phosphoribosyl binding site. The enzyme shows activity over a wide pH range, and the kinetic and ligand binding properties depend on both pH and the presence/absence of phosphate in the buffers. A slow hydrolysis of PRPP to ribose 5-phosphate and pyrophosphate, catalyzed by the enzyme, may be facilitated by elements in the C-terminal three-helix bundle part of the protein.

Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae.

Science.gov (United States)

Zhao, Jianzhi; Bao, Xiaoming; Li, Chen; Shen, Yu; Hou, Jin

2016-05-01

Monoterpenes have wide applications in the food, cosmetics, and medicine industries and have recently received increased attention as advanced biofuels. However, compared with sesquiterpenes, monoterpene production is still lagging in Saccharomyces cerevisiae. In this study, geraniol, a valuable acyclic monoterpene alcohol, was synthesized in S. cerevisiae. We evaluated three geraniol synthases in S. cerevisiae, and the geraniol synthase Valeriana officinalis (tVoGES), which lacked a plastid-targeting peptide, yielded the highest geraniol production. To improve geraniol production, synthesis of the precursor geranyl diphosphate (GPP) was regulated by comparing three specific GPP synthase genes derived from different plants and the endogenous farnesyl diphosphate synthase gene variants ERG20 (G) (ERG20 (K197G) ) and ERG20 (WW) (ERG20 (F96W-N127W) ), and controlling endogenous ERG20 expression, coupled with increasing the expression of the mevalonate pathway by co-overexpressing IDI1, tHMG1, and UPC2-1. The results showed that overexpressing ERG20 (WW) and strengthening the mevalonate pathway significantly improved geraniol production, while expressing heterologous GPP synthase genes or down-regulating endogenous ERG20 expression did not show positive effect. In addition, we constructed an Erg20p(F96W-N127W)-tVoGES fusion protein, and geraniol production reached 66.2 mg/L after optimizing the amino acid linker and the order of the proteins. The best strain yielded 293 mg/L geraniol in a fed-batch cultivation, a sevenfold improvement over the highest titer previously reported in an engineered S. cerevisiae strain. Finally, we showed that the toxicity of geraniol limited its production. The platform developed here can be readily used to synthesize other monoterpenes.

Human T cell recognition of the blood stage antigen Plasmodium hypoxanthine guanine xanthine phosphoribosyl transferase (HGXPRT in acute malaria

Directory of Open Access Journals (Sweden)

Woodberry Tonia

2009-06-01

Full Text Available Abstract Background The Plasmodium purine salvage enzyme, hypoxanthine guanine xanthine phosphoribosyl transferase (HGXPRT can protect mice against Plasmodium yoelii pRBC challenge in a T cell-dependent manner and has, therefore, been proposed as a novel vaccine candidate. It is not known whether natural exposure to Plasmodium falciparum stimulates HGXPRT T cell reactivity in humans. Methods PBMC and plasma collected from malaria-exposed Indonesians during infection and 7–28 days after anti-malarial therapy, were assessed for HGXPRT recognition using CFSE proliferation, IFNγ ELISPOT assay and ELISA. Results HGXPRT-specific T cell proliferation was found in 44% of patients during acute infection; in 80% of responders both CD4+ and CD8+ T cell subsets proliferated. Antigen-specific T cell proliferation was largely lost within 28 days of parasite clearance. HGXPRT-specific IFN-γ production was more frequent 28 days after treatment than during acute infection. HGXPRT-specific plasma IgG was undetectable even in individuals exposed to malaria for at least two years. Conclusion The prevalence of acute proliferative and convalescent IFNγ responses to HGXPRT demonstrates cellular immunogenicity in humans. Further studies to determine minimal HGXPRT epitopes, the specificity of responses for Plasmodia and associations with protection are required. Frequent and robust T cell proliferation, high sequence conservation among Plasmodium species and absent IgG responses distinguish HGXPRT from other malaria antigens.

ANALISIS CEMARAN BAKTERI Escherichia coli ANALISIS CEMARAN BAKTERI Escherichia coli ANALISIS CEMARAN BAKTERI Escherichia coli

OpenAIRE

ANGGREINI, RAHAYU

2015-01-01

2015 RAHAYU ANGGREINI coli Penelitian ini bertujuan untuk melakukan identifikasi cemaran bakteri E. coli O157:H7 pada daging sapi di kota Makassar. Sampel pada penelitian ini sebanyak 72 sampel Kata Kunci : Daging sapi, pasar tradisional, E. coli, E. coli O157:H7, kontaminasi bakteri, identifikasi E. coli O157:H7.

Tenofovir-induced Fanconi syndrome and osteomalacia in two HIV-infected patients: Role of intracellular tenofovir diphosphate levels and review of the literature

NARCIS (Netherlands)

Haverkort, M.E.; van der Spek, B.W.; Lips, P.T.A.M.; Slieker, W.A.; ter Heine, R.; Huitema, A.D.; Bronsveld, W.

2011-01-01

We present 2 human immunodeficiency virus-infected patients with tenofovir disoproxil fumarate-induced Fanconi syndrome, leading to osteomalacia. Intracellular tenofovir diphosphate levels were measured in 1 patient and were found to be very high, with plasma tenofovir levels just slightly elevated.

Study of the thorium phosphate-diphosphate (TPD) dissolution: kinetic aspect - thermodynamic aspect: analysis of the neo-formed phases; Etude de la dissolution du phosphate diphosphate de thorium: - aspect cinetique - aspect thermodynamique: analyse des phases neoformees

Energy Technology Data Exchange (ETDEWEB)

Thomas, A.Ch

2000-10-06

The aim of this work is to study the aqueous corrosion of the thorium phosphate-diphosphate (TPD), of the formula Th{sub 4}(PO{sub 4}){sub 4}P{sub 2}O{sub 7}, in the framework of the actinides immobilization. In order to complete the anterior studies concerning solid solutions where thorium is substituted by a tetravalent ion (uranium (IV) or plutonium (IV)) in the TPD structure, compounds of thorium and neptunium phosphate-diphosphate, of formula Th{sub 4-x}Np{sub x}(PO{sub 4}){sub 4}P{sub 2}O{sub 7}, have been prepared. Furthermore, a new chemical way of synthesis has been investigated in order to sinter solids solution of thorium and uranium phosphate-diphosphate (TUPD) in good conditions. The TPD dissolution study showed two principals steps. The first one corresponds to the control of element concentration by the material dissolution whereas the second corresponds to the formation of secondary precipitates for which thermodynamic equilibrium controls the concentration of the species in solution. Leaching tests have been performed varying several independent parameters in order to determine the TPD dissolution rate. The partial orders related to the protons or to the hydroxide ions have been found between 0.35 and 0.45 whereas the apparent dissolution rate constants are in the range 1.10{sup -5} for 9.10{sup -5} g.m{sup -2}.j{sup -1} for acidic and basic media. The neo-formed phases have been characterized after the dissolution of TPD and TUPD. We found that the TPD leaching in acidic medium leads to the formation of the crystallized thorium phosphate-hydrogen-phosphate (TPHP), of formula Th{sub 2}(PO{sub 4}){sub 2}(HPO{sub 4}), x H{sub 2}O, whereas the TUPD dissolution leads to the TPHP and an other compound, of formula (UO{sub 2}){sub 3}(PO{sub 4}){sub 2}, 5 H{sub 2}O. We calculated its solubility product which is in good agreement with those found in the literature. The phases formed during the leaching of solids containing plutonium; americium or curium (Th

A recruiting protein of geranylgeranyl diphosphate synthase controls metabolic flux toward chlorophyll biosynthesis in rice.

Science.gov (United States)

Zhou, Fei; Wang, Cheng-Yuan; Gutensohn, Michael; Jiang, Ling; Zhang, Peng; Zhang, Dabing; Dudareva, Natalia; Lu, Shan

2017-06-27

In plants, geranylgeranyl diphosphate (GGPP) is produced by plastidic GGPP synthase (GGPPS) and serves as a precursor for vital metabolic branches, including chlorophyll, carotenoid, and gibberellin biosynthesis. However, molecular mechanisms regulating GGPP allocation among these biosynthetic pathways localized in the same subcellular compartment are largely unknown. We found that rice contains only one functionally active GGPPS, OsGGPPS1, in chloroplasts. A functionally active homodimeric enzyme composed of two OsGGPPS1 subunits is located in the stroma. In thylakoid membranes, however, the GGPPS activity resides in a heterodimeric enzyme composed of one OsGGPPS1 subunit and GGPPS recruiting protein (OsGRP). OsGRP is structurally most similar to members of the geranyl diphosphate synthase small subunit type II subfamily. In contrast to members of this subfamily, OsGRP enhances OsGGPPS1 catalytic efficiency and specificity of GGPP production on interaction with OsGGPPS1. Structural biology and protein interaction analyses demonstrate that affinity between OsGRP and OsGGPPS1 is stronger than between two OsGGPPS1 molecules in homodimers. OsGRP determines OsGGPPS1 suborganellar localization and directs it to a large protein complex in thylakoid membranes, consisting of geranylgeranyl reductase (OsGGR), light-harvesting-like protein 3 (OsLIL3), protochlorophyllide oxidoreductase (OsPORB), and chlorophyll synthase (OsCHLG). Taken together, genetic and biochemical analyses suggest OsGRP functions in recruiting OsGGPPS1 from the stroma toward thylakoid membranes, thus providing a mechanism to control GGPP flux toward chlorophyll biosynthesis.

Analysis of quinocide in unprocessed primaquine diphosphate and primaquine diphosphate tablets using gas chromatography-mass spectrometry with supersonic molecular beams.

Science.gov (United States)

Brondz, Ilia; Fialkov, Alexander B; Amirav, Aviv

2009-01-30

Malaria is one of the most widespread and deadly diseases on the planet. Every year, about 500 million new cases are diagnosed, and the annual death toll is about 3 million. Primaquine has strong antiparasitic effects against gametocytes and can therefore prevent the spread of the parasite from treated patients to mosquitoes. It is also used in radical cures and prevents relapse. Consequently, primaquine is an often-used drug. In this study the separation of unprocessed primaquine from the contaminant quinocide based on gas chromatography-mass spectrometry with supersonic molecular beam (SMB) is presented and 7.5 mg primaquine diphosphate tablets were analyzed. We present a novel method for fast determination of quinocide which is an isomer of primaquine as the main contaminant in unprocessed primaquine and in its medical form as tablets by gas chromatography-mass spectrometry with SMB (also named supersonic GC-MS). Supersonic GC-MS provides enhanced molecular ion without any ion source related peak tailing plus extended range of compounds amenable for GC-MS analysis. In addition, major isomer mass spectral effects were revealed in the mass spectra of primaquine and quinocide which facilitated the unambiguous identification of quinocide in primaquine tablets. Fast GC-MS analysis is demonstrated with less then 2 min elution time of the drug and its main contaminants.

Acquisition of wild-type HIV-1 infection in a patient on pre-exposure prophylaxis with high intracellular concentrations of tenofovir diphosphate: a case report.

NARCIS (Netherlands)

Hoornenborg, Elske; Prins, Maria; Achterbergh, Roel C A; Woittiez, Lycke R; Cornelissen, Marion; Jurriaans, Suzanne; Kootstra, Neeltje A; Anderson, Peter L; Reiss, Peter; de Vries, Henry J C; Prins, Jan M; de Bree, Godelieve J

2017-01-01

Pre-exposure prophylaxis (PrEP) with emtricitabine and tenofovir disoproxil fumarate is highly effective against acquisition of HIV infection, and only two cases of infection with a multidrug-resistant virus have been reported under adequate long-term adherence, as evidenced by tenofovir diphosphate

Bornyl-diphosphate synthase from Lavandula angustifolia: A major monoterpene synthase involved in essential oil quality.

Science.gov (United States)

Despinasse, Yolande; Fiorucci, Sébastien; Antonczak, Serge; Moja, Sandrine; Bony, Aurélie; Nicolè, Florence; Baudino, Sylvie; Magnard, Jean-Louis; Jullien, Frédéric

2017-05-01

Lavender essential oils (EOs) of higher quality are produced by a few Lavandula angustifolia cultivars and mainly used in the perfume industry. Undesirable compounds such as camphor and borneol are also synthesized by lavender leading to a depreciated EO. Here, we report the cloning of bornyl diphosphate synthase of lavender (LaBPPS), an enzyme that catalyzes the production of bornyl diphosphate (BPP) and then by-products such as borneol or camphor, from an EST library. Compared to the BPPS of Salvia officinalis, the functional characterization of LaBPPS showed several differences in amino acid sequence, and the distribution of catalyzed products. Molecular modeling of the enzyme's active site suggests that the carbocation intermediates are more stable in LaBPPS than in SoBPPS leading probably to a lower efficiency of LaBPPS to convert GPP into BPP. Quantitative RT-PCR performed from leaves and flowers at different development stages of L. angustifolia samples show a clear correlation between transcript level of LaBPPS and accumulation of borneol/camphor, suggesting that LaBPPS is mainly responsible of in vivo biosynthesis of borneol/camphor in fine lavender. A phylogenetic analysis of terpene synthases (TPS) pointed out the basal position of LaBPPS in the TPSb clade, suggesting that LaBPPS could be an ancestor of others lavender TPSb. Finally, borneol could be one of the first monoterpenes to be synthesized in the Lavandula subgenus. Knowledge gained from these experiments will facilitate future studies to improve the lavender oils through metabolic engineering or plant breeding. Accession numbers: LaBPPS: KM015221. Copyright © 2017. Published by Elsevier Ltd.

Effect of fructose diphosphate combined with large-dose vitamin C therapy on the myocardial oxidative stress injury after neonatal asphyxia

Directory of Open Access Journals (Sweden)

Chun-Hua Liang1

2017-04-01

Full Text Available Objective: To study the effect of fructose diphosphate combined with large-dose vitamin C therapy on the myocardial oxidative stress injury after neonatal asphyxia. Methods: 40 patients with neonatal asphyxia who were treated in our hospital between June 2013 and April 2016 were collected and divided into the control group (n=20 who received large-dose vitamin C therapy and the observation group (n=20 who received fructose diphosphate combined with large-dose vitamin C therapy according to the double-blind randomized control method, and the treatment lasted for 10 d. Immediately after admission and after 10 d of treatment, RIA method was used to detect the serum levels of oxidative stress indexes, color Doppler diasonograph was used to determine left cardiac function parameters, and the myocardial enzyme spectrum detector was used to determine myocardial enzyme spectrum index levels. Results: Immediately after admission, the differences in the systemic oxidative stress degree, the left cardiac function damage degree and the myocardial enzyme spectrum index levels were not statistically significant between two groups of patients (P>0.05. After 10 d of treatment, serum malondialdehyde (MDA, advanced oxidation protein products (AOPP, creatine kinase isoenzyme (CK-MB, N-terminal pro-brain natriuretic peptide (Nt-proBNP, heart-type fatty acid-binding protein (H-FABP and troponin I (cTnI contents of observation group were lower than those of control group (P<0.05 while superoxide dismutase (SOD content was higher than that of control group (P<0.05, and the left cardiac function parameter ejection time (ET level was higher than that of control group (P<0.05 while left ventricular isovolumetric contraction time (ICT and left ventricular isovolumetric relaxation time (IRT levels were lower than those of control group (P<0.05. Conclusion: Fructose diphosphate combined with large-dose vitamin C can reduce the systemic oxidative stress of neonatal asphyxia

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...

Enhancing GDP-fucose production in recombinant Escherichia coli by metabolic pathway engineering.

Science.gov (United States)

Zhai, Yafei; Han, Donglei; Pan, Ying; Wang, Shuaishuai; Fang, Junqiang; Wang, Peng; Liu, Xian-wei

2015-02-01

Guanosine 5'-diphosphate (GDP)-fucose is the indispensible donor substrate for fucosyltransferase-catalyzed synthesis of fucose-containing biomolecules, which have been found involving in various biological functions. In this work, the salvage pathway for GDP-fucose biosynthesis from Bacterioides fragilis was introduced into Escherichia coli. Besides, the biosynthesis of guanosine 5'-triphosphate (GTP), an essential substrate for GDP-fucose biosynthesis, was enhanced via overexpression of enzymes involved in the salvage pathway of GTP biosynthesis. The production capacities of metabolically engineered strains bearing different combinations of recombinant enzymes were compared. The shake flask fermentation of the strain expressing Fkp, Gpt, Gmk and Ndk obtained the maximum GDP-fucose content of 4.6 ± 0.22 μmol/g (dry cell mass), which is 4.2 fold that of the strain only expressing Fkp. Through fed-batch fermentation, the GDP-fucose content further rose to 6.6 ± 0.14 μmol/g (dry cell mass). In addition to a better productivity than previous fermentation processes based on the de novo pathway for GDP-fucose biosynthesis, the established schemes in this work also have the advantage to be a potential avenue to GDP-fucose analogs encompassing chemical modification on the fucose residue. Copyright © 2014 Elsevier Inc. All rights reserved.

Electron density reactivity indexes of the tautomeric/ionization forms of thiamin diphosphate.

Science.gov (United States)

Jaña, Gonzalo A; Delgado, Eduardo J

2013-09-01

The generation of the highly reactive ylide in thiamin diphosphate catalysis is analyzed in terms of the nucleophilicity of key atoms, by means of density functional calculations at X3LYP/6-31++G(d,p) level of theory. The Fukui functions of all tautomeric/ionization forms are calculated in order to assess their reactivity. The results allow to conclude that the highly conserved glutamic residue does not protonate the N1' atom of the pyrimidyl ring, but it participates in a strong hydrogen bonding, stabilizing the eventual negative charge on the nitrogen, in all forms involved in the ylide generation. This condition provides the necessary reactivity on key atoms, N4' and C2, to carry out the formation of the ylide required to initiate the catalytic cycle of ThDP-dependent enzymes. This study represents a new approach for the ylide formation in ThDP catalysis.

Purification, crystallization and preliminary X-ray crystallographic analysis of human ppGpp hydrolase, Mesh1

Directory of Open Access Journals (Sweden)

Dawei Sun

2010-09-01

Full Text Available Bacterial SpoT is a Mn2+-dependent pyrophosphohydrolase to hydrolyze guanosine 3’-diphosphate-5’-diphosphate to guanosine diphosphate and pyrophosphate. In this study, SpoT ortholog from Homo sapiens (hMesh1, was over-expressed in Escherichia coli, purified and crystallized using hanging-drop vapour-diffusion method with polyethylene glycol and sodium citrate. The native crystal of hMesh1 was diffracted to 2.1 Å using a synchrotron-radiation source and belonged to the monoclinic space group P21 with cell dimensions of a = 53.27 Å, b = 62.61 Å, c = 52.45 Å and β= 94.96˚. The crystal contains two molecules in the asymmetric unit, with a solvent content of 44% and a Matthews coefficient VM value of 2.18 Å3/Da.

Kinetic study of the thorium phosphate - diphosphate dissolution

International Nuclear Information System (INIS)

Dacheux, N.; Thomas, A.C.; Brandel, V.; Genet, M.

2000-01-01

The thorium phosphate-diphosphate Th 4 (PO 4 ) 4 P 2 O 7 (TPD) structure allows the replacement of large amounts of thorium by tetravalent actinides leading to the formation of solid solutions. This compound was obtained in powdered or sintered form after pressing at room temperature at 300-800 MPa then heating at 1250 deg. C for 10-30 hours. The resistance of this material to aqueous corrosion was determined by varying several parameters such as surface, leaching flow, acidity or temperature. It was thus possible to independently determine the influence of each parameter on the leaching rate provided that the saturation of the solution was not obtained. In acidic media, the partial order related to [H 3 O + ] was found to be in the 0.31-0.35 range while, in basic media, the partial order related to [OH - ] was almost the same (0.45). The activation energy (42 kJ/mol) was determined between 4 deg. C and 120 deg. C. Moreover, the addition of phosphate in the leachate slightly increased the TPD dissolution rate. When the saturation of the solution is reached, a gelatinous precipitate controls the thorium and phosphate concentrations. The complete characterization of this solid led to the proposed general formula Th 2 (PO 4 ) 2 (HPO 4 ). n H 2 O which conventional solubility product (at I = 0 M) is very low: K * S,0 10 -66.6±1.2 even in very acidic media. (authors)

SIMULTANEOUS ANALYSIS OF AZIDOTHYMIDINE AND ITS MONOPHOSPHATE, DIPHOSPHATE AND TRIPHOSPHATE DERIVATIVES IN BIOLOGICAL-FLUIDS, TISSUE AND CULTURED-CELLS BY A RAPID HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC METHOD

NARCIS (Netherlands)

MOLEMA, G; JANSEN, RW; Visser, Jan; MEIJER, DKF

1992-01-01

A rapid high-performance liquid chromatographic (HPLC) method for the simultaneous analysis of the antiviral drug azidothymidine (AZT), AZT monophosphate, AZT diphosphate and AZT triphosphate, with ultraviolet detection in the nanomolar range, is described. Determination of these compounds in vitro

Acid dissociation constants of uridine-5 Prime -diphosphate compounds determined by {sup 31}phosphorus nuclear magnetic resonance spectroscopy and internal pH referencing

Energy Technology Data Exchange (ETDEWEB)

Jancan, Igor [Louisiana State University, Department of Chemistry, Baton Rouge, LA 70803 (United States); Macnaughtan, Megan A., E-mail: macnau@lsu.edu [Louisiana State University, Department of Chemistry, Baton Rouge, LA 70803 (United States)

2012-10-24

Highlights: Black-Right-Pointing-Pointer The first reported phosphate and imide pK{sub a} values of UDP-GlcNAc and UDP-S-GlcNAc. Black-Right-Pointing-Pointer New role for the monosaccharide in the imide pK{sub a} of uridine-5 Prime -phosphate compounds. Black-Right-Pointing-Pointer UDP-S-GlcNAc and UDP-GlcNAc have the same phosphate pK{sub a}, unlike thioyl analogs. Black-Right-Pointing-Pointer The {sup 31}P chemical shift of inorganic phosphate is a viable internal pH reference. Black-Right-Pointing-Pointer Stability of the external {sup 31}P chemical shift reference is essential. - Abstract: The acid dissociation constant (pK{sub a}) of small, biological molecules is an important physical property used for investigating enzyme mechanisms and inhibitor design. For phosphorus-containing molecules, the {sup 31}P nuclear magnetic resonance (NMR) chemical shift is sensitive to the local chemical environment, particularly to changes in the electronic state of the molecule. Taking advantage of this property, we present a {sup 31}P NMR approach that uses inorganic phosphate buffer as an internal pH reference to determine the pK{sub a} values of the imide and second diphosphate of uridine-5 Prime -diphosphate compounds, including the first reported values for UDP-GlcNAc and UDP-S-GlcNAc. New methods for using inorganic phosphate buffer as an internal pH reference, involving mathematical correction factors and careful control of the chemical shift reference sample, are illustrated. A comparison of the newly determined imide and diphosphate pK{sub a} values of UDP, UDP-GlcNAc, and UDP-S-GlcNAc with other nucleotide phosphate and thio-analogs reveals the significance of the monosaccharide and sulfur position on the pK{sub a} values.

Chemical interaction of potassium diphosphate with cadmium nitrate in aqueous solution

International Nuclear Information System (INIS)

Kokhanovskij, V.V.

1993-01-01

Formation of low-soluble compounds in 1.5 mol/l isomolar cross section of K 4 P 2 O 7 -Cd(NO 3 ) 2 -H 2 O system was studied. Liquid phases are studied by the methods of refractometry and pH value measuring, an solid ones - by the methods of chemical and X-ray phase analysis, IR spectroscopy, chromatography and microscopy. Three individual chemical compounds K 2 CdP 2 O 7 x 4H 2 O, K 2 Cd 3 (P 2 O 7 ) 2 x 3H 2 O and Cd 2 P 2 O 7 x 3.5H 2 O and some their mixtures were isolated and investigated. It is shown that doulble diphosphate K 6 Cd(P 2 O 7 ) 2 x 6H 2 O does not precipitate spontanously, but instead of it in wide region of system K 2 CdP 2 O 7 x 4H 2 O crystallizes as elongated acicular crystals or as thin plates of improper form

E coli enteritis

Science.gov (United States)

... coli; Food poisoning - E. coli; E. coli diarrhea; Hamburger disease ... coleslaw or potato salad) that have been out of the refrigerator too ... reheated Fish or oysters Raw fruits or vegetables that have ...

Multiple roles of mobile active center loops in the E1 component of the Escherichia coli pyruvate dehydrogenase complex - Linkage of protein dynamics to catalysis

Science.gov (United States)

Jordan, Frank; Arjunan, Palaniappa; Kale, Sachin; Nemeria, Natalia S.; Furey, William

2009-01-01

The region encompassing residues 401–413 on the E1 component of the pyruvate dehydrogenase multienzyme complex from Escherichia coli comprises a loop (the inner loop) which was not seen in the X-ray structure in the presence of thiamin diphosphate, the required cofactor for the enzyme. This loop is seen in the presence of a stable analogue of the pre-decarboxylation intermediate, the covalent adduct between the substrate analogue methyl acetylphosphonate and thiamin diphosphate, C2α-phosphonolactylthiamin diphosphate. It has been shown that the residue H407 and several other residues on this loop are required to reduce the mobility of the loop so electron density corresponding to it can be seen once the pre-decarboxylation intermediate is formed. Concomitantly, the loop encompassing residues 541–557 (the outer loop) appears to work in tandem with the inner loop and there is a hydrogen bond between the two loops ensuring their correlated motion. The inner loop was shown to: a) sequester the active center from carboligase side reactions; b) assist the interaction between the E1 and the E2 components, thereby affecting the overall reaction rate of the entire multienzyme complex; c) control substrate access to the active center. Using viscosity effects on kinetics it was shown that formation of the pre-decarboxylation intermediate is specifically affected by loop movement. A cysteine-less variant was created for the E1 component, onto which cysteines were substituted at selected loop positions. Introducing an electron spin resonance spin label and an 19F NMR label onto these engineered cysteines, the loop mobility was examined: a) both methods suggested that in the absence of ligand, the loop exists in two conformations; b) line-shape analysis of the NMR signal at different temperatures, enabled estimation of the rate constant for loop movement, and this rate constant was found to be of the same order of magnitude as the turnover number for the enzyme under the

Kinetic study of the thorium phosphate - diphosphate dissolution

Energy Technology Data Exchange (ETDEWEB)

Dacheux, N.; Thomas, A.C.; Brandel, V.; Genet, M. [Paris-11 Univ., 91 - Orsay (France). Inst. de Physique Nucleaire; Aupiais, J. [CEA/DAM-Ile de France, Dept. Analyse Surveillance Environnement, DASE, Service Radioanalyses Chimie Environnement, 91 - Bruyeres-Le-Chatel (France)

2000-07-01

The thorium phosphate-diphosphate Th{sub 4}(PO{sub 4}){sub 4}P{sub 2}O{sub 7} (TPD) structure allows the replacement of large amounts of thorium by tetravalent actinides leading to the formation of solid solutions. This compound was obtained in powdered or sintered form after pressing at room temperature at 300-800 MPa then heating at 1250 deg. C for 10-30 hours. The resistance of this material to aqueous corrosion was determined by varying several parameters such as surface, leaching flow, acidity or temperature. It was thus possible to independently determine the influence of each parameter on the leaching rate provided that the saturation of the solution was not obtained. In acidic media, the partial order related to [H{sub 3}O{sup +}] was found to be in the 0.31-0.35 range while, in basic media, the partial order related to [OH{sup -}] was almost the same (0.45). The activation energy (42 kJ/mol) was determined between 4 deg. C and 120 deg. C. Moreover, the addition of phosphate in the leachate slightly increased the TPD dissolution rate. When the saturation of the solution is reached, a gelatinous precipitate controls the thorium and phosphate concentrations. The complete characterization of this solid led to the proposed general formula Th{sub 2}(PO{sub 4}){sub 2}(HPO{sub 4}). n H{sub 2}O which conventional solubility product (at I = 0 M) is very low: K{sup *}{sub S,0} 10{sup -66.6{+-}}{sup 1.2} even in very acidic media. (authors)

E. Coli Infections

Science.gov (United States)

E. coli is the name of a type of bacteria that lives in your intestines. Most types of E. coli are harmless. However, some types can make you ... type causes travelers' diarrhea. The worst type of E. coli causes bloody diarrhea, and can sometimes cause kidney ...

E. Coli

Science.gov (United States)

... for the bacteria's medical name Escherichia coli . The strange thing about these bacteria — and lots of other ... In some cases, E. coli poisoning can cause life-threatening kidney problems. What Can Kids Do? Adults ...

Intestinal absorption of cytidine diphosphate choline and its changes in the digestive tract

International Nuclear Information System (INIS)

Yashima, Keisuke; Takamatsu, Masatoshi; Okuda, Kunio

1975-01-01

Intestinal absorption of cytidine diphosphate choline (CDP-choline), its structural changes in the digestive tract, and hepatic uptake have been investigated in rats using 14 C-labeled ( 14 CH 3 attached to N of choline) and 3 H-labeled (at C 5 of pyrimidine) compounds. The results indicate that: 1) CDP-choline is relatively stable in the stomach, but is quickly degraded into cytidine and choline in the intestine; 2) The hepatic uptakes of 14 C and 3 H reach the maximum in two to three hours after oral administration; 3) Whereas the amount of 14 C remaining in the gut is inversely related to the hepatic uptake, no similar correlation is seen with 3 H-labeled CDP-choline, and 4) Extrahepatic uptake of 14 C and 3 H is very small. The possibility of phosphorylation in the mucosa of choline and cytidine has been discussed, based on the differences in relative amount of radioactivity in individual broken-down products in the intestinal lumen and mucosa. (auth.)

Chromosomal features of Escherichia coli serotype O2:K2, an avian pathogenic E. coli

DEFF Research Database (Denmark)

Jørgensen, Steffen L; Kudirkiene, Egle; Li, Lili

2017-01-01

Escherichia coli causing infection outside the gastrointestinal system are referred to as extra-intestinal pathogenic E. coli. Avian pathogenic E. coli is a subgroup of extra-intestinal pathogenic E. coli and infections due to avian pathogenic E. coli have major impact on poultry production econo...

Pertussis toxin substrate is a guanosine 5'-[beta-thio]diphosphate-, N-ethylmaleimide-, Mg2+- and temperature-sensitive GTP-binding protein.

OpenAIRE

Wong, S K; Martin, B R; Tolkovsky, A M

1985-01-01

We compared the effects of guanine nucleotides and Mg2+ on ADP-ribosylation of rat brain and liver membrane proteins catalysed by Bordetella pertussis toxin (IAP) and cholera toxin (CT). Labelling of proteins in the presence of [alpha-32P]NAD+, ATP and CT required GTP or guanosine 5'-[gamma-thio]triphosphate (GTP [S]). In contrast, labelling of one (liver) or two (brain) polypeptides by IAP was enhanced by guanosine 5'-[beta-thio]diphosphate (GDP[S]) or GTP, but was blocked by GTP[S] or guano...

Theoretical pKa prediction of the α-phosphate moiety of uridine 5‧-diphosphate-GlcNAc

Science.gov (United States)

Vipperla, Bhavaniprasad; Griffiths, Thomas M.; Wang, Xingyong; Yu, Haibo

2017-01-01

The pKa value of the α-phosphate moiety of uridine 5‧-diphosphate-GlcNAc (UDP-GlcNAc) has been successfully calculated using density functional theory methods in conjunction with the Polarizable Continuum Models. Theoretical methods were benchmarked over a dataset comprising of alkyl phosphates. B3LYP/6-31+G(d,p) calculations using SMD solvation model provide excellent agreement with the experimental data. The predicted pKa for UDP-GlcNAc is consistent with most recent NMR studies but much higher than what it has long been thought to be. The importance of this study is evident that the predicted pKa for UDP-GlcNAc supports its potential role as a catalytic base in the substrate-assisted biocatalysis.

E. Coli and Pregnancy

Science.gov (United States)

... chat Live Help Fact Sheets Share Escherichia coli (E. coli) Friday, 01 September 2017 In every pregnancy, a ... risk. This sheet talks about whether exposure to E. coli may increase the risk for birth defects over ...

Novel concept of enzyme selective nicotinamide adenine dinucleotide (NAD)-modified inhibitors based on enzyme taxonomy from the diphosphate conformation of NAD.

Science.gov (United States)

Fujii, Mikio; Kitagawa, Yasuyuki; Iida, Shui; Kato, Keisuke; Ono, Machiko

2015-11-15

The dihedral angle θ of the diphosphate part of NAD(P) were investigated to distinguish the differences in the binding-conformation of NAD(P) to enzymes and to create an enzyme taxonomy. Furthermore, new inhibitors with fixed dihedral angles showed that enzymes could recognize the differences in the dihedral angle θ. We suggest the taxonomy and the dihedral angle θ are important values for chemists to consider when designing inhibitors and drugs that target enzymes. Copyright © 2015 Elsevier Ltd. All rights reserved.

Chromosomal features of Escherichia coli serotype O2:K2, an avian pathogenic E. coli.

Science.gov (United States)

Jørgensen, Steffen L; Kudirkiene, Egle; Li, Lili; Christensen, Jens P; Olsen, John E; Nolan, Lisa; Olsen, Rikke H

2017-01-01

Escherichia coli causing infection outside the gastrointestinal system are referred to as extra-intestinal pathogenic E. coli. Avian pathogenic E. coli is a subgroup of extra-intestinal pathogenic E. coli and infections due to avian pathogenic E. coli have major impact on poultry production economy and welfare worldwide. An almost defining characteristic of avian pathogenic E. coli is the carriage of plasmids, which may encode virulence factors and antibiotic resistance determinates. For the same reason, plasmids of avian pathogenic E. coli have been intensively studied. However, genes encoded by the chromosome may also be important for disease manifestation and antimicrobial resistance. For the E. coli strain APEC_O2 the plasmids have been sequenced and analyzed in several studies, and E. coli APEC_O2 may therefore serve as a reference strain in future studies. Here we describe the chromosomal features of E. coli APEC_O2. E. coli APEC_O2 is a sequence type ST135, has a chromosome of 4,908,820 bp (plasmid removed), comprising 4672 protein-coding genes, 110 RNA genes, and 156 pseudogenes, with an average G + C content of 50.69%. We identified 82 insertion sequences as well as 4672 protein coding sequences, 12 predicated genomic islands, three prophage-related sequences, and two clustered regularly interspaced short palindromic repeats regions on the chromosome, suggesting the possible occurrence of horizontal gene transfer in this strain. The wildtype strain of E. coli APEC_O2 is resistant towards multiple antimicrobials, however, no (complete) antibiotic resistance genes were present on the chromosome, but a number of genes associated with extra-intestinal disease were identified. Together, the information provided here on E. coli APEC_O2 will assist in future studies of avian pathogenic E. coli strains, in particular regarding strain of E. coli APEC_O2, and aid in the general understanding of the pathogenesis of avian pathogenic E. coli .

Human Meningitis-Associated Escherichia coli

Science.gov (United States)

KIM, KWANG SIK

2016-01-01

E. coli is the most common Gram-negative bacillary organism causing meningitis and E. coli meningitis continues to be an important cause of mortality and morbidity throughout the world. Our incomplete knowledge of its pathogenesis contributes to such mortality and morbidity. Recent reports of E. coli strains producing CTX-M-type or TEM-type extended-spectrum β-lactamases create a challenge. Studies using in vitro and in vivo models of the blood-brain barrier have shown that E. coli meningitis follows a high-degree of bacteremia and invasion of the blood-brain barrier. E. coli invasion of the blood-brain barrier, the essentials step in the development of E. coli meningitis, requires specific microbial and host factors as well as microbe- and host-specific signaling molecules. Blockade of such microbial and host factors contributing to E. coli invasion of the blood-brain barrier is shown to be efficient in preventing E. coli penetration into the brain. The basis for requiring a high-degree of bacteremia for E. coli penetration of the blood-brain barrier, however, remains unclear. Continued investigation on the microbial and host factors contributing to a high-degree of bacteremia and E. coli invasion of the blood-brain barrier is likely to identify new targets for prevention and therapy of E. coli meningitis. PMID:27223820

Diarrheagenic Escherichia coli Markers and Phenotypes among Fecal E. coli Isolates Collected from Nicaraguan Infants ▿

OpenAIRE

Reyes, Daniel; Vilchez, Samuel; Paniagua, Margarita; Colque-Navarro, Patricia; Weintraub, Andrej; Möllby, Roland; Kühn, Inger

2010-01-01

We analyzed the prevalence of diarrheagenic Escherichia coli (DEC) markers and common phenotypes in 2,164 E. coli isolates from 282 DEC-positive samples. Enteropathogenic E. coli (EPEC) and enteroaggregative E. coli (EAEC) were very diverse and were not correlated with diarrhea. Enterotoxigenic E. coli (ETEC) estA and enterohemorrhagic E. coli (EHEC) belonged to a few phenotypes and were significantly correlated with diarrhea.

Formation of nicotinamide ribose diphosphate ribose, a new metabolite of the NAD pathway, by growing mycelium of Aspergillus niger

International Nuclear Information System (INIS)

Kuwahara, Masaaki

1976-01-01

A new step of NAD metabolism was shown in Aspergillus niger. Radioactive nicotinic acid and nicotinamide were incorporated into nicotinamide ribose diphosphate ribose (NAm-RDPR), which had been isolated from the culture filtrate. Its content in the culture medium increased with an increase of culture time, and this compound was proved to be a terminal metabolite in the NAD pathway. The experimental results also showed that the Preiss-Handler pathway and the NAD cycling system function in the NAD biosynthesis in A. niger. A part of the radioactive precursors was also incorporated into an unknown compound. (auth.)

Effect of bile on growth, peritoneal absorption, and blood clearance of Escherichia coli in E coli peritonitis

International Nuclear Information System (INIS)

Andersson, R.; Schalen, C.; Tranberg, K.G.

1991-01-01

The effect of intraperitoneal bile on growth, peritoneal absorption, and clearance of Escherichia coli was determined in E coli peritonitis in the rat. In E coli peritonitis, intraperitoneal bacterial counts gradually decreased, whereas they increased (after 2 hours) with subsequent development of bacteremia in E coli plus bile peritonitis. After an intraperitoneal injection of labeled bacteria, blood radioactivity was only initially lower in E coli plus bile peritonitis compared with E coli peritonitis. Clearance from blood was lower in E coli plus bile peritonitis than in E coli peritonitis. Organ localization was similar in E coli peritonitis and E coli plus bile peritonitis with decreased splenic, increased pulmonary, and unchanged hepatic uptakes compared with controls. Impaired peritoneal absorption of bacteria, together with impaired local host defense, is likely to enhance the noxious effect of bile in E coli peritonitis

Remobilization of Phytol from Chlorophyll Degradation Is Essential for Tocopherol Synthesis and Growth of Arabidopsis

Science.gov (United States)

vom Dorp, Katharina; Hölzl, Georg; Plohmann, Christian; Eisenhut, Marion; Abraham, Marion

2015-01-01

Phytol from chlorophyll degradation can be phosphorylated to phytyl-phosphate and phytyl-diphosphate, the substrate for tocopherol (vitamin E) synthesis. A candidate for the phytyl-phosphate kinase from Arabidopsis thaliana (At1g78620) was identified via a phylogeny-based approach. This gene was designated VITAMIN E DEFICIENT6 (VTE6) because the leaves of the Arabidopsis vte6 mutants are tocopherol deficient. The vte6 mutant plants are incapable of photoautotrophic growth. Phytol and phytyl-phosphate accumulate, and the phytyl-diphosphate content is strongly decreased in vte6 leaves. Phytol feeding and enzyme assays with Arabidopsis and recombinant Escherichia coli cells demonstrated that VTE6 has phytyl-P kinase activity. Overexpression of VTE6 resulted in increased phytyl-diphosphate and tocopherol contents in seeds, indicating that VTE6 encodes phytyl-phosphate kinase. The severe growth retardation of vte6 mutants was partially rescued by introducing the phytol kinase mutation vte5. Double mutant plants (vte5 vte6) are tocopherol deficient and contain more chlorophyll, but reduced amounts of phytol and phytyl-phosphate compared with vte6 mutants, suggesting that phytol or phytyl-phosphate are detrimental to plant growth. Therefore, VTE6 represents the missing phytyl-phosphate kinase, linking phytol release from chlorophyll with tocopherol synthesis. Moreover, tocopherol synthesis in leaves depends on phytol derived from chlorophyll, not on de novo synthesis of phytyl-diphosphate from geranylgeranyl-diphosphate. PMID:26452599

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

Self irradiation effects on the thorium phosphate diphosphate dissolution (TPD): simulation by external irradiations

International Nuclear Information System (INIS)

Tamain, C.; Ozgumus, A.; Dacheux, N.; Garrido, F.; Thome, L.; Corbel, C.; Genet, M.

2004-01-01

The Thorium Phosphate Diphosphate (TPD), proposed as a ceramic for the long term immobilization of actinides, was externally irradiated with several ions and energies (but also with gamma rays) in order to simulate the self-irradiation. The influence of the electronic energy loss was first investigated. Thus, the XRD measurements have shown a complete amorphization of the material under 10 13 ions of Kr.cm -2 , while no significant structural change occurred after 5.10 13 S.cm -2 , 2.10 16 He.cm -2 or 320 kGy of dose of gamma rays. The dissolution of the raw and irradiated pellets was studied versus several parameters such as amorphized fraction, energy loss of incident ions, radiolytic species produced in situ in the leachate during irradiation (such as H 2 O 2 ), temperature and acidity. The results reveal an important increase of the dissolution kinetics for amorphized pellets compared to raw ceramic. (authors)

Activation of G-proteins by receptor-stimulated nucleoside diphosphate kinase in Dictyostelium.

Science.gov (United States)

Bominaar, A A; Molijn, A C; Pestel, M; Veron, M; Van Haastert, P J

1993-01-01

Recently, interest in the enzyme nucleoside diphosphate kinase (EC2.7.4.6) has increased as a result of its possible involvement in cell proliferation and development. Since NDP kinase is one of the major sources of GTP in cells, it has been suggested that the effects of an altered NDP kinase activity on cellular processes might be the result of altered transmembrane signal transduction via guanine nucleotide-binding proteins (G-proteins). In the cellular slime mould Dictyostelium discoideum, extracellular cAMP induces an increase of phospholipase C activity via a surface cAMP receptor and G-proteins. In this paper it is demonstrated that part of the cellular NDP kinase is associated with the membrane and stimulated by cell surface cAMP receptors. The GTP produced by the action of NDP kinase is capable of activating G-proteins as monitored by altered G-protein-receptor interaction and the activation of the effector enzyme phospholipase C. Furthermore, specific monoclonal antibodies inhibit the effect of NDP kinase on G-protein activation. These results suggest that receptor-stimulated NDP kinase contributes to the mediation of hormone action by producing GTP for the activation of GTP-binding proteins. Images PMID:8389692

Suppression of the E. coli SOS response by dNTP pool changes.

Science.gov (United States)

Maslowska, Katarzyna H; Makiela-Dzbenska, Karolina; Fijalkowska, Iwona J; Schaaper, Roel M

2015-04-30

The Escherichia coli SOS system is a well-established model for the cellular response to DNA damage. Control of SOS depends largely on the RecA protein. When RecA is activated by single-stranded DNA in the presence of a nucleotide triphosphate cofactor, it mediates cleavage of the LexA repressor, leading to expression of the 30(+)-member SOS regulon. RecA activation generally requires the introduction of DNA damage. However, certain recA mutants, like recA730, bypass this requirement and display constitutive SOS expression as well as a spontaneous (SOS) mutator effect. Presently, we investigated the possible interaction between SOS and the cellular deoxynucleoside triphosphate (dNTP) pools. We found that dNTP pool changes caused by deficiencies in the ndk or dcd genes, encoding nucleoside diphosphate kinase and dCTP deaminase, respectively, had a strongly suppressive effect on constitutive SOS expression in recA730 strains. The suppression of the recA730 mutator effect was alleviated in a lexA-deficient background. Overall, the findings suggest a model in which the dNTP alterations in the ndk and dcd strains interfere with the activation of RecA, thereby preventing LexA cleavage and SOS induction. Published by Oxford University Press on behalf of Nucleic Acids Research 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Clinical implications of thymidylate synthetase, dihydropyrimidine dehydrogenase and orotate phosphoribosyl transferase activity levels in colorectal carcinoma following radical resection and administration of adjuvant 5-FU chemotherapy

International Nuclear Information System (INIS)

Ishikawa, Masashi; Miyauchi, Takayuki; Kashiwagi, Yutaka

2008-01-01

A number of studies have investigated whether the activity levels of enzymes involved in 5-fluorouracil (5-FU) metabolism are prognostic factors for survival in patients with colorectal carcinoma. Most reports have examined thymidylate synthetase (TS) and dihydropyrimidine dehydrogenase (DPD) in unresectable or metastatic cases, therefore it is unclear whether the activity of these enzymes is of prognostic value in colorectal cancer patients treated with radical resection and adjuvant chemotherapy with 5-FU. This study examined fresh frozen specimens of colorectal carcinoma from 40 patients who had undergone curative operation and were orally administered adjuvant tegafur/uracil (UFT) chemotherapy. TS, DPD and orotate phosphoribosyl transferase (OPRT) activities were assayed in cancer tissue and adjacent normal tissue and their association with clinicopathological variables was investigated. In addition, the relationships between TS, DPD and OPRT activities and patient survival were examined to determine whether any of these enzymes could be useful prognostic factors. While there was no clear relationship between pathological findings and TS or DPD activity, OPRT activity was significantly lower in tumors with lymph node metastasis than in tumors lacking lymph node metastasis. Postoperative survival was significantly better in the groups with low TS activity and/or high OPRT activity. TS and OPRT activity levels in tumor tissue may be important prognostic factors for survival in Dukes' B and C colorectal carcinoma with radical resection and adjuvant chemotherapy with UFT

Biosynthesis of the Common Polysaccharide Antigen of Pseudomonas aeruginosa PAO1: Characterization and Role of GDP-D-Rhamnose:GlcNAc/GalNAc-Diphosphate-Lipid α1,3-D-Rhamnosyltransferase WbpZ.

Science.gov (United States)

Wang, Shuo; Hao, Youai; Lam, Joseph S; Vlahakis, Jason Z; Szarek, Walter A; Vinnikova, Anna; Veselovsky, Vladimir V; Brockhausen, Inka

2015-06-15

The opportunistic pathogen Pseudomonas aeruginosa produces two major cell surface lipopolysaccharides, characterized by distinct O antigens, called common polysaccharide antigen (CPA) and O-specific antigen (OSA). CPA contains a polymer of D-rhamnose (D-Rha) in α1-2 and α1-3 linkages. Three putative glycosyltransferase genes, wbpX, wbpY, and wbpZ, are part of the CPA biosynthesis cluster. To characterize the enzymatic function of the wbpZ gene product, we chemically synthesized the donor substrate GDP-D-Rha and enzymatically synthesized GDP-D-[(3)H]Rha. Using nuclear magnetic resonance (NMR) spectroscopy, we showed that WbpZ transferred one D-Rha residue from GDP-D-Rha in α1-3 linkage to both GlcNAc- and GalNAc-diphosphate-lipid acceptor substrates. WbpZ is also capable of transferring D-mannose (D-Man) to these acceptors. Therefore, WbpZ has a relaxed specificity with respect to both acceptor and donor substrates. The diphosphate group of the acceptor, however, is required for activity. WbpZ does not require divalent metal ion for activity and exhibits an unusually high pH optimum of 9. WbpZ from PAO1 is therefore a GDP-D-Rha:GlcNAc/GalNAc-diphosphate-lipid α1,3-D-rhamnosyltransferase that has significant activity of GDP-D-Man:GlcNAc/GalNAc-diphosphate-lipid α1,3-D-mannosyltransferase. We used site-directed mutagenesis to replace the Asp residues of the two DXD motifs with Ala. Neither of the mutant constructs of wbpZ (D172A or D254A) could be used to rescue CPA biosynthesis in the ΔwbpZ knockout mutant in a complementation assay. This suggested that D172 and D254 are essential for WbpZ function. This work is the first detailed characterization study of a D-Rha-transferase and a critical step in the development of CPA synthesis inhibitors. This is the first characterization of a D-rhamnosyltransferase and shows that it is essential in Pseudomonas aeruginosa for the synthesis of the common polysaccharide antigen. Copyright © 2015, American Society for

Annual Surveillance Summary: Escherichia coli (E. coli) Infections in the Military Health System (MHS), 2015

Science.gov (United States)

2017-03-01

Annual Surveillance Summary: Escherichia coli ( E . coli ) Infections in the Military Health System (MHS...or position of the Department of the Navy, Department of Defense, nor the U.S. Government. i i E . coli in the MHS: Annual Summary 2015 Prepared...March 2017 EpiData Center Department NMCPHC-EDC-TR-187-2017 ii ii E . coli in the MHS: Annual Summary 2015 Prepared March 2017 EpiData

Protective effects of indigenous Escherichia coli against a pathogenic E. coli challenge strain in pigs.

Science.gov (United States)

Vahjen, W; Cuisiniere, T; Zentek, J

2017-10-13

To investigate the inhibitory effect of indigenous enterobacteria on pathogenic Escherichia coli, a challenge trial with postweaning pigs was conducted. A pathogenic E. coli strain was administered to all animals and their health was closely monitored thereafter. Faecal samples were taken from three healthy and three diarrhoeic animals. Samples were cultivated on MacConkey agar and isolates were subcultured. A soft agar overlay assay was used to determine the inhibitory activity of the isolates. A total of 1,173 enterobacterial isolates were screened for their ability to inhibit the E. coli challenge strain. Colony forming units of enterobacteria on MacConkey agar were not different between healthy and diarrhoeic animals in the original samples. Furthermore, numbers of isolates per animal were also not significantly different between healthy (482 isolates) and diarrhoeic animals (691 isolates). A total of 43 isolates (3.7%) with inhibitory activity against the pathogenic E. coli challenge strain were detected. All inhibitory isolates were identified as E. coli via MALDI-TOF. The isolates belonged to the phylotypes A, C and E. Many isolates (67.4%) were commensal E. coli without relevant porcine pathogenic factors, but toxin- and fimbrial genes (stx2e, fae, estIb, elt1a, fas, fan) were detected in 14 inhibitory isolates. Healthy animals showed significantly (P=0.003) more inhibitory isolates (36 of 482 isolates; 7.5%) than diseased animals (7 of 691 isolates; 1.0%). There were no significant correlations regarding phylotype or pathogenic factors between healthy and diseased animals. This study has shown that a small proportion of indigenous E. coli is able to inhibit in vitro growth of a pathogenic E. coli strain in pigs. Furthermore, healthy animals possess significantly more inhibitory E. coli strains than diarrhoeic animals. The inhibition of pathogenic E. coli by specific indigenous E. coli strains may be an underlying principle for the containment of pathogenic

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

Impact of dry chilling on the genetic diversity of Escherichia coli on beef carcasses and on the survival of E. coli and E. coli O157.

Science.gov (United States)

Visvalingam, Jeyachchandran; Liu, Yang; Yang, Xianqin

2017-03-06

The objective of this study was to examine the effect of dry chilling on the genetic diversity of naturally occurring Escherichia coli on beef carcasses, and to examine whether two populations of E. coli recovered from carcasses during chilling and E. coli O157 differed in their response to desiccation. Isolates of E. coli were obtained from beef carcasses during a 67h dry chilling process and were genotyped using multiple-locus variable-number tandem-repeat analysis (MLVA). Ten E. coli genotypes found only at 0h (group A) and found more than once (group B), as well as five strains of E. coli O157 (group C) were inoculated on stainless steel coupons and their survival was examined after exposure to 75 and 100% relative humidity (RH) at 0 or 35°C for 67h. A total of 450 E. coli isolates were obtained, with 254, 49, 49, 51, 23, 20, and 4 from 0, 1, 2, 4, 6, 8 and 24h of chilling, respectively. No E. coli were recovered at 67h. MLVA of the isolates revealed 173 distinct genotypes. Genetic diversity of E. coli isolates, defined as ratio of the number of isolates to the number of genotypes, remained between 2.3 and 1.3 during the 24h of chilling. All strains inoculated on stainless steel coupons and exposed to 75% RH at 35°C were completely inactivated, irrespective of their groups. Inactivation of E. coli of the three groups was not significantly (P>0.05) different by exposure to 75% RH at 0°C. The findings indicate that the genetic diversity of E. coli on beef carcasses was not affected by dry chilling. In addition, inactivation of E. coli genotypes and E. coli O157 by desiccation on stainless steel simulating dry chilling conditions did not differ significantly (P>0.05). Thus, dry chilling may be used as an effective antimicrobial intervention for beef carcasses. Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.

Can E. coli fly?

DEFF Research Database (Denmark)

Lindeberg, Yrja Lisa; Egedal, Karen; Hossain, Zenat Zebin

2018-01-01

, and the numbers of flies landing on the exposed rice were counted. Following exposure, the surface of the rice was microbiologically and molecularly analysed for the presence of E. coli and genes of diarrheagenic E. coli and Shigella strains. RESULTS: Rice was at greater risk (p ... with E. coli if flies landed on the rice than if no flies landed on the rice (odds ratio 5·4 (p ...-landings, the average CFU per fly-landing was > 0·6 x 103 CFU. Genes of diarrheagenic E. coli and Shigella species were detected in 39 of 60 (65%) of exposed rice samples. Two fly species were identified; the common housefly (Musca domestica) and the oriental latrine fly (Chrysomya megacephala). CONCLUSION: Flies may...

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

The oxygen effect in E. coli cells

International Nuclear Information System (INIS)

Myasnik, M.N.; Skvortsov, V.G.; Sokolov, V.A.

1982-01-01

In experiments on E. coli strains deficient in some stages of DNA repair from radiation damages, it was demonstrated that the value of the oxygen effect, under optimal conditions for manifestation thereof, decreases in the following order: E. coli WP2 (the wild type) → E. coli WP2 exr - and E. coli B → E. coli WP2 uvr A6 → E. coli WP2 rec Al and E. coli WP2 hcr - exr - . It was detected that 0.14 M NaCl solution sensitizes the anoxic cells of some E. coli strains to the effect of γ-radiation. It was established that mutation of the uvr A-gene increases sharply the sensitivity of cells to iradiation under the anoxic conditions in the presence of NaCl, the reverse'' oxygen effect being observed

The radioprotector WR-2721 reduces neutron-induced mutations at the hypoxanthine-guanine phosphoribosyl transferase locus in mouse splenocytes when administered prior to or following irradiation

International Nuclear Information System (INIS)

Grdina, D.J.; Basic, I.

1992-01-01

An in vitro T-lymphocyte cloning technique has been applied to study the effects of JANUS fission-spectrum neutron irradiation and the radioprotector S-2-(3-aminopropylamino) ethylphosphorothioic acid (WR-2721) on the subsequent development of somatic mutations at the hypoxanthine-guanine phosphoribosyl transferase (hprt) locus in hybrid B6CF1 male mice. In control studies performed to establish an in vitro cloning technique, the mutant frequencies of splenic T-lymphocytes, as a result of exposure to a 100 cGy dose of neutrons, increased with time from a control level of 9 x 10 -7 to a maximum value of 1.7 x 10 -5 at 56 days following irradiation. Between 56 and 150 days after irradiation, mutant frequencies were observed to plateau and remain stable. All subsequent determinations were performed at 56 days following the experimental treatment of animals. WR-2721 at a dose of 400 mg/kg was effective in protecting against the induction of hprt mutants (i.e. a mutant frequency reduction factor, MFRF) following the largest dose of neutrons used (i.e. 150 cGy). The antimutagenic effectiveness of WR-2721 administered 30 min prior to irradiation was unaffected, even when the dose was reduced to 200 mg/kg. These findings confirm our earlier report using the radioprotector N-(2-mercaptoethyl)-1,2-diaminopropane (WR-1065) under in vitro conditions, and demonstrate that these agents can be used as effective antimutagens even when they are administered up to 3 h following radiation exposure. (Author)

Hemolytic porcine intestinal Escherichia coli without virulence-associated genes typical of intestinal pathogenic E. coli.

Science.gov (United States)

Schierack, Peter; Weinreich, Joerg; Ewers, Christa; Tachu, Babila; Nicholson, Bryon; Barth, Stefanie

2011-12-01

Testing 1,666 fecal or intestinal samples from healthy and diarrheic pigs, we obtained hemolytic Escherichia coli isolates from 593 samples. Focusing on hemolytic E. coli isolates without virulence-associated genes (VAGs) typical for enteropathogens, we found that such isolates carried a broad variety of VAGs typical for extraintestinal pathogenic E. coli.

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.

Prevalence and behavior of multidrug-resistant shiga toxin-producing Escherichia coli, enteropathogenic E. coli and enterotoxigenic E. coli on coriander.

Science.gov (United States)

Gómez-Aldapa, Carlos A; Segovia-Cruz, Jesús A; Cerna-Cortes, Jorge F; Rangel-Vargas, Esmeralda; Salas-Rangel, Laura P; Gutiérrez-Alcántara, Eduardo J; Castro-Rosas, Javier

2016-10-01

The prevalence and behavior of multidrug-resistant diarrheagenic Escherichia coli pathotypes on coriander was determined. One hundred coriander samples were collected from markets. Generic E. coli were determined using the most probable number procedure. Diarrheagenic E. coli pathotypes (DEPs) were identified using two multiplex polymerase chain reaction procedures. Susceptibility to sixteen antibiotics was tested for the isolated DEPs strains by standard test. The behavior of multidrug-resistant DEPs isolated from coriander was determined on coriander leaves and chopped coriander at 25°± 2 °C and 3°± 2 °C. Generic E. coli and DEPs were identified, respectively, in 43 and 7% of samples. Nine DEPs strains were isolated from positive coriander samples. The identified DEPs included Shiga toxin-producing E. coli (STEC, 4%) enterotoxigenic E. coli (ETEC, 2%) and enteropathogenic E. coli (EPEC, 1%). All isolated DEPs strains exhibited multi-resistance to antibiotics. On inoculated coriander leaves stored at 25°± 2 °C or 3°± 2 °C, no growth was observed for multidrug-resistant DEPs strains. However, multidrug-resistant DEPs strains grew in chopped coriander: after 24 h at 25° ± 2 °C, DEPs strains had grown to approximately 3 log CFU/g. However, at 3°± 2 °C the bacterial growth was inhibited. To the best of our knowledge, this is the first report of the presence and behavior of multidrug-resistant STEC, ETEC and EPEC on coriander and chopped coriander. Copyright © 2016 Elsevier Ltd. All rights reserved.

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.

Quantitative and molecular analyses of mutation in a pSV2gpt transformed CHO cell line

International Nuclear Information System (INIS)

Stankowski, L.F. Jr.; Tindall, K.R.; Hsie, A.W.

1983-01-01

Following NDA-mediated gene transfer we have isolated a cell line useful for studying gene mutation at the molecular level. This line, AS52, derived from a hypoxanthine-guanine phosphoribosyl transferase (HGPRT) deficient Chinese hamster ovary (CHO) cell line, carries a single copy of the E. coli xanthine-guanine phosphoribosyl transferase (XGPRT) gene (gpt) and exhibits a spontaneous mutant frequency of 20 TG/sup r/ mutants/10 6 clonable cells. As with HGPRT - mutants, XGPRT - mutants can be selected in 6-thioguanine. AS52 (XGPRT + ) and wild type CHO (HGPRT + ) cell exhibit almost identical cytotoxic responses to various agents. We observed significant differences in mutation induction by UV light and ethyl methanesulfonate (EMS). Ratios of XGPRT - to HGPRT - mutants induced per unit dose (J/m 2 for UV light and μg/ml for EMS) are 1.4 and 0.70, respectively. Preliminary Southern blot hybridization analyses has been performed on 30 XGPRT - AS52 mutants. A majority of spontaneous mutants have deletions ranging in size from 1 to 4 kilobases (9/19) to complete loss of gpt sequences (4/19); the remainder have no detectable (5/19) or only minor (1/19) alterations. 5/5 UV-induced and 5/6 EMS-induced mutants do not show a detectable change. Similar analyses are underway for mutations induced by x-irradiation and ICR 191 treatment

The asymptomatic bacteriuria Escherichia coli strain 83972 outcompetes uropathogenic E. coli strains in human urine

DEFF Research Database (Denmark)

Hancock, Viktoria; Ulett, G.C.; Schembri, M.A.

2006-01-01

Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU). In contrast to uropathogenic E. coli (UPEC), which causes symptomatic urinary tract infections (UTI), very little is known about the mechanisms by which these strains colonize the human urinary tract....... The prototype ABU E. coli strain 83972 was originally isolated from a girl who had carried it asymptomatically for 3 years. Deliberate colonization of UTI-susceptible individuals with E. coli 83972 has been used successfully as an alternative approach for the treatment of patients who are refractory...... to conventional therapy. Colonization with strain 83972 appears to prevent infection with UPEC strains in such patients despite the fact that this strain is unable to express the primary adhesins involved in UTI, viz. P and type 1 fimbriae. Here we investigated the growth characteristics of E. coli 83972 in human...

Opportunities and challenges for the sustainable production of structurally complex diterpenoids in recombinant microbial systems.

Science.gov (United States)

Kemper, Katarina; Hirte, Max; Reinbold, Markus; Fuchs, Monika; Brück, Thomas

2017-01-01

With over 50.000 identified compounds terpenes are the largest and most structurally diverse group of natural products. They are ubiquitous in bacteria, plants, animals and fungi, conducting several biological functions such as cell wall components or defense mechanisms. Industrial applications entail among others pharmaceuticals, food additives, vitamins, fragrances, fuels and fuel additives. Central building blocks of all terpenes are the isoprenoid compounds isopentenyl diphosphate and dimethylallyl diphosphate. Bacteria like Escherichia coli harbor a native metabolic pathway for these isoprenoids that is quite amenable for genetic engineering. Together with recombinant terpene biosynthesis modules, they are very suitable hosts for heterologous production of high value terpenes. Yet, in contrast to the number of extracted and characterized terpenes, little is known about the specific biosynthetic enzymes that are involved especially in the formation of highly functionalized compounds. Novel approaches discussed in this review include metabolic engineering as well as site-directed mutagenesis to expand the natural terpene landscape. Focusing mainly on the validation of successful integration of engineered biosynthetic pathways into optimized terpene producing Escherichia coli , this review shall give an insight in recent progresses regarding manipulation of mostly diterpene synthases.

Opportunities and challenges for the sustainable production of structurally complex diterpenoids in recombinant microbial systems

Directory of Open Access Journals (Sweden)

Katarina Kemper

2017-05-01

Full Text Available With over 50.000 identified compounds terpenes are the largest and most structurally diverse group of natural products. They are ubiquitous in bacteria, plants, animals and fungi, conducting several biological functions such as cell wall components or defense mechanisms. Industrial applications entail among others pharmaceuticals, food additives, vitamins, fragrances, fuels and fuel additives. Central building blocks of all terpenes are the isoprenoid compounds isopentenyl diphosphate and dimethylallyl diphosphate. Bacteria like Escherichia coli harbor a native metabolic pathway for these isoprenoids that is quite amenable for genetic engineering. Together with recombinant terpene biosynthesis modules, they are very suitable hosts for heterologous production of high value terpenes. Yet, in contrast to the number of extracted and characterized terpenes, little is known about the specific biosynthetic enzymes that are involved especially in the formation of highly functionalized compounds. Novel approaches discussed in this review include metabolic engineering as well as site-directed mutagenesis to expand the natural terpene landscape. Focusing mainly on the validation of successful integration of engineered biosynthetic pathways into optimized terpene producing Escherichia coli, this review shall give an insight in recent progresses regarding manipulation of mostly diterpene synthases.

A genomically modified Escherichia coli strain carrying an orthogonal E. coli histidyl-tRNA synthetase•tRNAHis pair.

Science.gov (United States)

Englert, Markus; Vargas-Rodriguez, Oscar; Reynolds, Noah M; Wang, Yane-Shih; Söll, Dieter; Umehara, Takuya

2017-11-01

Development of new aminoacyl-tRNA synthetase (aaRS)•tRNA pairs is central for incorporation of novel non-canonical amino acids (ncAAs) into proteins via genetic code expansion (GCE). The Escherichia coli and Caulobacter crescentus histidyl-tRNA synthetases (HisRS) evolved divergent mechanisms of tRNA His recognition that prevent their cross-reactivity. Although the E. coli HisRS•tRNA His pair is a good candidate for GCE, its use in C. crescentus is limited by the lack of established genetic selection methods and by the low transformation efficiency of C. crescentus. E. coli was genetically engineered to use a C. crescentus HisRS•tRNA His pair. Super-folder green fluorescent protein (sfGFP) and chloramphenicol acetyltransferase (CAT) were used as reporters for read-through assays. A library of 313 ncAAs coupled with the sfGFP reporter system was employed to investigate the specificity of E. coli HisRS in vivo. A genomically modified E. coli strain (named MEOV1) was created. MEVO1 requires an active C. crescentus HisRS•tRNA His pair for growth, and displays a similar doubling time as the parental E. coli strain. sfGFP- and CAT-based assays showed that the E. coli HisRS•tRNA His pair is orthogonal in MEOV1 cells. A mutation in the anticodon loop of E. coli tRNA His CUA elevated its suppression efficiency by 2-fold. The C. crescentus HisRS•tRNA His pair functionally complements an E. coli ΔhisS strain. The E. coli HisRS•tRNA His is orthogonal in MEOV1 cells. E. coli tRNA His CUA is an efficient amber suppressor in MEOV1. We developed a platform that allows protein engineering of E. coli HisRS that should facilitate GCE in E. coli. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2017 Elsevier B.V. All rights reserved.

Silver nanoparticle-E. coli colloidal interaction in water and effect on E. coli survival.

Science.gov (United States)

Dror-Ehre, A; Mamane, H; Belenkova, T; Markovich, G; Adin, A

2009-11-15

Silver nanoparticles exhibit antibacterial properties via bacterial inactivation and growth inhibition. The mechanism is not yet completely understood. This work was aimed at elucidating the effect of silver nanoparticles on inactivation of Escherichia coli, by studying particle-particle interactions in aqueous suspensions. Stable, molecularly capped, positively or negatively charged silver nanoparticles were mixed at 1 to 60microgmL(-1) with suspended E. coli cells to examine their effect on inactivation of the bacteria. Gold nanoparticles with the same surfactant were used as a control, being of similar size but made up of a presumably inert metal. Log reduction of 5log(10) and complete inactivation were obtained with the silver nanoparticles while the gold nanoparticles did not show any inactivation ability. The effect of molecularly capped nanoparticles on E. coli survival was dependent on particle number. Log reduction of E. coli was associated with the ratio between the number of nanoparticles and the initial bacterial cell count. Electrostatic attraction or repulsion mechanisms in silver nanoparticle-E. coli cell interactions did not contribute to the inactivation process.

Surface complexation modeling of uranium (Vi) retained onto zirconium diphosphate in presence of organic acids

International Nuclear Information System (INIS)

Almazan T, M. G.; Garcia G, N.; Ordonez R, E.

2010-10-01

In the field of nuclear waste disposal, predictions regarding radionuclide migration through the geosphere, have to take account the effects of natural organic matter. This work presents an investigation of interaction mechanisms between U (Vi) and zirconium diphosphate (ZrP 2 O 7 ) in presence of organic acids (citric acid and oxalic acid). The retention reactions were previously examined using a batch equilibrium method. Previous results showed that U (Vi) retention was more efficient when citric acid or oxalic acid was present in solid surface at lower ph values. In order to determine the retention equilibria for both systems studied, a phosphorescence spectroscopy study was carried out. The experimental data were then fitted using the Constant Capacitance Model included in the FITEQL4.0 code. Previous results concerning surface characterization of ZrP 2 O 7 (surface sites density and surface acidity constants) were used to constraint the modeling. The best fit for U (Vi)/citric acid/ZrP 2 O 7 and U (Vi)/oxalic acid/ZrP 2 O 7 systems considered the formation of a ternary surface complex. (Author)

Direct incorporation of guanosine 5'-diphosphate into microtubules without guanosine 5'-triphosphate hydrolysis

International Nuclear Information System (INIS)

Hamel, E.; Batra, J.K.; Lin, C.M.

1986-01-01

Using highly purified calf brain tubulin bearing [8- 14 C]guanosine 5'-diphosphate (GDP) in the exchangeable nucleotide site and heat-treated microtubule-associated proteins, the authors have found that a significant proportion of exchangeable-site GDP in microtubules can be incorporated directly during guanosine 5'-triphosphate (GTP) dependent polymerization of tubulin, without an initial exchange of GDP for GTP and subsequent GTP hydrolysis during assembly. The precise amount of GDP incorporated directly into microtubules is highly dependent on specific reaction conditions, being favored by high tubulin concentrations, low GTP and Mg 2+ concentrations, and exogenous GDP in the reaction mixture. Minimum effects were observed with changes in reaction pH or temperature, changes in concentration of microtubule-associated proteins, alteration of the sulfonate buffer, or the presence of a calcium chelator in the reaction mixture. Under conditions most favorable for direct GDP incorporation, about one-third of the GDP in microtubules is incorporated directly (without GTP hydrolysis) and two-thirds is incorporated hydrolytically (as a consequence of GTP hydrolysis). Direct incorporation of GDP occurs in a constant proportion throughout elongation, and the amount of direct incorporation probably reflects the rapid equilibration of GDP and GTP at the exchangeable site that occurs before the onset of assembly

Identification and Functional Characterization of Monofunctional ent-Copalyl Diphosphate and ent-Kaurene Synthases in White Spruce Reveal Different Patterns for Diterpene Synthase Evolution for Primary and Secondary Metabolism in Gymnosperms1[W][OA

Science.gov (United States)

Keeling, Christopher I.; Dullat, Harpreet K.; Yuen, Mack; Ralph, Steven G.; Jancsik, Sharon; Bohlmann, Jörg

2010-01-01

The biosynthesis of the tetracyclic diterpene ent-kaurene is a critical step in the general (primary) metabolism of gibberellin hormones. ent-Kaurene is formed by a two-step cyclization of geranylgeranyl diphosphate via the intermediate ent-copalyl diphosphate. In a lower land plant, the moss Physcomitrella patens, a single bifunctional diterpene synthase (diTPS) catalyzes both steps. In contrast, in angiosperms, the two consecutive cyclizations are catalyzed by two distinct monofunctional enzymes, ent-copalyl diphosphate synthase (CPS) and ent-kaurene synthase (KS). The enzyme, or enzymes, responsible for ent-kaurene biosynthesis in gymnosperms has been elusive. However, several bifunctional diTPS of specialized (secondary) metabolism have previously been characterized in gymnosperms, and all known diTPSs for resin acid biosynthesis in conifers are bifunctional. To further understand the evolution of ent-kaurene biosynthesis as well as the evolution of general and specialized diterpenoid metabolisms in gymnosperms, we set out to determine whether conifers use a single bifunctional diTPS or two monofunctional diTPSs in the ent-kaurene pathway. Using a combination of expressed sequence tag, full-length cDNA, genomic DNA, and targeted bacterial artificial chromosome sequencing, we identified two candidate CPS and KS genes from white spruce (Picea glauca) and their orthologs in Sitka spruce (Picea sitchensis). Functional characterization of the recombinant enzymes established that ent-kaurene biosynthesis in white spruce is catalyzed by two monofunctional diTPSs, PgCPS and PgKS. Comparative analysis of gene structures and enzyme functions highlights the molecular evolution of these diTPSs as conserved between gymnosperms and angiosperms. In contrast, diTPSs for specialized metabolism have evolved differently in angiosperms and gymnosperms. PMID:20044448

The Biosynthetic Origin of Irregular Monoterpenes in Lavandula

Science.gov (United States)

Demissie, Zerihun A.; Erland, Lauren A. E.; Rheault, Mark R.; Mahmoud, Soheil S.

2013-01-01

Lavender essential oils are constituted predominantly of regular monoterpenes, for example linalool, 1,8-cineole, and camphor. However, they also contain irregular monoterpenes including lavandulol and lavandulyl acetate. Although the majority of genes responsible for the production of regular monoterpenes in lavenders are now known, enzymes (including lavandulyl diphosphate synthase (LPPS)) catalyzing the biosynthesis of irregular monoterpenes in these plants have not been described. Here, we report the isolation and functional characterization of a novel cis-prenyl diphosphate synthase cDNA, termed Lavandula x intermedia lavandulyl diphosphate synthase (LiLPPS), through a homology-based cloning strategy. The LiLPPS ORF, encoding for a 305-amino acid long protein, was expressed in Escherichia coli, and the recombinant protein was purified by nickel-nitrilotriacetic acid affinity chromatography. The approximately 34.5-kDa bacterially produced protein specifically catalyzed the head-to-middle condensation of two dimethylallyl diphosphate units to LPP in vitro with apparent Km and kcat values of 208 ± 12 μm and 0.1 s−1, respectively. LiLPPS is a homodimeric enzyme with a sigmoidal saturation curve and Hill coefficient of 2.7, suggesting a positive co-operative interaction among its catalytic sites. LiLPPS could be used to modulate the production of lavandulol and its derivatives in plants through metabolic engineering. PMID:23306202

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

Comparative genomics and transcriptomics of Escherichia coli isolates carrying virulence factors of both enteropathogenic and enterotoxigenic E. coli.

Science.gov (United States)

Hazen, Tracy H; Michalski, Jane; Luo, Qingwei; Shetty, Amol C; Daugherty, Sean C; Fleckenstein, James M; Rasko, David A

2017-06-14

Escherichia coli that are capable of causing human disease are often classified into pathogenic variants (pathovars) based on their virulence gene content. However, disease-associated hybrid E. coli, containing unique combinations of multiple canonical virulence factors have also been described. Such was the case of the E. coli O104:H4 outbreak in 2011, which caused significant morbidity and mortality. Among the pathovars of diarrheagenic E. coli that cause significant human disease are the enteropathogenic E. coli (EPEC) and enterotoxigenic E. coli (ETEC). In the current study we use comparative genomics, transcriptomics, and functional studies to characterize isolates that contain virulence factors of both EPEC and ETEC. Based on phylogenomic analysis, these hybrid isolates are more genomically-related to EPEC, but appear to have acquired ETEC virulence genes. Global transcriptional analysis using RNA sequencing, demonstrated that the EPEC and ETEC virulence genes of these hybrid isolates were differentially-expressed under virulence-inducing laboratory conditions, similar to reference isolates. Immunoblot assays further verified that the virulence gene products were produced and that the T3SS effector EspB of EPEC, and heat-labile toxin of ETEC were secreted. These findings document the existence and virulence potential of an E. coli pathovar hybrid that blurs the distinction between E. coli pathovars.

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

Influence of the temperature in the uranium (Vi) sorption in zirconium diphosphate; Influencia de la temperatura en la sorcion de uranio (VI) en difosfato de circonio

Energy Technology Data Exchange (ETDEWEB)

Garcia G, N.; Solis, D. [Universidad Autonoma del Estado de Mexico, Facultad de Quimica, Paseo Colon y Paseo Tollocan, 50120 Toluca, Estado de Mexico (Mexico); Ordonez R, E., E-mail: nidgg@yahoo.com.mx [ININ, Departamento de Quimica, Carretera Mexico-Toluca s/n, 52750 Ocoyoacac, Estado de Mexico (Mexico)

2012-10-15

In the present work was evaluated the uranium (Vi) sorption at 10, 20, 30, 40 and 60 C on the zirconium diphosphate (ZrP{sub 2}O{sub 7}). They were carried out kinetic and isotherms using the method by lots, these will allow to fix the sorption time (kinetic) and to explain the behavior of this sorption in different ph conditions and temperature (isotherm). The quantity of retained uranium in the surface was quantified by means of the fluorescence technique. (Author)

Hemolytic Porcine Intestinal Escherichia coli without Virulence-Associated Genes Typical of Intestinal Pathogenic E. coli ▿ †

Science.gov (United States)

Schierack, Peter; Weinreich, Joerg; Ewers, Christa; Tachu, Babila; Nicholson, Bryon; Barth, Stefanie

2011-01-01

Testing 1,666 fecal or intestinal samples from healthy and diarrheic pigs, we obtained hemolytic Escherichia coli isolates from 593 samples. Focusing on hemolytic E. coli isolates without virulence-associated genes (VAGs) typical for enteropathogens, we found that such isolates carried a broad variety of VAGs typical for extraintestinal pathogenic E. coli. PMID:21965399

Revisiting the Latency of Uridine Diphosphate-Glucuronosyltransferases (UGTs—How Does the Endoplasmic Reticulum Membrane Influence Their Function?

Directory of Open Access Journals (Sweden)

Yuejian Liu

2017-08-01

Full Text Available Uridine diphosphate-glucuronosyltransferases (UGTs are phase 2 conjugation enzymes mainly located in the endoplasmic reticulum (ER of the liver and many other tissues, and can be recovered in artificial ER membrane preparations (microsomes. They catalyze glucuronidation reactions in various aglycone substrates, contributing significantly to the body’s chemical defense mechanism. There has been controversy over the last 50 years in the UGT field with respect to the explanation for the phenomenon of latency: full UGT activity revealed by chemical or physical disruption of the microsomal membrane. Because latency can lead to inaccurate measurements of UGT activity in vitro, and subsequent underprediction of drug clearance in vivo, it is important to understand the mechanisms behind this phenomenon. Three major hypotheses have been advanced to explain UGT latency: compartmentation, conformation, and adenine nucleotide inhibition. In this review, we discuss the evidence behind each hypothesis in depth, and suggest some additional studies that may reveal more information on this intriguing phenomenon.

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...

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...

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...

Cloning of a sesquiterpene synthase from Lavandula x intermedia glandular trichomes.

Science.gov (United States)

Sarker, Lukman S; Demissie, Zerihun A; Mahmoud, Soheil S

2013-11-01

The essential oil (EO) of Lavandula is dominated by monoterpenes, but can also contain small amounts of sesquiterpenes, depending on species and environmental conditions. For example, the sesquiterpene 9-epi-caryophyllene can make up to 8 % of the EO in a few species, including those commercially propagated for EO production. Here, we report the cloning and functional characterization of 9-epi-caryophyllene synthase (LiCPS) from the glandular trichomes of Lavandula x intermedia, cv. Grosso. The 1,617 bp open reading frame of LiCPS, which did not encode a transit peptide, was expressed in Escherichia coli and the recombinant protein purified by Ni-NTA agarose affinity chromatography. The ca. 60 kDa recombinant protein specifically converted farnesyl diphosphate to 9-epi-caryophyllene. LiCPS also produced a few monoterpenes when assayed with the monoterpene precursor geranyl diphosphate (GPP), but--unlike most monoterpene synthases--was not able to derive detectable amounts of any products from the cis isomer of GPP, neryl diphosphate. The LiCPS transcripts accumulated in developing L. x intermedia flowers and were highly enriched in glandular trichomes, but were not detected in leaves suggesting that the transcriptional expression of this gene is spatially and developmentally regulated.

Molecular cloning and characterization of a cDNA encoding the gibberellin biosynthetic enzyme ent-kaurene synthase B from pumpkin (Cucurbita maxima L.).

Science.gov (United States)

Yamaguchi, S; Saito, T; Abe, H; Yamane, H; Murofushi, N; Kamiya, Y

1996-08-01

The first committed step in the formation of diterpenoids leading to gibberellin (GA) biosynthesis is the conversion of geranylgeranyl diphosphate (GGDP) to ent-kaurene. ent-Kaurene synthase A (KSA) catalyzes the conversion of GGDP to copalyl diphosphate (CDP), which is subsequently converted to ent-kaurene by ent-kaurene synthase B (KSB). A full-length KSB cDNA was isolated from developing cotyledons in immature seeds of pumpkin (Cucurbita maxima L.). Degenerate oligonucleotide primers were designed from the amino acid sequences obtained from the purified protein to amplify a cDNA fragment, which was used for library screening. The isolated full-length cDNA was expressed in Escherichia coli as a fusion protein, which demonstrated the KSB activity to cyclize [3H]CDP to [3H]ent-kaurene. The KSB transcript was most abundant in growing tissues, but was detected in every organ in pumpkin seedlings. The deduced amino acid sequence shares significant homology with other terpene cyclases, including the conserved DDXXD motif, a putative divalent metal ion-diphosphate complex binding site. A putative transit peptide sequence that may target the translated product into the plastids is present in the N-terminal region.

The uridine diphosphate glucuronosyltransferases: quantitative structure-activity relationships for hydroxyl polychlorinated biphenyl substrates

Energy Technology Data Exchange (ETDEWEB)

Wang, Degao [Dalian University of Technology, Department of Environmental Science and Technology, Dalian (China)

2005-10-01

Quantitative structure-activity relationships (QSARs), which relate the glucuronidation of hydroxyl polychlorinated biphenyls (OH-PCBs) - catalyzed by the uridine diphosphate glucuronosyltransferases (UGTs) - to their physicochemical properties and molecular structural parameters, can be used to predict the rate constants and interpret the mechanism of glucuronidation. In this study, QSARs have been developed that use 23 semi-empirical calculated quantum chemical descriptors to predict the logarithms of the constants 1/K{sub m} and V{sub max}, related to enzyme kinetics. A partial least squares regression method was used to select the optimal set of descriptors to minimize the multicollinearity between the descriptors, as well as to maximize the cross-validated coefficient (Q{sup 2} {sub cum}) values. The key descriptors affecting log(1/K{sub m}) were E{sub lumo}- E{sub homo} (the energy gap between the lowest unoccupied molecular orbital and the highest occupied molecular orbital) and q{sub C}{sup -} (the largest negative net atomic charge on a carbon atom), while the key descriptors affecting log V{sub max} were the polarizability {alpha}, the Connolly solvent-excluded volume (CSEV), and logP (the logarithm of the partition coefficient for octanol/water). From the results obtained it can be concluded that hydrophobic and electronic aspects of OH-PCBs are important in the glucuronidation of OH-PCBs. (orig.)

Surface and micellar properties of Chloroquine Diphosphate and its interactions with surfactants and Human Serum Albumin

International Nuclear Information System (INIS)

Usman, Muhammad; Siddiq, Mohammad

2013-01-01

Highlights: ► Free energy of adsorption is more negative than free energy of micellization. ► Shifts in UV/Visible spectra in presence of SDS indicated interaction of CLQ with SDS. ► The decrease in fluorescence intensity of HSA by CLQ shows its binding with HSA. -- Abstract: This manuscript addresses the physicochemical behavior of an antimalarial drug Chloroquine Diphosphate (CLQ) as well as its interaction with anionic surfactants and Human Serum Albumin (HSA). Surface tension and specific conductivity were employed to detect the critical micelle concentration (CMC) and thus its surface and thermodynamic parameters were calculated. Solubilization of this drug within micelles of anionic surfactant sodium dodecyl sulfate (SDS) has also been studied. UV/Visible spectroscopy was used to calculate partition coefficient (K x ), free energy of partition and number of drug molecules per micelle. The complexation of drug with HSA at physiological conditions (pH 7.4) has also been analyzed by using UV/Visible and fluorescence spectroscopy. The values of drug-protein binding constant, number of binding sites and free energy of binding were calculated

Escherichia coli EDL933 Requires Gluconeogenic Nutrients To Successfully Colonize the Intestines of Streptomycin-Treated Mice Precolonized with E. coli Nissle 1917

Science.gov (United States)

Schinner, Silvia A. C.; Mokszycki, Matthew E.; Adediran, Jimmy; Leatham-Jensen, Mary; Conway, Tyrrell

2015-01-01

Escherichia coli MG1655, a K-12 strain, uses glycolytic nutrients exclusively to colonize the intestines of streptomycin-treated mice when it is the only E. coli strain present or when it is confronted with E. coli EDL933, an O157:H7 strain. In contrast, E. coli EDL933 uses glycolytic nutrients exclusively when it is the only E. coli strain in the intestine but switches in part to gluconeogenic nutrients when it colonizes mice precolonized with E. coli MG1655 (R. L. Miranda et al., Infect Immun 72:1666–1676, 2004, http://dx.doi.org/10.1128/IAI.72.3.1666-1676.2004). Recently, J. W. Njoroge et al. (mBio 3:e00280-12, 2012, http://dx.doi.org/10.1128/mBio.00280-12) reported that E. coli 86-24, an O157:H7 strain, activates the expression of virulence genes under gluconeogenic conditions, suggesting that colonization of the intestine with a probiotic E. coli strain that outcompetes O157:H7 strains for gluconeogenic nutrients could render them nonpathogenic. Here we report that E. coli Nissle 1917, a probiotic strain, uses both glycolytic and gluconeogenic nutrients to colonize the mouse intestine between 1 and 5 days postfeeding, appears to stop using gluconeogenic nutrients thereafter in a large, long-term colonization niche, but continues to use them in a smaller niche to compete with invading E. coli EDL933. Evidence is also presented suggesting that invading E. coli EDL933 uses both glycolytic and gluconeogenic nutrients and needs the ability to perform gluconeogenesis in order to colonize mice precolonized with E. coli Nissle 1917. The data presented here therefore rule out the possibility that E. coli Nissle 1917 can starve the O157:H7 E. coli strain EDL933 of gluconeogenic nutrients, even though E. coli Nissle 1917 uses such nutrients to compete with E. coli EDL933 in the mouse intestine. PMID:25733524

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.

Crystallization and preliminary X-ray diffraction study of recombinant adenine phosphoribosyltransferase from the thermophilic bacterium Thermus thermophilus strain HB27

Science.gov (United States)

Sinitsyna, E. V.; Timofeev, V. I.; Tuzova, E. S.; Kostromina, M. A.; Murav'eva, T. I.; Esipov, R. S.; Kuranova, I. P.

2017-07-01

Adenine phosphoribosyltransferase (APRT) belongs to the type I phosphoribosyltransferase family and catalyzes the formation of adenosine monophosphate via transfer of the 5-phosphoribosyl group from phosphoribosyl pyrophosphate to the nitrogen atom N9 of the adenine base. Proteins of this family are involved in a salvage pathway of nucleotide synthesis, thus providing purine base utilization and maintaining the optimal level of purine bases in the body. Adenine phosphoribosyltransferase from the extremely thermophilic Thermus thermophilus strain HB27 was produced using a highly efficient E. coli producer strain and was then purified by affinity and gel-filtration chromatography. This enzyme was successfully employed as a catalyst for the cascade biosynthesis of biologically important nucleotides. The screening of crystallization conditions for recombinant APRT from T. thermophilus HB27 was performed in order to determine the enzyme structure by X-ray diffraction. The crystallization conditions, which were found by the vapor-diffusion technique, were then optimized to apply the counter-diffusion technique. The crystals of the enzyme were grown by the capillary counter-diffusion method. The crystals belong to sp. gr. P1211 and have the following unitcell parameters: a = 69.86 Å, b = 82.16 Å, c = 91.39 Å, α = γ = 90°, β = 102.58°. The X-ray diffraction data set suitable for the determination of the APRT structure at 2.6 Å resolution was collected from the crystals at the SPring-8 synchrotron facility (Japan).

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.

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.

Experimental induced avian E. coli salpingitis

DEFF Research Database (Denmark)

Olsen, Rikke Heidemann; Thøfner, Ida; Pors, Susanne Elisabeth

2016-01-01

Several types of Escherichia coli have been associated with extra-intestinal infections in poultry, however, they may vary significantly in their virulence potential. The aim of the present study was to investigate the virulence of five strains of E. coli obtained from different disease......) had a distinct ability to cause disease. Results of the study shows major differences in virulence of different strains of E. coli in ascending infections; however, there was no indication of tissue-specific adaptation, since strains obtained from lesions unrelated to the reproductive system were...... fully capable of causing experimental infection. In conclusion, the study provides evidence for the clinical outcome of infection with E. coli in poultry is largely influenced by the specific strain as well as individual host factors....

Inactivation and Gene Expression of a Virulent WastewaterEscherichia coliStrain and the Nonvirulent CommensalEscherichia coliDSM1103 Strain upon Solar Irradiation

KAUST Repository

Aljassim, Nada I.; Mantilla-Calderon, David; Wang, Tiannyu; Hong, Pei-Ying

2017-01-01

This study examined the decay kinetics and molecular responses of two Escherichia coli strains upon solar irradiation. The first is E. coli PI-7, a virulent and antibiotic-resistant strain that was isolated from wastewater and carries the emerging NDM-1 antibiotic resistance gene. The other strain, E. coli DSM1103, displayed lower virulence and antibiotic resistance than E. coli PI-7. In a buffer solution, E. coli PI-7 displayed a longer lag phase prior to decay and a longer half-life compared with E. coli DSM1103 (6.64 ± 0.63 h and 2.85 ± 0.46 min vs 1.33 ± 0.52 h and 2.04 ± 0.36 min). In wastewater, both E. coli strains decayed slower than they did in buffer. Although solar irradiation remained effective in reducing the numbers of both strains by more than 5-log10 in <24 h, comparative genomics and transcriptomics revealed differences in the genomes and overall regulation of genes between the two E. coli strains. A wider arsenal of genes related to oxidative stress, cellular repair and protective mechanisms were upregulated in E. coli PI-7. Subpopulations of E. coli PI-7 expressed genes related to dormancy and persister cell formation during the late decay phase, which may have accounted for its prolonged persistence. Upon prolonged solar irradiation, both E. coli strains displayed upregulation of genes related to horizontal gene transfer and antibiotic resistance. Virulence functions unique to E. coli PI-7 were also upregulated. Our findings collectively indicated that, whereas solar irradiation is able to reduce total cell numbers, viable E. coli remained and expressed genes that enable survival despite solar treatment. There remains a need for heightened levels of concern regarding risks arising from the dissemination of E. coli that may remain viable in wastewater after solar irradiation.

Inactivation and Gene Expression of a Virulent WastewaterEscherichia coliStrain and the Nonvirulent CommensalEscherichia coliDSM1103 Strain upon Solar Irradiation

KAUST Repository

Aljassim, Nada I.

2017-03-06

This study examined the decay kinetics and molecular responses of two Escherichia coli strains upon solar irradiation. The first is E. coli PI-7, a virulent and antibiotic-resistant strain that was isolated from wastewater and carries the emerging NDM-1 antibiotic resistance gene. The other strain, E. coli DSM1103, displayed lower virulence and antibiotic resistance than E. coli PI-7. In a buffer solution, E. coli PI-7 displayed a longer lag phase prior to decay and a longer half-life compared with E. coli DSM1103 (6.64 ± 0.63 h and 2.85 ± 0.46 min vs 1.33 ± 0.52 h and 2.04 ± 0.36 min). In wastewater, both E. coli strains decayed slower than they did in buffer. Although solar irradiation remained effective in reducing the numbers of both strains by more than 5-log10 in <24 h, comparative genomics and transcriptomics revealed differences in the genomes and overall regulation of genes between the two E. coli strains. A wider arsenal of genes related to oxidative stress, cellular repair and protective mechanisms were upregulated in E. coli PI-7. Subpopulations of E. coli PI-7 expressed genes related to dormancy and persister cell formation during the late decay phase, which may have accounted for its prolonged persistence. Upon prolonged solar irradiation, both E. coli strains displayed upregulation of genes related to horizontal gene transfer and antibiotic resistance. Virulence functions unique to E. coli PI-7 were also upregulated. Our findings collectively indicated that, whereas solar irradiation is able to reduce total cell numbers, viable E. coli remained and expressed genes that enable survival despite solar treatment. There remains a need for heightened levels of concern regarding risks arising from the dissemination of E. coli that may remain viable in wastewater after solar irradiation.

Treatment of inflammatory bowel disease associated E. coli with ciprofloxacin and E. coli Nissle in the streptomycin-treated mouse intestine

DEFF Research Database (Denmark)

Petersen, Andreas Munk; Schjørring, Susanne; Gerstrøm, Sarah Choi

2011-01-01

E. coli belonging to the phylogenetic group B2 are linked to Inflammatory Bowel Disease (IBD). Studies have shown that antimicrobials have some effect in the treatment of IBD, and it has been demonstrated that E. coli Nissle has prophylactic abilities comparable to 5-aminosalicylic acid (5-ASA......) therapy in ulcerative colitis. The objective of this study was to test if ciprofloxacin and/or E. coli Nissle could eradicate IBD associated E. coli in the streptomycin-treated mouse intestine....

(5'-32P)-8-azidoguanosine-3',5'-monophosphate. I. Synthesis and properties. II. Interaction with E. coli proteins

International Nuclear Information System (INIS)

Owens, J.R.

1983-01-01

Under certain conditions of nutritional deprivation, microorganisms produce the magic spot nucleotides guanosine-3'-diphosphate-5'-triphosphate(pppGpp) and the tetraphosphate ppGpp. The latter is known to be a pleiotypic effector, i.e. it inhibits (and sometimes stimulates) many biological processes including transcription, translation, and metabolic pathways. It is unknown whether pppGpp, ppGp, pGpp, and pGp, other members of this family of guanosine-3',5'-phosphates, also have regulatory properties. To begin to investigate this question, a radioactive photoaffinity analog of pGp was prepared: (5' 32 P)pN 3 Gp. The interaction of this photoprobe with E. coli sonicates and a purified protein (RNA polymerase) was examined. At physiological salt concentrations two proteins (RNA polymerase) was examined. At physiological salt concentrations two proteins of 86,000 and 65,000 daltons (p86 and p65) were primarily photolabeled. Competition studies with guanosine and adenosine nucleotides indicated (5 32 P)pN 3 Gp was labeling a ppGpp binding site on p86, and a pGp (or GMP) site on p65. ATP phosphorylation of p86 increased photoincorporation, while it decreased labeling of p65. The data also provide evidence of a different type of regulatory mechanism, i.e. phosphorylation modulates binding of an allosteric effector (ppGpp) to a protein(enzyme). Both ATP and GTP were found to phosphorylate the same proteins, although GTP was the preferred substrate in some cases

Lipocalin 2 is protective against E. coli pneumonia

DEFF Research Database (Denmark)

Wu, Hong; Santoni-Rugiu, Eric; Ralfkiaer, Elisabeth

2010-01-01

Lipocalin 2 is a bacteriostatic protein that binds the siderophore enterobactin, an iron-chelating molecule produced by Escherichia coli (E. coli) that is required for bacterial growth. Infection of the lungs by E. coli is rare despite a frequent exposure to this commensal bacterium. Lipocalin 2...... is an effector molecule of the innate immune system and could therefore play a role in hindering growth of E. coli in the lungs....

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.

Cloning and expression analysis of two dehydrodolichyl diphosphate synthase genes from Tripterygium wilfordii

Directory of Open Access Journals (Sweden)

Lin-Hui Gao

2018-01-01

Full Text Available Objective: To clone and investigate two dehydrodolichyl diphosphate synthase genes of Tripterygium wilfordii by bioinformatics and tissue expression analysis. Materials and Methods: According to the T. wifordii transcriptome database, specific primers were designed to clone the TwDHDDS1 and TwDHDDS2 genes via PCR. Based on the cloned sequences, protein structure prediction, multiple sequence alignment and phylogenetic tree construction were performed. The expression levels of the genes in different tissues of T. wilfordii were measured by real-time quantitative PCR. Results: The TwDHDDS1 gene encompassed a 873 bp open reading frame (ORF and encoded a protein of 290 amino acids. The calculated molecular weight of the translated protein was about 33.46 kDa, and the theoretical isoelectric point (pI was 8.67. The TwDHDDS2 encompassed a 768 bp ORF, encoding a protein of 255 amino acids with a calculated molecular weight of about 21.19 kDa, and a theoretical isoelectric point (pI of 7.72. Plant tissue expression analysis indicated that TwDHDDS1 and TwDHDDS2 both have relatively ubiquitous expression in all sampled organ tissues, but showed the highest transcription levels in the stems. Conclusions: The results of this study provide a basis for further functional studies of TwDHDDS1 and TwDHDDS2. Most importantly, these genes are promising genetic targets for the regulation of the biosynthetic pathways of important bioactive terpenoids such as triptolide.

Escherichia coli Probiotic Strain ED1a in Pigs Has a Limited Impact on the Gut Carriage of Extended-Spectrum-β-Lactamase-Producing E. coli

Science.gov (United States)

Mourand, G.; Paboeuf, F.; Fleury, M. A.; Jouy, E.; Bougeard, S.; Denamur, E.

2016-01-01

ABSTRACT Four trials were conducted to evaluate the impact of Escherichia coli probiotic strain ED1a administration to pigs on the gut carriage or survival in manure of extended-spectrum-β-lactamase-producing E. coli. Groups of pigs were orally inoculated with strain E. coli M63 carrying the blaCTX-M-1 gene (n = 84) or used as a control (n = 26). In the first two trials, 24 of 40 E. coli M63-inoculated pigs were given E. coli ED1a orally for 6 days starting 8 days after oral inoculation. In the third trial, 10 E. coli M63-inoculated pigs were given either E. coli ED1a or probiotic E. coli Nissle 1917 for 5 days. In the fourth trial, E. coli ED1a was given to a sow and its 12 piglets, and these 12 piglets plus 12 piglets that had not received E. coli ED1a were then inoculated with E. coli M63. Fecal shedding of cefotaxime-resistant Enterobacteriaceae (CTX-RE) was studied by culture, and blaCTX-M-1 genes were quantified by PCR. The persistence of CTX-RE in manure samples from inoculated pigs or manure samples inoculated in vitro with E. coli M63 with or without probiotics was studied. The results showed that E. coli M63 and ED1a were good gut colonizers. The reduction in the level of fecal excretion of CTX-RE in E. coli ED1a-treated pigs compared to that in nontreated pigs was usually less than 1 log10 CFU and was mainly observed during the probiotic administration period. The results obtained with E. coli Nissle 1917 did not differ significantly from those obtained with E. coli ED1a. CTX-RE survival did not differ significantly in manure samples with or without probiotic treatment. In conclusion, under our experimental conditions, E. coli ED1a and E. coli Nissle 1917 could not durably prevent CTX-RE colonization of the pig gut. PMID:27795372

Escherichia coli Probiotic Strain ED1a in Pigs Has a Limited Impact on the Gut Carriage of Extended-Spectrum-β-Lactamase-Producing E. coli.

Science.gov (United States)

Mourand, G; Paboeuf, F; Fleury, M A; Jouy, E; Bougeard, S; Denamur, E; Kempf, I

2017-01-01

Four trials were conducted to evaluate the impact of Escherichia coli probiotic strain ED1a administration to pigs on the gut carriage or survival in manure of extended-spectrum-β-lactamase-producing E. coli Groups of pigs were orally inoculated with strain E. coli M63 carrying the bla CTX-M-1 gene (n = 84) or used as a control (n = 26). In the first two trials, 24 of 40 E. coli M63-inoculated pigs were given E. coli ED1a orally for 6 days starting 8 days after oral inoculation. In the third trial, 10 E. coli M63-inoculated pigs were given either E. coli ED1a or probiotic E. coli Nissle 1917 for 5 days. In the fourth trial, E. coli ED1a was given to a sow and its 12 piglets, and these 12 piglets plus 12 piglets that had not received E. coli ED1a were then inoculated with E. coli M63. Fecal shedding of cefotaxime-resistant Enterobacteriaceae (CTX-RE) was studied by culture, and bla CTX-M-1 genes were quantified by PCR. The persistence of CTX-RE in manure samples from inoculated pigs or manure samples inoculated in vitro with E. coli M63 with or without probiotics was studied. The results showed that E. coli M63 and ED1a were good gut colonizers. The reduction in the level of fecal excretion of CTX-RE in E. coli ED1a-treated pigs compared to that in nontreated pigs was usually less than 1 log 10 CFU and was mainly observed during the probiotic administration period. The results obtained with E. coli Nissle 1917 did not differ significantly from those obtained with E. coli ED1a. CTX-RE survival did not differ significantly in manure samples with or without probiotic treatment. In conclusion, under our experimental conditions, E. coli ED1a and E. coli Nissle 1917 could not durably prevent CTX-RE colonization of the pig gut. Copyright © 2016 American Society for Microbiology.

Metabolic engineering for the high-yield production of isoprenoid-based C5 alcohols in E. coli

Science.gov (United States)

George, Kevin W.; Thompson, Mitchell G.; Kang, Aram; Baidoo, Edward; Wang, George; Chan, Leanne Jade G.; Adams, Paul D.; Petzold, Christopher J.; Keasling, Jay D.; Soon Lee, Taek

2015-01-01

Branched five carbon (C5) alcohols are attractive targets for microbial production due to their desirable fuel properties and importance as platform chemicals. In this study, we engineered a heterologous isoprenoid pathway in E. coli for the high-yield production of 3-methyl-3-buten-1-ol, 3-methyl-2-buten-1-ol, and 3-methyl-1-butanol, three C5 alcohols that serve as potential biofuels. We first constructed a pathway for 3-methyl-3-buten-1-ol, where metabolite profiling identified NudB, a promiscuous phosphatase, as a likely pathway bottleneck. We achieved a 60% increase in the yield of 3-methyl-3-buten-1-ol by engineering the Shine-Dalgarno sequence of nudB, which increased protein levels by 9-fold and reduced isopentenyl diphosphate (IPP) accumulation by 4-fold. To further optimize the pathway, we adjusted mevalonate kinase (MK) expression and investigated MK enzymes from alternative microbes such as Methanosarcina mazei. Next, we expressed a fusion protein of IPP isomerase and the phosphatase (Idi1~NudB) along with a reductase (NemA) to diversify production to 3-methyl-2-buten-1-ol and 3-methyl-1-butanol. Finally, we used an oleyl alcohol overlay to improve alcohol recovery, achieving final titers of 2.23 g/L of 3-methyl-3-buten-1-ol (~70% of pathway-dependent theoretical yield), 150 mg/L of 3-methyl-2-buten-1-ol, and 300 mg/L of 3-methyl-1-butanol. PMID:26052683

Enhanced target-specific signal detection using an Escherichia coli lysate in multiplex microbead immunoassays with E. coli-derived recombinant antigens.

Science.gov (United States)

Crestani, Sandra; Leitolis, Amanda; Lima, Lucianna Freitas Oliveira; Krieger, Marco A; Foti, Leonardo

2016-08-01

Diverse techniques have been developed to analyze antibody-mediated responses to infections. However, the most common tests, i.e., enzyme-linked immunosorbent assays, require separate reactions for each antigen and consequently necessitate large sample volumes. Luminex technology allows the detection of multiple antibodies in a single experiment, but nonspecific binding can impair the results. Therefore, we examined the use of Escherichia coli lysates to reduce nonspecific binding and improve the results of liquid microarrays based on Luminex technology. Anti-bacteria antibodies were detected in human serum samples, as evidenced by high median fluorescence intensity (MFI) in assays performed with paramagnetic microspheres coupled with E. coli lysates. Moreover, the addition of an E. coli lysate as a blocker reduced the nonspecific binding of antigens produced by E. coli in a concentration-dependent manner. Tris-HCl reduced MFI values in negative samples, but did not affect MFI for positive samples. For microspheres coupled with different antigens, an E. coli lysate blocker significantly improved the fluorescence signals from positive samples. The addition of Tris-HCl and the E. coli lysate induced antigen-specific differences in MFI. This combination of the E. coli lysate blocker and Tris-HCl yielded a statistically significant improvement in MFI in the assays for Chagas disease and hepatitis C virus samples. However, for the Treponema pallidum p47 antigen improvement in MFI was only observed for the preparation with the E. coli blocker at a concentration of 3%. In conclusion, the addition of an E. coli lysate and Tris-HCl to the microarray assay reduced the nonspecific binding of human anti-bacteria antibodies and, therefore, increased the specific MFI. Copyright © 2016 Elsevier B.V. All rights reserved.

The effect of cytidine-diphosphate choline (CDP-choline) on brain lipid changes during aging

International Nuclear Information System (INIS)

De Medio, G.E.; Trovarelli, G.; Piccinin, G.L.; Porcellati, G.

1984-01-01

Lipid synthesis has been tested in vivo in different brain areas of 12-month-old male rats. Cortex, striatum, brainstem, and subcortex of brain have been examined. The cerebellum was discarded. Mixtures of (2- 3 H)glycerol and (Me- 14 C)choline were injected into the lateral ventricle of the brain as lipid precursors, and their incorporation into total lipid, water-soluble intermediates and choline-containing phospholipids was examined 1 hr after isotope injection. In another series of experiments cytidine-5'-diphosphate choline (CDP-choline) was injected intraventricularly to the aged rats 10 min before sacrifice with a simultaneous injection, and radioactivity assays were performed as above. Distribution of radioactivity content of CDP-choline among brain areas 10 min after its administration showed a noticeable enrichment of the nucleotide and water-soluble-related compounds in the examined areas, but to a lesser degree in the cerebral cortex. The incorporation of labelled glycerol, which is severely depressed in aged rats in all four areas [Gaiti et al, 1982, 1983], was increased only in the cortex, and apparently decreased in the other areas. This last result is probably due to a dilution effect brought about by the administered cold CDP-choline upon the ( 14 C)-containing water-soluble metabolites. As a consequence, the ( 3 H)/( 14 C) ratio in total lipid and in isolated phosphatidylcholine and choline plasmalogen increased after CDP-choline treatment

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...

Temperature effects on the interaction mechanisms between the europium (III) and uranyl ions and zirconium diphosphate

International Nuclear Information System (INIS)

Finck, N.

2006-10-01

Temperature should remain higher than 25 C in the near field environment of a nuclear waste repository for thousands years. In this context, the aim of this work is to study the temperature influence on the interaction mechanisms between europium (III) and uranyl ions and zirconium diphosphate, as well as the influence of a complexing medium (nitrate) on the sorption of the lanthanide. The experimental definition of the equilibria was achieved by combining a structural investigation with the macroscopic sorption data. Surface complexes were characterized at all temperatures (25 C to 90 C) by TRLFS experiments carried out on dry and in situ samples using an oven. This characterization was completed by XPS experiments carried out at 25 C on samples prepared at 25 C and 90 C. The reaction constants (surface hydration and cations sorption) were obtained by simulating the experimental data with the constant capacitance surface complexation model. The reaction constants temperature dependency allowed one to characterize thermodynamically the different reactions by application of the van't Hoff relation. The validity of this law was tested by performing microcalorimetric measurements of the sorption heat for both cations. (author)

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.

Evaluation of Petrifilm™ Select E. coli Count Plate medium to discriminate antimicrobial resistant Escherichia coli

Directory of Open Access Journals (Sweden)

Jensen Lars

2008-09-01

Full Text Available Abstract Background Screening and enumeration of antimicrobial resistant Escherichia coli directly from samples is needed to identify emerging resistant clones and obtain quantitative data for risk assessment. Aim of this study was to evaluate the performance of 3M™ Petrifilm™ Select E. coli Count Plate (SEC plate supplemented with antimicrobials to discriminate antimicrobial-resistant and non-resistant E. coli. Method A range of E. coli isolates were tested by agar dilution method comparing the Minimal Inhibitory Concentration (MIC for eight antimicrobials obtained by Mueller-Hinton II agar, MacConkey agar and SEC plates. Kappa statistics was used to assess the levels of agreement when classifying strains as resistant, intermediate or susceptible. Results SEC plate showed that 74% of all strains agreed within ± 1 log2 dilution when comparing MICs with Mueller-Hinton II media. High agreement levels were found for gentamicin, ampicillin, chloramphenicol and cefotaxime, resulting in a kappa value of 0.9 and 100% agreement within ± 1 log2 dilution. Significant variances were observed for oxytetracycline and sulphamethoxazole. Further tests showed that the observed discrepancy in classification of susceptibility to oxytetracycline by the two media could be overcome when a plate-dependent breakpoint of 64 mg/L was used for SEC plates. For sulphamethoxazole, SEC plates provided unacceptably high MICs. Conclusion SEC plates showed good agreement with Mueller-Hinton II agar in MIC studies and can be used to screen and discriminate resistant E. coli for ampicillin, cephalothin, streptomycin, chloramphenicol, cefotaxime and gentamicin using CLSI standardized breakpoints, but not for sulphamethoxazole. SEC plates can also be used to discriminate oxytetracycline-resistant E. coli if a plate-dependent breakpoint value of 64 mg/L is used.

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...

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...

Glucose uptake regulation in E. coli by the small RNA SgrS: comparative analysis of E. coli K-12 (JM109 and MG1655 and E. coli B (BL21

Directory of Open Access Journals (Sweden)

Ng Weng-Ian

2010-09-01

Full Text Available Abstract Background The effect of high glucose concentration on the transcription levels of the small RNA SgrS and the messenger RNA ptsG, (encoding the glucose transporter IICBGlc, was studied in both E. coli K-12 (MG1655 and JM109 and E. coli B (BL21. It is known that the transcription level of sgrS increases when E. coli K-12 (MG1655 and JM109 is exposed to the non-metabolized glucose alpha methyl glucoside (αMG or when the bacteria with a defective glycolysis pathway is grown in presence of glucose. The increased level of sRNA SgrS reduces the level of the ptsG mRNA and consequently lowers the level of the glucose transporter IICBGlc. The suggested trigger for this action is the accumulation of the corresponding phospho-sugars. Results In the course of the described work, it was found that E. coli B (BL21 and E. coli K-12 (JM109 and MG1655 responded similarly to αMG: both strains increased SgrS transcription and reduced ptsG transcription. However, the two strains reacted differently to high glucose concentration (40 g/L. E. coli B (BL21 reacted by increasing sgrS transcription and reducing ptsG transcription while E. coli K-12 (JM109 and MG1655 did not respond to the high glucose concentration, and, therefore, transcription of sgrS was not detected and ptsG mRNA level was not affected. Conclusions The results suggest that E. coli B (BL21 tolerates high glucose concentration not only by its more efficient central carbon metabolism, but also by controlling the glucose transport into the cells regulated by the sRNA SgrS, which may suggest a way to control glucose consumption and increase its efficient utilization.

Endogenous E. coli endophthalmitis.

Science.gov (United States)

Shammas, H F

1977-01-01

A case of Escherichia coli septicemia with associated metastatic en dophthalmitis and endocarditis is presented. The ocular signs and symptoms were the initial manifestations of sepsis. Irreversible damage to the eye occurred in less than 24 hours. The pattern of metastatic bacterial endophthalmitis has changed since the introduction of potent antimicrobial agents, with an increased incidence of Gram-negative bacillemia. E. coli endophthalmitis carries a poor prognosis. Early diagnosis and systemic treatment will prevent the life-threatening complications of sepsis.

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.

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 ...

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.

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...

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 ...

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...

Temperature effects on the interaction mechanisms between the europium (III) and uranyl ions and zirconium diphosphate; Effets de la temperature sur les mecanismes d'interaction entre les ions europium (3) et uranyle et le diphosphate de zirconium

Energy Technology Data Exchange (ETDEWEB)

Finck, N

2006-10-15

Temperature should remain higher than 25 C in the near field environment of a nuclear waste repository for thousands years. In this context, the aim of this work is to study the temperature influence on the interaction mechanisms between europium (III) and uranyl ions and zirconium diphosphate, as well as the influence of a complexing medium (nitrate) on the sorption of the lanthanide. The experimental definition of the equilibria was achieved by combining a structural investigation with the macroscopic sorption data. Surface complexes were characterized at all temperatures (25 C to 90 C) by TRLFS experiments carried out on dry and in situ samples using an oven. This characterization was completed by XPS experiments carried out at 25 C on samples prepared at 25 C and 90 C. The reaction constants (surface hydration and cations sorption) were obtained by simulating the experimental data with the constant capacitance surface complexation model. The reaction constants temperature dependency allowed one to characterize thermodynamically the different reactions by application of the van't Hoff relation. The validity of this law was tested by performing microcalorimetric measurements of the sorption heat for both cations. (author)

Faecal Escherichia coli from patients with E. coli urinary tract infection and healthy controls who have never had a urinary tract infection

DEFF Research Database (Denmark)

Nielsen, Karen L; Dynesen, Pia; Larsen, Preben

2014-01-01

Urinary tract infections (UTIs) are primarily caused by Escherichia coli with the patient's own faecal flora acting as a reservoir for the infecting E. coli. Here we sought to characterize the E. coli faecal flora of UTI patients and healthy controls who had never had a UTI. Up to 20 E. coli...... colonies from each rectal swab were random amplified polymorphic DNA (RAPD) typed for clonality, dominance in the sample and correlation to the infecting UTI isolate in patients. Each distinct clone was phylotyped and tested for antimicrobial susceptibility. Eighty-seven per cent of the UTI patients...... carried the infecting strain in their faecal flora, and faecal clones causing UTI were more often dominant in the faecal flora. Patients had a larger diversity of E. coli in their gut flora by carrying more unique E. coli clones compared to controls, and patient faecal clones were more often associated...

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...

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

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...

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.

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.

Survival of pathogenic Escherichia coli on basil, lettuce, and spinach

Science.gov (United States)

The contamination of lettuce, spinach and basil with pathogenic E. coli has caused numerous illnesses over the past decade. E. coli O157:H7, E. coli O104:H4 and avian pathogenic E. coli (APECstx- and APECstx+) were inoculated on basil plants and in promix soiless substrate using drip and overhead ir...

Biphasic Elimination of Tenofovir Diphosphate and Nonlinear Pharmacokinetics of Zidovudine Triphosphate in a Microdosing Study

Science.gov (United States)

Chen, Jianmeng; Flexner, Charles; Liberman, Rosa G.; Skipper, Paul L.; Louissaint, Nicolette; Tannenbaum, Steven R.; Hendrix, Craig; Fuchs, Edward

2012-01-01

Objective Phase 0 studies can provide initial pharmacokinetics (PK) data in humans and help to facilitate early drug development, but their predictive value for standard dosing is controversial. To evaluate the prediction of microdosing for active intracellular drug metabolites, we compared the PK profile of two antiretroviral drugs, zidovudine (ZDV) and tenofovir (TFV), in microdose and standard dosing regimens. Study Design We administered a microdose (100 μg) of 14C-labeled drug (ZDV or tenofovir disoproxil fumarate (TDF)) with or without a standard unlabelled dose (300 mg) to healthy volunteers. Both the parent drug in plasma and the active metabolite, ZDV-triphosphate (ZDV-TP) or TFV-diphosphate (TFV-DP) in PBMCs and CD4+ cells were measured by AMS. Results The intracellular ZDV-TP concentration increased less than proportionally over the dose range studied (100 μg to 300 mg), while the intracellular TFV-DP PK were linear over the same dose range. ZDV-TP concentrations were lower in CD4+ cells versus total peripheral blood mononuclear cells (PBMCs), while TFV-DP concentrations were not different in CD4+ cells and PBMCs. Conclusion Our data were consistent with a rate-limiting step in the intracellular phosphorylation of ZDV but not TFV. AMS shows promise for predicting the PK of active intracellular metabolites of nucleosides, but nonlinearity of PK may be seen with some drugs. PMID:23187888

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.

The molecular origin of the thiamin diphosphate-induced spectral bands of ThDP-dependent enzymes

NARCIS (Netherlands)

Kovina, M.V.; Kok, A.; Sevostyanova, I.A.; Khailova, L.S.; Belkina, N.V.; Kochetov, G.A.

2004-01-01

New and previously published data on a variety of ThDP-dependent enzymes such as baker's yeast transketolase, yeast pyruvate decarboxylase and pyruvate dehydrogenase from pigeon breast muscle, bovine heart, bovine kidney, Neisseria meningitidis and E. coli show their spectral sensitivity to ThDP

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 ...

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.

A scoping study on the prevalence of Escherichia coli and ...

African Journals Online (AJOL)

In the Eastern Cape Province E. coli and enterococci were detected in 7 and 4 of the 15 RHRW tanks, respectively. Enterococci were detected only from one river whereas E. coli was detected in all three rivers; in spring water neither enterococci nor E. coli were detected. Samples from GRWH tanks were positive for E. coli ...

Asymptomatic bacteriuria Escherichia coli are live biotherapeutics for UTI.

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Rudick, Charles N; Taylor, Aisha K; Yaggie, Ryan E; Schaeffer, Anthony J; Klumpp, David J

2014-01-01

Urinary tract infections (UTI) account for approximately 8 million clinic visits annually with symptoms that include acute pelvic pain, dysuria, and irritative voiding. Empiric UTI management with antimicrobials is complicated by increasing antimicrobial resistance among uropathogens, but live biotherapeutics products (LBPs), such as asymptomatic bacteriuria (ASB) strains of E. coli, offer the potential to circumvent antimicrobial resistance. Here we evaluated ASB E. coli as LBPs, relative to ciprofloxacin, for efficacy against infection and visceral pain in a murine UTI model. Visceral pain was quantified as tactile allodynia of the pelvic region in response to mechanical stimulation with von Frey filaments. Whereas ciprofloxacin promoted clearance of uropathogenic E. coli (UPEC), it did not reduce pelvic tactile allodynia, a measure of visceral pain. In contrast, ASB E. coli administered intravesically or intravaginally provided comparable reduction of allodynia similar to intravesical lidocaine. Moreover, ASB E. coli were similarly effective against UTI allodynia induced by Proteus mirabilis, Enterococccus faecalis and Klebsiella pneumoniae. Therefore, ASB E. coli have anti-infective activity comparable to the current standard of care yet also provide superior analgesia. These studies suggest that ASB E. coli represent novel LBPs for UTI symptoms.

Modeling the inactivation of Escherichia coli 0157:H7 and uropathogenic E.coli in ground chicken by high pressure processing and thymol

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Disease causing Escherichia coli commonly found in meat and poultry include intestinal pathogenic E. coli (iPEC) as well as extraintestinal types such as the Uropathogenic E. coli (UPEC). In this study we compare the resistance of iPEC (O157:H7) to UPEC in chicken meat using High Pressure Processing...

Modeling the inactivatin of Escherichia coli 0157:H7 and uropathogenic E. coli in ground beef by high pressure processing and citral

Science.gov (United States)

Disease causing Escherichia coli commonly found in meat and poultry include intestinal pathogenic E. coli (iPEC) as well as extraintestinal types such as the Uropathogenic E. coli (UPEC). In this study we compared the resistance of iPEC (O157:H7) to UPEC in ground beef using High Pressure Processing...

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....

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.

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 ...

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 ...

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.

Non-Escherichia coli versus Escherichia coli community-acquired urinary tract infections in children hospitalized in a tertiary center: relative frequency, risk factors, antimicrobial resistance and outcome.

Science.gov (United States)

Marcus, Nir; Ashkenazi, Shai; Yaari, Arnon; Samra, Zmira; Livni, Gilat

2005-07-01

Currently hospitalization for children with urinary tract infections (UTIs) is reserved for severe or complicated cases. Changes may have taken place in the characteristics and causative uropathogens of hospital-treated community-acquired UTI. To study children hospitalized in a tertiary center with community-acquired UTI, compare Escherichia coli and non-E. coli UTI, define predictors for non-E. coli UTI and elucidate the appropriate therapeutic approach. A prospective clinical and laboratory study from 2001 through 2002 in a tertiary pediatric medical center. Patients were divided by results of the urine culture into E. coli and non-E. coli UTI groups, which were compared. Of 175 episodes of culture-proved UTI, 70 (40%) were caused by non-E. coli pathogens. Non-E. coli UTI was more commonly found in children who were male (P = 0.005), who had underlying renal abnormalities (P = 0.0085) and who had received antibiotic therapy in the prior month (P = 0.0009). Non-E. coli uropathogens were often resistant to antibiotics usually recommended for initial therapy for UTI, including cephalosporins and aminoglycosides; 19% were initially treated with inappropriate empiric intravenous antibiotics (compared with 2% for E. coli UTI, P = 0.0001), with a longer hospitalization. Current treatment routines are often inappropriate for hospitalized children with non-E. coli UTI, which is relatively common in this population. The defined risk factors associated with non-E. coli UTIs and its antimicrobial resistance patterns should be considered to improve empiric antibiotic therapy for these infections.

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.

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 ...

Insights into the evolution of pathogenicity of Escherichia coli from genomic analysis of intestinal E. coli of Marmota himalayana in Qinghai-Tibet plateau of China.

Science.gov (United States)

Lu, Shan; Jin, Dong; Wu, Shusheng; Yang, Jing; Lan, Ruiting; Bai, Xiangning; Liu, Sha; Meng, Qiong; Yuan, Xuejiao; Zhou, Juan; Pu, Ji; Chen, Qiang; Dai, Hang; Hu, Yuanyuan; Xiong, Yanwen; Ye, Changyun; Xu, Jianguo

2016-12-07

Escherichia coli is both of a widespread harmless gut commensal and a versatile pathogen of humans. Domestic animals are a well-known reservoir for pathogenic E. coli. However, studies of E. coli populations from wild animals that have been separated from human activities had been very limited. Here we obtained 580 isolates from intestinal contents of 116 wild Marmot Marmota himalayana from Qinghai-Tibet plateau, China, with five isolates per animal. We selected 125 (hereinafter referred to as strains) from the 580 isolates for genome sequencing, based on unique pulse field gel electrophoresis patterns and at least one isolate per animal. Whole genome sequence analysis revealed that all 125 strains carried at least one and the majority (79.2%) carried multiple virulence genes based on the analysis of 22 selected virulence genes. In particular, the majority of the strains carried virulence genes from different pathovars as potential 'hybrid pathogens'. The alleles of eight virulence genes from the Marmot E. coli were found to have diverged earlier than all known alleles from human and other animal E. coli. Phylogenetic analysis of the 125 Marmot E. coli genomes and 355 genomes selected from 1622 human and other E. coli strains identified two new phylogroups, G and H, both of which diverged earlier than the other phylogroups. Eight of the 12 well-known pathogenic E. coli lineages were found to share a most recent common ancestor with one or more Marmot E. coli strains. Our results suggested that the intestinal E. coli of the Marmots contained a diverse virulence gene pool and is potentially pathogenic to humans. These findings provided a new understanding of the evolutionary origin of pathogenic E. coli.

Ischemic colitis or melanosis coli: a case report

Directory of Open Access Journals (Sweden)

Nadeem Mohammed

2007-09-01

Full Text Available Abstract Background Melanosis Coli is described as black or brown discolouration of the mucosa of the colon. Its a benign condition, which arises from anthraquinone laxative abuse and has no symptoms of its own. The main importance of diagnosing Melanosis Coli correctly lies in the fact that if its extensive, there may be difficulty in differentiating it from ischemic colitis. Case presentation We present a case of extensive Melanosis Coli involving the whole of large bowel that appeared gangrenous. A sub total colectomy was performed on presumed diagnosis of ischemic bowel. Conclusion This report reminds the clinicians that extensive Melanosis Coli may mimic ischemic colitis and thus must be considered as a differential diagnosis.

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…

Presence of non-O157 Shiga toxin-producing Escherichia coli, enterotoxigenic E. coli, enteropathogenic E. coli and Salmonella in fresh beetroot (Beta vulgaris L.) juice from public markets in Mexico.

Science.gov (United States)

Gómez-Aldapa, Carlos A; Rangel-Vargas, Esmeralda; Bautista-De León, Haydee; Castro-Rosas, Javier

2014-10-01

Unpasteurized juice has been associated with foodborne illness outbreaks for many years. Beetroot is a vegetable grown all over the world in temperate areas. In Mexico beetroot is consumed cooked in salads or raw as fresh unpasteurized juices. No data about the microbiological quality or safety of unpasteurized beetroot juices are available. Indicator bacteria, diarrheagenic Escherichia coli pathotypes (DEP) and Salmonella frequencies were determined for fresh unpasteurized beetroot juice from restaurants. One hundred unpasteurized beetroot juice samples were collected from public markets in Pachuca, Mexico. Frequencies in these samples were 100%, 75%, 53%, 9% and 4% of positive samples, for coliform bacteria, fecal coliforms, E. coli, DEP and Salmonella, respectively. Identified DEP included enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC) and non-O157 Shiga toxin-producing E. coli (STEC). Identified Salmonella serotypes included Typhimurium and Enteritidis. This is the first report of microbiological quality and atypical EPEC, ETEC, non-O157 STEC and Salmonella isolation from fresh raw beetroot juice in Mexico. Fresh raw beetroot juice from markets is very probably an important factor contributing to the endemicity of atypical EPEC, ETEC, non-O157 STEC and Salmonella-related gastroenteritis in Mexico. © 2014 Society of Chemical Industry.

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...

Environmental Escherichia coli: ecology and public health implications-a review.

Science.gov (United States)

Jang, J; Hur, H-G; Sadowsky, M J; Byappanahalli, M N; Yan, T; Ishii, S

2017-09-01

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 faeces or wastewater effluent. The presence of E. coli in environmental waters has long been considered as an indicator of recent faecal 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 extraintestinal 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 faecal 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 revealed the diversity and complexity of E. coli strains in various environments, which are affected by multiple environmental factors. This review examines the current knowledge on the ecology of E. coli strains in various environments with regard 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. © 2017 The Society for Applied Microbiology.

Fecal colonization with P-fimbriated Escherichia coli in newborn children and relation to development of extraintestinal E. coli infections.

Science.gov (United States)

Tullus, K

1987-01-01

The incidence of E. coli pyelonephritis before the age of one year among the children born at Danderyd Hospital during a ten year period was studied. During the study period, 4 or 5 outbreaks of E. coli pyelonephritis occurred among the children who had previously been staying in the hospital's neonatal ward. These outbreaks seemed to have been caused by nosocomial spread of and fecal colonization with certain virulent E. coli strains among the children staying in the ward during certain periods of time. The strains that were spread in the ward seemed to belong to certain pyelonephritogenic E. coli clones of the serotypes O6:K5, O4:K3 and possibly O6:K2. Although the children became fecally colonized with the strains in the neonatal ward, most fell ill some time after they had left the ward. The mean age at the development of their first pyelonephritis was 3.4 months for the boys and 6.2 months for the girls, who had been cared for in this ward. A correlation between the number of infections and the bed occupancy of the ward could be found (p less than 0.01). The risk for a child staying in the ward during an outbreak to develop pyelonephritis was about 5-10%. There was a baseline incidence rate of 0.6-0.7% during non-epidemic periods. During one of the outbreaks there was also an increased incidence rate of E. coli septicemia among the children staying in the neonatal ward. The predictive value of fecal colonization with P-fimbriated E. coli for the later development of extraintestinal E. coli infections was studied in a 2.5 year prospective study. During this study period there was a baseline incidence rate of 10-20% fecal colonization with P-fimbriated E. coli among the children staying in both the neonatal and maternity wards, interrupted only by minor peaks of colonization with such strains. Length of stay in the neonatal ward and a high bed occupancy of the neonatal ward were statistically correlated to fecal colonization with P-fimbriated E. coli strains (p

Ontology-based literature mining of E. coli vaccine-associated gene interaction networks.

Science.gov (United States)

Hur, Junguk; Özgür, Arzucan; He, Yongqun

2017-03-14

Pathogenic Escherichia coli infections cause various diseases in humans and many animal species. However, with extensive E. coli vaccine research, we are still unable to fully protect ourselves against E. coli infections. To more rational development of effective and safe E. coli vaccine, it is important to better understand E. coli vaccine-associated gene interaction networks. In this study, we first extended the Vaccine Ontology (VO) to semantically represent various E. coli vaccines and genes used in the vaccine development. We also normalized E. coli gene names compiled from the annotations of various E. coli strains using a pan-genome-based annotation strategy. The Interaction Network Ontology (INO) includes a hierarchy of various interaction-related keywords useful for literature mining. Using VO, INO, and normalized E. coli gene names, we applied an ontology-based SciMiner literature mining strategy to mine all PubMed abstracts and retrieve E. coli vaccine-associated E. coli gene interactions. Four centrality metrics (i.e., degree, eigenvector, closeness, and betweenness) were calculated for identifying highly ranked genes and interaction types. Using vaccine-related PubMed abstracts, our study identified 11,350 sentences that contain 88 unique INO interactions types and 1,781 unique E. coli genes. Each sentence contained at least one interaction type and two unique E. coli genes. An E. coli gene interaction network of genes and INO interaction types was created. From this big network, a sub-network consisting of 5 E. coli vaccine genes, including carA, carB, fimH, fepA, and vat, and 62 other E. coli genes, and 25 INO interaction types was identified. While many interaction types represent direct interactions between two indicated genes, our study has also shown that many of these retrieved interaction types are indirect in that the two genes participated in the specified interaction process in a required but indirect process. Our centrality analysis of

Melanosis coli in patients with colon cancer

Directory of Open Access Journals (Sweden)

Dorota Biernacka-Wawrzonek

2016-12-01

Full Text Available Intoduction: Melanosis coli is a benign lesion affecting the mucosa of the large intestine. There is a relationship between the presence of melanosis and anthraquinone laxative use. Melanosis coli is also observed in patients with colon cancer, but there is doubt whether these two conditions are related. Aim : To analyze the correlation between melanosis and colon cancer. Material and methods: We analyzed retrospectively 436 patients undergoing colon cancer surgery. There were 246 women and 190 men. Patients were divided into three age groups: under 50 years, between 51 and 65 years, and over 66 years. We analyzed sections of the cancer and intestinal mucosa from the tumor’s proximal (2–5 cm and distal (8–10 cm zone. Results : Melanosis coli was present in 52 patients, which represents 11.9% of patients with colon cancer. More often it was present in women. The most common location of melanosis and colon cancer was the terminal part of the large intestine. In patients below 50 years of age in both sexes melanosis coli did not occur. In men, melanosis was more common in the age group over 66 years. Intensity of pigmentation was higher in the tumor’s distal zone. Conclusions : The incidence of melanosis coli increases with age, similar to that of colon cancer. Melanosis was not present inside tumors, in almost half of the cases it was not present in the proximal zone, and the degree of pigmentation increased in distal zone. The cause-effect relationship between melanosis coli and colon cancer remains uncertain.

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

The role of Cra in regulating acetate excretion and osmotic tolerance in E. coli K-12 and E. coli B at high density growth.

Science.gov (United States)

Son, Young-Jin; Phue, Je-Nie; Trinh, Loc B; Lee, Sang Jun; Shiloach, Joseph

2011-06-30

E. coli B (BL21), unlike E.coli K-12 (JM109) is insensitive to glucose concentration and, therefore, grows faster and produces less acetate than E. coli K-12, especially when growing to high cell densities at high glucose concentration. By performing genomic analysis, it was demonstrated that the cause of this difference in sensitivity to the glucose concentration is the result of the differences in the central carbon metabolism activity. We hypothesized that the global transcription regulator Cra (FruR) is constitutively expressed in E. coli B and may be responsible for the different behaviour of the two strains. To investigate this possibility and better understand the function of Cra in the two strains, cra - negative E. coli B (BL21) and E. coli K-12 (JM109) were prepared and their growth behaviour and gene expression at high glucose were evaluated using microarray and real-time PCR. The deletion of the cra gene in E. coli B (BL21) minimally affected the growth and maximal acetate accumulation, while the deletion of the same gene in E.coli K-12 (JM109) caused the cells to stop growing as soon as acetate concentration reached 6.6 g/L and the media conductivity reached 21 mS/cm. ppsA (gluconeogenesis gene), aceBA (the glyoxylate shunt genes) and poxB (the acetate producing gene) were down-regulated in both strains, while acs (acetate uptake gene) was down-regulated only in E.coli B (BL21). These transcriptional differences had little effect on acetate and pyruvate production. Additionally, it was found that the lower growth of E. coli K-12 (JM109) strain was the result of transcription inhibition of the osmoprotectant producing bet operon (betABT). The transcriptional changes caused by the deletion of cra gene did not affect the activity of the central carbon metabolism, suggesting that Cra does not act alone; rather it interacts with other pleiotropic regulators to create a network of metabolic effects. An unexpected outcome of this work is the finding that cra

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...

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).

Can Escherichia coli fly? The role of flies as transmitters of E. coli to food in an urban slum in Bangladesh.

Science.gov (United States)

Lindeberg, Yrja Lisa; Egedal, Karen; Hossain, Zenat Zebin; Phelps, Matthew; Tulsiani, Suhella; Farhana, Israt; Begum, Anowara; Jensen, Peter Kjaer Mackie

2018-01-01

To investigate the transmission of faecal bacteria by flies to food under natural settings. Over a period of 2 months, paired (exposed and non-exposed) containers with cooked rice were placed on the ground in kitchen areas in an urban slum area in Dhaka, Bangladesh, and the numbers of flies landing on the exposed rice were counted. Following exposure, the surface of the rice was microbiologically and molecularly analysed for the presence of Escherichia coli and genes of diarrhoeagenic E. coli and Shigella strains. Rice was at greater risk (P E. coli if flies landed on the rice than if no flies landed on the rice (odds ratio 5·4 (P 0·6 × 103 CFU. Genes of diarrhoeagenic E. coli and Shigella species were detected in 39 of 60 (65%) of exposed rice samples. Two fly species were identified: the common housefly (Musca domestica) and the oriental latrine fly (Chrysomya megacephala). Flies may transmit large quantities of E. coli to food under field settings. The findings highlight the importance of implementing control measures to minimise exposure of food to flies to ensure food safety. Fly control measures should be considered for the prevention of diarrhoeal diseases caused by E. coli. © 2017 John Wiley & Sons Ltd.

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)

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.

High temperature in combination with UV irradiation enhances horizontal transfer of stx2 gene from E. coli O157:H7 to non-pathogenic E. coli.

Directory of Open Access Journals (Sweden)

Wan-Fu Yue

Full Text Available Shiga toxin (stx genes have been transferred to numerous bacteria, one of which is E. coli O157:H7. It is a common belief that stx gene is transferred by bacteriophages, because stx genes are located on lambdoid prophages in the E. coli O157:H7 genome. Both E. coli O157:H7 and non-pathogenic E. coli are highly enriched in cattle feedlots. We hypothesized that strong UV radiation in combination with high temperature accelerates stx gene transfer into non-pathogenic E. coli in feedlots.E. coli O157:H7 EDL933 strain were subjected to different UV irradiation (0 or 0.5 kJ/m(2 combination with different temperature (22, 28, 30, 32, and 37 °C treatments, and the activation of lambdoid prophages was analyzed by plaque forming unit while induction of Stx2 prophages was quantified by quantitative real-time PCR. Data showed that lambdoid prophages in E. coli O157:H7, including phages carrying stx2, were activated under UV radiation, a process enhanced by elevated temperature. Consistently, western blotting analysis indicated that the production of Shiga toxin 2 was also dramatically increased by UV irradiation and high temperature. In situ colony hybridization screening indicated that these activated Stx2 prophages were capable of converting laboratory strain of E. coli K12 into new Shiga toxigenic E. coli, which were further confirmed by PCR and ELISA analysis.These data implicate that high environmental temperature in combination with UV irradiation accelerates the spread of stx genes through enhancing Stx prophage induction and Stx phage mediated gene transfer. Cattle feedlot sludge are teemed with E. coli O157:H7 and non-pathogenic E. coli, and is frequently exposed to UV radiation via sunlight, which may contribute to the rapid spread of stx gene to non-pathogenic E. coli and diversity of shiga toxin producing E. coli.

The presence of verotoxinogenic E. coli in some foods

Directory of Open Access Journals (Sweden)

Bostan K.

2005-01-01

Full Text Available In this study 30 samples each of ready-to-cook meatballs and white cheese as well as 96 samples of various ready-to-eat foods, obtained from different sales outlets in Istanbul, were analyzed for the presence of verotoxins (consequently vemtoxigenic E. coli E. coli with the aid of the enzyme immunoassay technique. Additionally, total coli-form and chromogenic E. coli count were determined by cultural methods for all food samples. E. coli growth was detected in all ready-to-cook meatballs (100%, in 27 of the white cheese samples (90% and in 69 of the other ready-to-eat food samples (71.9%. Verotoxins, however, could not be detected in any of the samples examined with the aid of the ELISA technique. The findings of this study indicate a low microbiological quality of the analyzed meatball, white cheese and ready-to-eat food samples; a considerable part of them did not conform to legal standards. However, within the sensitivity limits of the method applied no verotoxinogenic E. coli could be detected.

Lethality prediction for Escherichia coli 0157:H7 and Uropathogenic E. coli in ground chicken treated with high pressure processing and trans-cinnamaldehyde

Science.gov (United States)

Pathogenic Escherichia coli, intestinal (O157:H7) as well as extraintestinal types (Uropathogenic E. coli (UPEC)) are commonly found in many foods including chicken meat. In this study we compared the resistance of E. coli O157:H7 to UPEC in chicken meat under the stresses of high hydrostatic pressu...

Molecular epidemiology of extended-spectrum beta-lactamase-producing Escherichia coli

Directory of Open Access Journals (Sweden)

Catherine Ludden

2014-09-01

Full Text Available Objectives: E. coli O25b-ST131 has disseminated worldwide in hospitals and the community. The objective of this study was to determine the extent to which E. coli O25b-ST131 accounts for extended-spectrum beta-lactamase (ESBLproducing E. coli from clinical samples from all sources in this region. Methods: Between January and June 2010 ESBL-producing E. coli were collected from 94 routine samples including 47 from residents of 25 nursing homes, 15 categorized as hospital acquired and 32 others. PCR was performed for detection of bla CTX-M, bla OXA-1, bla TEM, bla SHV and for the identification of members of the E. coli O25b:ST131 clonal group. PFGE was carried out using Xba I in accordance with PulseNet protocols. Results: The majority (97% of isolates harbored a bla CTX-M gene.E. coli O25b-ST131 accounted for 87% of all ESBL-producing E. coliand for 96% of isolates from nursing home residents. Conclusion:The E. coli O25b-ST131 clonal group predominated in the collection of ESBL-producing E. coli, particularly in nursing home isolates. J Microbiol Infect Dis 2014; 4(3: 92-96

Third International E. coli genome meeting

Energy Technology Data Exchange (ETDEWEB)

NONE

1994-12-31

Proceedings of the Third E. Coli Genome Meeting are provided. Presentations were divided into sessions entitled (1) Large Scale Sequencing, Sequence Analysis; (2) Databases; (3) Sequence Analysis; (4) Sequence Divergence in E. coli Strains; (5) Repeated Sequences and Regulatory Motifs; (6) Mutations, Rearrangements and Stress Responses; and (7) Origins of New Genes. The document provides a collection of abstracts of oral and poster presentations.

Impact of persistent and nonpersistent generic Escherichia coli and Salmonella sp. recovered from a beef packing plant on biofilm formation by E. coli O157.

Science.gov (United States)

Visvalingam, J; Ells, T C; Yang, X

2017-12-01

To examine the influence of meat plant Escherichia coli and Salmonella sp. isolates on E. coli O157 biofilm formation. Biofilm formation was quantified by crystal violet staining (A 570 nm ) and viable cell numbers for up to 6 days at 15°C. All five persistent E. coli genotypes formed strong biofilms when cultured alone or co-cultured with E. coli O157, with A 570 nm values reaching ≥4·8 at day 4, while only two of five nonpersistent genotypes formed such biofilms. For E. coli O157:H7 co-culture biofilms with E. coli genotypes 136 and 533, its numbers were ≥1·5 and ≥1 log CFU per peg lower than those observed for its mono-culture biofilm at days 2 and 4, respectively. The number of E. coli O157:NM in similar co-culture biofilms was 1 log CFU per peg lower than in its mono-culture biofilm at day 4 and 6, respectively. Salmonella sp. lowered the number of E. coli O157:NM by 0·5 log unit, once, at day 6. Generic E. coli may outcompete E. coli O157 strains while establishing biofilms. Findings advance knowledge regarding inter-strain competition for a similar ecological niche and may aid development of biocontrol strategies for E. coli O157 in food processing environments. © 2017 Her Majesty the Queen in Right of Canada. Journal of Applied Microbiology © 2017 The Society for Applied Microbiology Reproduced with the permission of the Minister of the Department of Agriculture and Agri-Food Canada.

Protection against an infectious disease by enterohaemorrhagic E. coli 0-157.

Science.gov (United States)

Ota, A

1999-07-01

Preventive measures against infection by enterohaemorrhagic E. coli 0-157 are described. Eating yoghurt and Kefir supposedly induces more bifid bacteria and lactic acid bacteria to colonize in the intestines, thereby protecting humans from infection by E. coli 0-157. Some foods, such as plum extract, act as a mild antibiotic and produce an acidic environment within the intestine, thus interfering with growth of the E. coli 0-157. The natural colonization of harmless E. coli or other bacteria that are more powerful than E. coli 0-157 can possibly protect against infection. A vaccination against E. coli 0-157 H7 may also be effective. In addition, it has been suggested that the correct levels of nitric oxide and calcium in the blood may activate immunity and protect against infection by E. coli 0-157.

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.

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.

Enteroaggregative Escherichia coli is the predominant diarrheagenic E. coli pathotype among irrigation water and food sources in South Africa.

Science.gov (United States)

Aijuka, Matthew; Santiago, Araceli E; Girón, Jorge A; Nataro, James P; Buys, Elna M

2018-08-02

Diarrheagenic E. coli (DEC) has been implicated in foodborne outbreaks worldwide and have been associated with childhood stunting in the absence of diarrhoea. Infection is extraordinarily common, but the routes of transmission have not been determined. Therefore, determining the most prevalent pathotypes in food and environmental sources may help provide better guidance to various stakeholders in ensuring food safety and public health and advancing understanding of the epidemiology of enteric disease. We characterized 205 E. coli strains previously isolated from producer distributor bulk milk (PDBM)(118), irrigation water (48), irrigated lettuce (29) and street vendor coleslaw (10) in South Africa. Enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroaggregative E. coli (EAEC) and diffusely adherent E. coli (DAEC) were sought. We used PCR and partial gene sequencing for all 205 strains while 46 out of 205 that showed poor resolution were subsequently characterized using cell adherence (HeLa cells). PCR and partial gene sequencing of aatA and/or aaiC genes confirmed EAEC (2%, 5 out of 205) as the only pathotype. Phylogenetic analysis of sequenced EAEC strains with E. coli strains in GenBank showing ≥80% nucleotide sequence similarity based on possession of aaiC and aatA generated distinct clusters of strains separated predominantly based on their source of isolation (food source or human stool) suggesting a potential role of virulence genes in source tracking. EAEC 24%, 11 out of 46 strains (PDBM = 15%, irrigation water = 7%, irrigated lettuce = 2%) was similarly the predominant pathotype followed by strains showing invasiveness to HeLa cells, 4%, 2 out of 46 (PDBM = 2%, irrigated lettuce = 2%), among stains characterized using cell adherence. Therefore, EAEC may be the leading cause of DEC associated food and water-borne enteric infection in South Africa. Additionally, solely using molecular based methods targeting virulence

Fluoroquinolone-resistant Escherichia coli carriage in long-term care facility.

Science.gov (United States)

Maslow, Joel N; Lee, Betsy; Lautenbach, Ebbing

2005-06-01

We conducted a cross-sectional study to determine the prevalence of, and risk factors for, colonization with fluoroquinolone (FQ)-resistant Escherichia coli in residents in a long-term care facility. FQ-resistant E. coli were identified from rectal swabs for 25 (51%) of 49 participants at study entry. On multivariable analyses, prior FQ use was the only independent risk factor for FQ-resistant E. coli carriage and was consistent for FQ exposures in the previous 3, 6, 9, or 12 months. Pulsed-field gel electrophoresis of FQ-resistant E. coli identified clonal spread of 1 strain among 16 residents. Loss (6 residents) or acquisition (7 residents) of FQ-resistant E. coli was documented and was associated with de novo colonization with genetically distinct strains. Unlike the case in the hospital setting, FQ-resistant E. coli carriage in long-term care facilities is associated with clonal spread.

Expression in E. coli systems

DEFF Research Database (Denmark)

Krogsdam, Anne-M; Kristiansen, Karsten; Nøhr, Jane

2003-01-01

intracellularly in soluble form. In E. coli, proteins containing disulfide bonds are best produced by secretion because the disulfide forming foldases reside in the periplasm. Likewise, a correct N-terminus is more likely to be obtained upon secretion. Moreover, potentially toxic proteins are more likely......Owing to cost advantage, speed of production, and often high product yield (up to 50% of total cell protein), expression in Escherichia coli is generally the first choice when attempting to express a recombinant protein. Expression systems exist to produce recombinant protein intracellularly...

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…

Triglyceride kinetics in fasted and fed E.coli septic rats

International Nuclear Information System (INIS)

Lanza-Jacoby, S.; Tabares, A.

1990-01-01

The mechanism for the development of hypertriglyceridemia during gram-negative sepsis was studies by examining the liver production and clearance of very-low-density lipoprotein (VLDL) triglyceride (TG). To assess the liver output and peripheral clearance the kinetics of VLDL-TG were determined by a constant intravenous infusion of [2- 3 H] glycerol-labeled VLDL in fasted control, fasted E. coli-treated, fed control, and fed E.coli-treated rats. Lewis inbred rats, 275-300 g, were made septic with 8 x 10 7 live E.coli colonies per 100 g body weight. Twenty-four hours following E.coli injection serum TG of fasted E.coli-treated rats was elevated by 170% which was attributed to a 67% decrease in the clearance rate of VLDL-TG in fasted E.coli-treated rats compared with their fasted controls. The secretion of VLDL-TG declined by 31% in the livers of the fasted E.coli-treated rats which was accompanied by a 2-fold increase in the composition of liver TG. In a second series of experiments control and E.coli-treated rats were fed intragastrically (IG) a balanced solution containing glucose plus fat as the sources of nonprotein calories. Serum TG were 26% lower in the fed E.coli-treated rats because the clearance rate increased by 86%. The secretion of TG in the fed septic rats increased by 40% but this difference was not significant. In the septic rat the ability to clear triglycerides from the plasma depends upon the nutritional state

ECMDB: The E. coli Metabolome Database

OpenAIRE

Guo, An Chi; Jewison, Timothy; Wilson, Michael; Liu, Yifeng; Knox, Craig; Djoumbou, Yannick; Lo, Patrick; Mandal, Rupasri; Krishnamurthy, Ram; Wishart, David S.

2012-01-01

The Escherichia coli Metabolome Database (ECMDB, http://www.ecmdb.ca) is a comprehensively annotated metabolomic database containing detailed information about the metabolome of E. coli (K-12). Modelled closely on the Human and Yeast Metabolome Databases, the ECMDB contains >2600 metabolites with links to ?1500 different genes and proteins, including enzymes and transporters. The information in the ECMDB has been collected from dozens of textbooks, journal articles and electronic databases. E...

Metabolic Modeling of Common Escherichia coli Strains in Human Gut Microbiome

Directory of Open Access Journals (Sweden)

Yue-Dong Gao

2014-01-01

Full Text Available The recent high-throughput sequencing has enabled the composition of Escherichia coli strains in the human microbial community to be profiled en masse. However, there are two challenges to address: (1 exploring the genetic differences between E. coli strains in human gut and (2 dynamic responses of E. coli to diverse stress conditions. As a result, we investigated the E. coli strains in human gut microbiome using deep sequencing data and reconstructed genome-wide metabolic networks for the three most common E. coli strains, including E. coli HS, UTI89, and CFT073. The metabolic models show obvious strain-specific characteristics, both in network contents and in behaviors. We predicted optimal biomass production for three models on four different carbon sources (acetate, ethanol, glucose, and succinate and found that these stress-associated genes were involved in host-microbial interactions and increased in human obesity. Besides, it shows that the growth rates are similar among the models, but the flux distributions are different, even in E. coli core reactions. The correlations between human diabetes-associated metabolic reactions in the E. coli models were also predicted. The study provides a systems perspective on E. coli strains in human gut microbiome and will be helpful in integrating diverse data sources in the following study.

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 ...

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.

Anti E. coli Activity of Herbal Medicines: a Review

Directory of Open Access Journals (Sweden)

Nahid Rezaei

2017-12-01

Full Text Available Escherichia coli is the gram negative bacilli of Entrobacteriaceae family that commonly found in intestinal infections and many infections outside the intestine, like urinary tract infections (UTI, cholecystitis, wound infections, meningitis, septicemia, pulmonary infections, and many more. Plants are rich sources of bioactive compounds, hence they can be effective in a wide variety of diseases. The pandemic spread of multidrug-resistant (MDR bacteria (i.e., extended-spectrum b-lactamase-producing Enterobacteriaceae (ESBLPE, Carbapenemase-producing Enterobacteriaceae (CPE threaten healthcare Worldwide. The present review is a report of the most effective medicinal plants against E. coli. In this research, the required online database searches were conducted using the key words such as bacteria, E. coli and medicinal plants. Databases of Web of Science, PubMed, Scopus, Google Scholar, and ScienceDirect were explored to find and explore related articles. Since the incidence of E. coli is high, the aim of this study is to identify and report anti E. coli medicinal plants in Iran. The obtained results showed that there were 51 medicinal herbs that could be considered as the main medicinal plants capable of affecting E. coli.

Biochemical and serological characterization of Escherichia coli ...

African Journals Online (AJOL)

This study was designed to determine the isolation rate, serotypes and biochemical profiles of E. coli from colibacillosis and dead-in-shell embryos in Zaria, Northern-Nigeria. The isolation rate of E. coli from hatcheries studied were 4.67% and 7.50% from farms of Simtu Agricultural Company and National Animal Production ...

Cytolysin a expressing E. coli a promising candidate for imageable therapeutic probe

International Nuclear Information System (INIS)

Nguyen, Vu Hong; Phan, Thuy Xuan; Hong, Yeoung Jin; Min, Jung Joon

2007-01-01

Using bacteria for cancer treatment has a long history. Discovery of optical reporter genes consisting of fluorescent and luminescent protein facilitates the monitor of bacteria in vivo, non-invasively and repeatedly. E. coli, the natural enteric bacteria possessing capacity of tumor-targeting ability, seems to be suitable candidate for cancer treatment. In this study, we established the strain light-emitting E. coli for diagnostic purpose and Cytolysin A (Cly A) expressing E. coli for therapeutic purpose. E. coli (MG1655, wild type strain) was transformed plasmid pUC19 carrying lux gene to create the light expressing bacteria and test the tumor targeting-capacity by injecting the bacteria into CT26-tumor bearing mice via tail vein. On the other hand, for therapeutic purpose, plasmid containing Cly A gene, which is encoded for a pore-forming protein toxin, was introduced into E. coli. The toxicity of Cly A was evaluated in vitro by inoculating the bacteria with various cultured cancer cell lines. On the other hand, to test the therapeutic effect, the bacteria were injected intratumorally and intravenously into s.c.CT26-bearing as well as CT26-lung metastasized Balb/c mice. In vivo imaging data showed that the E. coli strains selectively located in the tumor. The in vitro result showed that the number of death cells were significantly higher in the samples containing E. coli expressing Cly A (E. coli Cly A) compared with the samples containing wild type strain. The growth of tumors was repressed in mice injected with either E. coli Cly A (significantly) or wild type E. coli (mildly), while tumors in no treatment group still grew fast. Furthermore, the tumors inoculated with E. coli cly A were necrotized but not with wild type E. coli. In the CT26-lung metastasized mouse model, the life span of mice was elongated when inject E. coli and longer in the group injected with E. coli cly A. Cly A expressing E. coli can become an effective candidate for imageable

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

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.

E.coli O157:H7

Directory of Open Access Journals (Sweden)

Treor Francis Fernandez

2008-06-01

Full Text Available The serious nature of the symptoms of haemorrhagic colitis and HUS and the apparent low infectious dose (<100 cells of E.coli O157:H7 places this food borne pathogen a most serious of known food borne pathogens. Persuasive evidence suggests that healthy cattle are a reservoir of O157 and they can enter the food chain to provide a source of exposure for humans. A possible route of transmission of O157 VTEC may involve infections initially in calves that shed their organism into faecal slurry that may be used on grazing grass. This provides potential for infection of other animals from which the organism may contaminate milk or carcasses at slaughter. Possible sources of VTEC in healthy animals other E.coli O157:H7 than cattle and a wider range of foodstuffs require further investigation. Many features of E.coli O157:H7 strains remain poorly understood. It includes: (i Role of virulent genes in the animal, (ii Mechanism of evolution of the organism, (iii The progress of individual cases of E.coli O157:H7 infection, and (iv The difference in incidence of infection in different geographical areas. [Veterinary World 2008; 1(3.000: 83-87

Brote causado por Escherichia coli en Chalco, México Outbreak caused by Escherichia coli in Chalco, México

Directory of Open Access Journals (Sweden)

Iliana Alejandra Cortés-Ortiz

2002-07-01

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.Objective. To identify the etiologic agent responsible for a disease outbreak following an overflow of sewage water in Valle de Chalco, Mexico. Material and Methods. A retrospective cross-sectional study was carried out. Rectal samples were collected from the population of Chalco valley, who suffered from diarrhea and vomiting during a natural disaster that took place on May 31, 2000. The Instituto de Diagnóstico y Referencia Epidemiológicos (Epidemic Reference and Diagnosis Institute, InDRE, Ministry of Health, received 1521 rectal

Method for enzyme synthesis of radioactive thymine 5'-deoxyribonucleotides

International Nuclear Information System (INIS)

Nejedly, Z.; Ekl, J.; Hybs, K.; Kolina, J.; Filip, J.; Votruba, I.; Skoda, J.

1978-01-01

The enzyme synthesis is described for thymidine-5'-monophosphate, thymidine-5'-diphosphate and thymidine-5'-triphosphate specifically or nonspecifically labelled with 14 C or 3 H. The anabolic transformation of radioactive thymine to radioactive thymine 5'-deoxyribonucleotides is catalyzed by the action of enzyme preparations separated from Escherichia coli bacteria. It is achieved by the action of nonpurified cell-free extracts on special auxotrophic mutants of the thymine-dependent Escherichia coli SPT - strain in the presence of deoxyriboso-1-phosphate and adenosine-5'-triphosphate. The radioactive thymidine-5'-monophosphate may further be phosphorylated. In reaction mixtures, radioactive thymine, deoxyriboso-1-phosphate and adenosine-5'-triphosphate are used in molar ratios of 1:1:2 to 1:10:100, the optimum molar ratio being 1:5:10. (B.S.)

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.

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.

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

Assessment of the efficiency of ColiSure™ for coliforms Escherichia coli enumeration in pasteurizad milk / Avaliação do desempenho do ColiSure™ na enumeração de coliformes totais e Escherichia coli em leite pasteurizado

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Elsa Helena Walter de Santana

2000-12-01

Full Text Available In the dairy industry the coliforms detection can he used as indicative of hygiene production of the raw milk and the contamination after-pasteurization. The traditional methods for the enumeration of the total and faecal coliforms are laborious and needs an incubation time of 96 hours. Rapid methods for detection of these microorganisms have been developed and among them the ColsSuroit is a rapid method that gives results in 24 hours and involves defined substrates for simultaneous determination of total conforms and E. coli in water based in specific enzymatic reactions of these microorganisms. The objective of this study was to evaluate its utilization in milk. Ninety-five samples of pasteurized milk were collected from the markets in Londrina city, Parana and analyzed by the Most Probable Number (NMP enumeration using the Brilliant Green Bile Lactose Broth (BGBL and ColiSure™. There was a correlation of 0.80 betwee-n the mediums when the incubation time was 43 hours for total conforms. The low down occurrence of E. coffin the analyzed samples made impossible to comparate the performance of methods for this microorganism. Compfementary analysis showed a greater sensibility and especifity of the ColiSure™ in comparation with the BGBL. The CofiSure™ can be indicated as a substitute for the traditional method, with the advantage to be faster and easier.Na indústria láctea a detecção de microrganismos do grupo coliformes é utilizada como indicativo da higiene na produção do leite e de contaminação pós-pasteurizaçüo. Os métodos tradicionais para a enumeração de coliformes totais e fecais são trabalhosos, com tempo de incubação longo, de até 96 horas. Métodos rápidos para a detecção destes microrganismos têm sido desenvolvidos na área de microbiologia de alimentos. O ColiSure™ é um método rápido, que fornece resultados simultaneamente para a presença ou a ausência de coliformes totais e Escherichia coli em �

Escherichia Coli Removal from Water Using Electrophotocatalytic ...

African Journals Online (AJOL)

Michael Horsfall

inactivation of bacterial microorganisms in areas with low ... disinfection of water contaminated with fecal indicators such as E. coli ... media, brain heart infusion, sodium chloride, sodium hydroxide ... furnace at temperature 105 and 320°C f0r 60 min. For 2- and .... charge of E. coli logarithmic growth phase might affect the ...

The Escherichia coli transcriptome linked to growth fitness

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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

E. coli Surface Properties Differ between Stream Water and Sediment Environments

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Xiao Liang

2016-11-01

Full Text Available The importance of E. coli as an indicator organism in fresh water has led to numerous studies focusing on cell properties and transport behavior. However, previous studies have been unable to assess if differences in E. coli cell surface properties and genomic variation are associated with different environmental habitats. In this study, we investigated the variation in characteristics of E. coli obtained from stream water and stream bottom sediments. Cell properties were measured for 77 genomically different E. coli strains (44 strains isolated from sediments and 33 strains isolated from water under common stream conditions in the Upper Midwestern United States: pH 8.0, ionic strength 10mM and 22˚C. Measured cell properties include hydrophobicity, zeta potential, net charge, total acidity and extracellular polymeric substance (EPS composition. Our results indicate that stream sediment E. coli had significantly greater hydrophobicity, greater EPS protein content and EPS sugar content, less negative net charge, and higher point of zero charge than stream water E. coli. A significant positive correlation was observed between hydrophobicity and EPS protein for stream sediment E. coli but not for stream water E. coli. Additionally, E. coli surviving in the same habitat tended to have significantly larger (GTG5 genome similarity. After accounting for the intrinsic impact from the genome, environmental habitat was determined to be a factor influencing some cell surface properties, such as hydrophobicity. The diversity of cell properties and its resulting impact on particle interactions should be considered for environmental fate and transport modeling of aquatic indicator organisms such as E. coli.

E. coli Surface Properties Differ between Stream Water and Sediment Environments.

Science.gov (United States)

Liang, Xiao; Liao, Chunyu; Thompson, Michael L; Soupir, Michelle L; Jarboe, Laura R; Dixon, Philip M

2016-01-01

The importance of E. coli as an indicator organism in fresh water has led to numerous studies focusing on cell properties and transport behavior. However, previous studies have been unable to assess if differences in E. coli cell surface properties and genomic variation are associated with different environmental habitats. In this study, we investigated the variation in characteristics of E. coli obtained from stream water and stream bottom sediments. Cell properties were measured for 77 genomically different E. coli strains (44 strains isolated from sediments and 33 strains isolated from water) under common stream conditions in the Upper Midwestern United States: pH 8.0, ionic strength 10 mM and 22°C. Measured cell properties include hydrophobicity, zeta potential, net charge, total acidity, and extracellular polymeric substance (EPS) composition. Our results indicate that stream sediment E. coli had significantly greater hydrophobicity, greater EPS protein content and EPS sugar content, less negative net charge, and higher point of zero charge than stream water E. coli . A significant positive correlation was observed between hydrophobicity and EPS protein for stream sediment E. coli but not for stream water E. coli . Additionally, E. coli surviving in the same habitat tended to have significantly larger (GTG) 5 genome similarity. After accounting for the intrinsic impact from the genome, environmental habitat was determined to be a factor influencing some cell surface properties, such as hydrophobicity. The diversity of cell properties and its resulting impact on particle interactions should be considered for environmental fate and transport modeling of aquatic indicator organisms such as E. coli .

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.

Antimicrobial susceptibility patterns of E. coli from clinical sources in ...

African Journals Online (AJOL)

Background: Escherichia coli is the leading cause of urinary tract, ear, wound and other infections in humans. Increasing rates of antimicrobial resistance among E. coli is a growing concern worldwide. Objectives: The aim of this study was to determine the prevalence and antimicrobial susceptibility of E. coli from clinical ...

Meta-Analysis of Transcriptional Responses to Mastitis-Causing Escherichia coli.

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Sidra Younis

Full Text Available Bovine mastitis is a widespread disease in dairy cows, and is often caused by bacterial mammary gland infection. Mastitis causes reduced milk production and leads to excessive use of antibiotics. We present meta-analysis of transcriptional profiles of bovine mastitis from 10 studies and 307 microarrays, allowing identification of much larger sets of affected genes than any individual study. Combining multiple studies provides insight into the molecular effects of Escherichia coli infection in vivo and uncovers differences between the consequences of E. coli vs. Staphylococcus aureus infection of primary mammary epithelial cells (PMECs. In udders, live E. coli elicits inflammatory and immune defenses through numerous cytokines and chemokines. Importantly, E. coli infection causes downregulation of genes encoding lipid biosynthesis enzymes that are involved in milk production. Additionally, host metabolism is generally suppressed. Finally, defensins and bacteria-recognition genes are upregulated, while the expression of the extracellular matrix protein transcripts is silenced. In PMECs, heat-inactivated E. coli elicits expression of ribosomal, cytoskeletal and angiogenic signaling genes, and causes suppression of the cell cycle and energy production genes. We hypothesize that heat-inactivated E. coli may have prophylactic effects against mastitis. Heat-inactivated S. aureus promotes stronger inflammatory and immune defenses than E. coli. Lipopolysaccharide by itself induces MHC antigen presentation components, an effect not seen in response to E. coli bacteria. These results provide the basis for strategies to prevent and treat mastitis and may lead to the reduction in the use of antibiotics.

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.

Adsorption, sedimentation, and inactivation of E. coli within wastewater treatment wetlands.

Science.gov (United States)

Boutilier, L; Jamieson, R; Gordon, R; Lake, C; Hart, W

2009-09-01

Bacteria fate and transport within constructed wetlands must be understood if engineered wetlands are to become a reliable form of wastewater treatment. This study investigated the relative importance of microbial treatment mechanisms in constructed wetlands treating both domestic and agricultural wastewater. Escherichia coli (E. coli) inactivation, adsorption, and settling rates were measured in the lab within two types of wastewater (dairy wastewater lagoon effluent and domestic septic tank effluent). In situ E. coli inactivation was also measured within a domestic wastewater treatment wetland and the adsorption of E. coli was also measured within the wetland effluent. Inactivation of E. coli appears to be the most significant contributor to E. coli removal within the wastewaters and wetland environments examined in this study. E. coli survived longer within the dairy wastewater (DW) compared to the domestic wastewater treatment wetland water (WW). First order rate constants for E. coli inactivation within the WW in the lab ranged from 0.09 day(-1) (d(-1)) at 7.6 degrees C to 0.18d(-1) at 22.8 degrees C. The average in situ rate constant observed within the domestic wetland ranged from 0.02 d(-1) to 0.03 d(-1) at an average water temperature of 17 degrees C. First order rate constants for E. coli inactivation within the DW ranged from 0.01 d(-1) at 7.7 degrees C to 0.04 d(-1) at 24.6 degrees C. Calculated distribution coefficients (K(d)) were 19,000 mL g(-1), 324,000 mL g(-1), and 293 mL g(-1) for E. coli with domestic septic tank effluent (STE), treated wetland effluent (WLE), and DW, respectively. Approximately 50%, 20%, and 90% of E. coli were "free floating" or associated with particles 5 microm within both the STE and DW, settling did not appear to contribute to E. coli removal within sedimentation experiments, indicating that the particles the bacteria were associated with had very small settling velocities. The results of this study highlight the

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....

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...

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.

Escherichia coli in broiler chickens with airsacculitis

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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

Escherichia coli pathotypes in Pakistan from consecutive floods in 2010 and 2011.

Science.gov (United States)

Bokhari, Habib; Shah, Muhammad Ali; Asad, Saba; Akhtar, Sania; Akram, Muhammad; Wren, Brendan W

2013-03-01

This study compares Escherichia coli pathotypes circulating among children in Pakistan during the floods of 2010 and 2011 and from sporadic cases outside flood affected areas. Using multiplex polymerase chain reaction 115 of 205 stool samples (56.29%) were positive for diarrheagenic E. coli from specimens taken during the floods compared with 50 of 400 (12.5%) stool samples being positive for sporadic cases. The E. coli pathotypes were categorized as Enteropathogenic E. coli 33 (28.69%) and 13 (26%), Enterotoxigenic E. coli 29 (25.21%) and 15 (30%), Enteroaggregative E. coli 21 (18.2%) and 18 (36%), Enterohemorrhagic E. coli 5 (4.34%) and 1 (2%) from flood and sporadic cases, respectively. Furthermore, patients co-infected with more than one pathotype were 26 (22.60%) and 3 (6%) from flood and sporadic cases, respectively. The study shows an unexpectedly high rate of isolation of E. coli pathotypes suggesting Pakistan as an endemic region that requires active surveillance particularly during flood periods.

Experimental evolution of E. coli

Science.gov (United States)

Zhang, Mengshi

The evolution from unicellular to multicellular behavior is an essential step in the history of life. Our aim is to investigate the emergence of collective behavior in the model organism Escherichia coli (E. coli) and its selection advantages, such as better utilization of public goods. Our preliminary results suggest that the evolution of collective behavior may be a natural response to stressed conditions. Mailing address: Room 306 Science Centre North Block, The Chinese University of Hong Kong, Shatin, N.T. Hong Kong SAR. Phone: +852-3943-6354. Fax: +852-2603-5204. E-mail: mengshi0928@gmail.com.

Production and Purification Immunoglobulin against E. coli in Egg Yolk

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Mohammadreza Nassiri

2016-08-01

Full Text Available Introduction Chicken is the only avian species in which polyclonal antibodies, like IgG is transported from the hen to the egg yolk in a similar manner as the transport of mammalian IgG from the mother to the fetus. Immunoglobulin Y in the chicken is transported to the egg and accumulates in the egg yolk in large quantities. IgY is an egg yolk antibody that has been used widely for treatment and prevention of infections in humans and animal. IgY is used for passive protection of the pathogen infections such as Escherichia coli, bovine and human rotavirus, bovine coronavirus, salmonella, staphylococcus and Pseudomonas. IgY is a promising candidate as an alternative to antibiotics. Eschericha coli strains of serotype O157: H7 belongs to a family of pathogenic E. coli called enterohemorrhagic E. coli (EHEC strains responsible for hemorrhagic colitis, bloody or non-bloody diarrhea, and hemolytic uremic syndrome in humans. This strain of E. coli pathogenises by adhering to host intestinal epithelium and forming bacterial colonies. The purpose of this study was to produce and purify immunoglobulin Y against E. coli O157:H7 and develop specific polyclonal anti E. coli antibody in the egg yolk. Materials and Methods Sixteen-week-old laying hens (Mashhad, Iran were kept in individual cages with food and water ad libitum. Immunization of hens was performed by intramuscularly injecting killed E. coli O157: H7 with an equal volume of Freund’s complete adjuvant into two sides of chest area (Sigma, USA for the first immunization. Two booster immunizations followed up using complete and incomplete Freund’s adjuvants in two weeks interval. Freund’s adjuvant without antigen was injected to the control group. Two weeks after the last injection, the eggs were collected daily for eight weeks, marked and stored at 4 ºC. In order to IgY purification, eggs were collected. Purification of IgY from egg yolk was based on Polson and using PEG6000. Finally, the

Physicochemical Factors: Impact on Spermagglutination Induced by Escherichia coli

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Kiranjeet Kaur

2014-02-01

Full Text Available Motility is a sensitive parameter of sperm function which is predictive of its fertilization potential in vitro. The decrease in sperm motility may be associated with sperm agglutination and immobilization due to mere presence of bacteria or excretion of bacterial toxic products. Supplementation with various agents like sucrose, mannitol, calcium, and EDTA is well known to improve the sperm motility in vitro. The present study was designed to check any protective role exerted by the addition of different agents on spermatozoal motility against E. coli induced sperm agglutination. 52 semen specimens were screened for the presence of sperm-agglutinating strain of E. coli. Further, influence of various factors, namely, sugars, salts, and chelating agents was studied. Also, the impact of exposure to high temperature and alcohol on sperm-agglutinating efficiency of E. coli was observed. None of the factors could inhibit the sperm agglutination induced by E. coli, except high temperature suggesting the involvement of protein moiety. In addition, it was observed that agglutinating efficiency of E. coli was limited to spermatozoa and RBCs. It may be concluded that sperm-agglutinating property of E. coli is quite stable as various physicochemical factors tested did not show any negative effect on the same except high temperature.

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.

Quantitative Brightness Analysis of Fluorescence Intensity Fluctuations in E. Coli.

Directory of Open Access Journals (Sweden)

Kwang-Ho Hur

Full Text Available The brightness measured by fluorescence fluctuation spectroscopy specifies the average stoichiometry of a labeled protein in a sample. Here we extended brightness analysis, which has been mainly applied in eukaryotic cells, to prokaryotic cells with E. coli serving as a model system. The small size of the E. coli cell introduces unique challenges for applying brightness analysis that are addressed in this work. Photobleaching leads to a depletion of fluorophores and a reduction of the brightness of protein complexes. In addition, the E. coli cell and the point spread function of the instrument only partially overlap, which influences intensity fluctuations. To address these challenges we developed MSQ analysis, which is based on the mean Q-value of segmented photon count data, and combined it with the analysis of axial scans through the E. coli cell. The MSQ method recovers brightness, concentration, and diffusion time of soluble proteins in E. coli. We applied MSQ to measure the brightness of EGFP in E. coli and compared it to solution measurements. We further used MSQ analysis to determine the oligomeric state of nuclear transport factor 2 labeled with EGFP expressed in E. coli cells. The results obtained demonstrate the feasibility of quantifying the stoichiometry of proteins by brightness analysis in a prokaryotic cell.

Molecular photonic imaging of cancer using light-emitting e. coli

International Nuclear Information System (INIS)

Park, Jae Hyo; Min, Jung Joon; Moon, Sung Min; Kim, Hyun Ju; Hong, Yeong Jin; Choy, Hyon E.; Bom, Hee Seung; Jeong, Jae Ho; Cho, Kyoung Oh

2005-01-01

Cancer research has long sought a magic bullet that would selectively target and destroy malignant cells. In this study, we exploited that E. coli injected into tumor-bearing mice selectively target and proliferate in solid tumors by employing optical imaging technique. Lux operon or GFP has been cloned into pUC19 plasmid to engineer pUC19Lux or pUC19gfp which was transformed into varying kinds of wild type (MG1655) or mutant E.coli strains. For stable expression, lux operon was cloned with asd (aspartate β-semialdehyde dehydrogenase) gene and transformed into asd defective E. coli (MG1655asd-/asd+lux). These bacteria were i.v. injected into tumor mice or directly into central necrosis of tumor. The imaging signal from wild type E.coli was detected initially at liver (20min), then migrated to and shine in the tumor mass until 2 weeks of injection which was consistently observed in immuno-defective (nude) and -competent (Balb/c) mice. Imaging signal of stbaly transformed strain (MG1655asd-/asd+lux) was stronger and longer-lasting than that of transiently transformed strain (MG1655lux). Flagella defective E. coli strain failed to reach tumor loci. Only a few amounts of stress regulatory defective E. coli strain arrived at but couldn't survive at the tumor loci. E. coli colonies expressing GFP was mostly observed at the border of central necrosis and peripheral proliferative areas in immunofluorescence studies. Directly injected MG1655ad-/asd+lux was transiently observed at central necrosis followed by spreading to the peripheral tumor mass which was consistent with the finding by tail vein injection. We successfully engineered E. coli strain stably expressing lux reporter gene. E. coli strongly targeted solid tumor regardless of host immune status. Our results support that the targeting of tumor by E.coli is an active process and would be applied as a delivery vehicle of varying imaging markers or therapeutic molecules

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...

Rare Case of Polymicrobial Keratitis With Balantidium coli.

Science.gov (United States)

Hazarika, Manali; Pai H, Vijaya; Khanna, Vinay; Reddy, Harish; Tilak, Kriti; Chawla, Kiran

2016-12-01

To report a rare case of polymicrobial keratitis due to Balantidium coli and gram-negative bacteria, Pseudomonas aeruginosa and Klebsiella pneumoniae, in a soft contact lens (CL) wearer. We report a case of CL-related keratitis due to B. coli, P. aeruginosa, and K. pneumoniae. The culture of the corneal scrapings, the CL cleaning solution, and the CL revealed the growth of a rare ciliated parasite, B. coli, along with gram-negative bacteria, namely, P. aeruginosa and K. pneumoniae. The patient was successfully treated with topical broad-spectrum antibiotics and intravenous metronidazole. Polymicrobial keratitis has seldom been reported with B. coli as the causative agent. CL wear can be a risk factor for this infection. Treatment with topical antibiotics may not suffice, and the intravenous route of antiprotozoal drugs may be a useful adjunct. Increasing awareness, early diagnosis, and treatment may improve the final visual outcome.

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)

Reconstitution of active mycobacterial binuclear iron monooxygenase complex in Escherichia coli.

Science.gov (United States)

Furuya, Toshiki; Hayashi, Mika; Kino, Kuniki

2013-10-01

Bacterial binuclear iron monooxygenases play numerous physiological roles in oxidative metabolism. Monooxygenases of this type found in actinomycetes also catalyze various useful reactions and have attracted much attention as oxidation biocatalysts. However, difficulties in expressing these multicomponent monooxygenases in heterologous hosts, particularly in Escherichia coli, have hampered the development of engineered oxidation biocatalysts. Here, we describe a strategy to functionally express the mycobacterial binuclear iron monooxygenase MimABCD in Escherichia coli. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of the mimABCD gene expression in E. coli revealed that the oxygenase components MimA and MimC were insoluble. Furthermore, although the reductase MimB was expressed at a low level in the soluble fraction of E. coli cells, a band corresponding to the coupling protein MimD was not evident. This situation rendered the transformed E. coli cells inactive. We found that the following factors are important for functional expression of MimABCD in E. coli: coexpression of the specific chaperonin MimG, which caused MimA and MimC to be soluble in E. coli cells, and the optimization of the mimD nucleotide sequence, which led to efficient expression of this gene product. These two remedies enabled this multicomponent monooxygenase to be actively expressed in E. coli. The strategy described here should be generally applicable to the E. coli expression of other actinomycetous binuclear iron monooxygenases and related enzymes and will accelerate the development of engineered oxidation biocatalysts for industrial processes.

[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.

Pathogenic Potential to Humans of Bovine Escherichia coli O26, Scotland

Science.gov (United States)

Rosser, Tracy; Allison, Lesley J.; Courcier, Emily; Evans, Judith; McKendrick, Iain J.; Pearce, Michael C.; Handel, Ian; Caprioli, Alfredo; Karch, Helge; Hanson, Mary F.; Pollock, Kevin G.J.; Locking, Mary E.; Woolhouse, Mark E.J.; Matthews, Louise; Low, J. Chris; Gally, David L.

2012-01-01

Escherichia coli O26 and O157 have similar overall prevalences in cattle in Scotland, but in humans, Shiga toxin–producing E. coli O26 infections are fewer and clinically less severe than E. coli O157 infections. To investigate this discrepancy, we genotyped E. coli O26 isolates from cattle and humans in Scotland and continental Europe. The genetic background of some strains from Scotland was closely related to that of strains causing severe infections in Europe. Nonmetric multidimensional scaling found an association between hemolytic uremic syndrome (HUS) and multilocus sequence type 21 strains and confirmed the role of stx2 in severe human disease. Although the prevalences of E. coli O26 and O157 on cattle farms in Scotland are equivalent, prevalence of more virulent strains is low, reducing human infection risk. However, new data on E. coli O26–associated HUS in humans highlight the need for surveillance of non-O157 enterohemorrhagic E. coli and for understanding stx2 phage acquisition. PMID:22377426

Thiamin diphosphate-dependent enzymes: from enzymology to metabolic regulation, drug design and disease models.

Science.gov (United States)

Bunik, Victoria I; Tylicki, Adam; Lukashev, Nikolay V

2013-12-01

Bringing a knowledge of enzymology into research in vivo and in situ is of great importance in understanding systems biology and metabolic regulation. The central metabolic significance of thiamin (vitamin B1 ) and its diphosphorylated derivative (thiamin diphosphate; ThDP), and the fundamental differences in the ThDP-dependent enzymes of metabolic networks in mammals versus plants, fungi and bacteria, or in health versus disease, suggest that these enzymes are promising targets for biotechnological and medical applications. Here, the in vivo action of known regulators of ThDP-dependent enzymes, such as synthetic structural analogs of the enzyme substrates and thiamin, is analyzed in light of the enzymological data accumulated during half a century of research. Mimicking the enzyme-specific catalytic intermediates, the phosphonate analogs of 2-oxo acids selectively inhibit particular ThDP-dependent enzymes. Because of their selectivity, use of these compounds in cellular and animal models of ThDP-dependent enzyme malfunctions improves the validity of the model and its predictive power when compared with the nonselective and enzymatically less characterized oxythiamin and pyrithiamin. In vitro studies of the interaction of thiamin analogs and their biological derivatives with potential in vivo targets are necessary to identify and attenuate the analog selectivity. For both the substrate and thiamin synthetic analogs, in vitro reactivities with potential targets are highly relevant in vivo. However, effective concentrations in vivo are often higher than in vitro studies would suggest. The significance of specific inihibition of the ThDP-dependent enzymes for the development of herbicides, antibiotics, anticancer and neuroprotective strategies is discussed. © 2013 FEBS.

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 ...

77 FR 58091 - Risk-Based Sampling of Beef Manufacturing Trimmings for Escherichia coli (E. coli) O157:H7 and...

Science.gov (United States)

2012-09-19

... for exposure of carcasses and parts to any contamination or food safety hazard during the removal of... DEPARTMENT OF AGRICULTURE Food Safety and Inspection Service [Docket No. FSIS-2012-0020] Risk-Based Sampling of Beef Manufacturing Trimmings for Escherichia coli (E. coli) O157:H7 and Plans for Beef...

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

Genetic-Metabolic Coupling for Targeted Metabolic Engineering

DEFF Research Database (Denmark)

Cardinale, Stefano; Tueros Farfan, Felipe Gonzalo; Sommer, Morten Otto Alexander

2017-01-01

pertur-bations, thus limiting their relevance to specific conditions. Here, we address this issue by coupling cell fitness to the production of thiamine diphosphate in Escherichia coli using a synthetic RNA biosensor. We use this strategy to interrogate a library of transposon mutants and elucidate...... the native gene network influencing both cell fitness and thiamine production. Ultimately, we identify effectors of the OxyR-Fur stress response that limit thiamine biosynthesis via alternative regulation of iron storage and Fe-S cluster inclusion in enzymes. This study presents a new approach...

High-throughput enzyme screening platform for the IPP-bypass mevalonate pathway for isopentenol production

DEFF Research Database (Denmark)

Kang, Aram; Meadows, Corey W.; Canu, Nicolas

2017-01-01

Isopentenol (or isoprenol, 3-methyl-3-buten-1-ol) is a drop-in biofuel and a precursor for commodity chemicals such as isoprene. Biological production of isopentenol via the mevalonate pathway has been optimized extensively in Escherichia coli, yielding 70% of its theoretical maximum. However, high...... ATP requirements and isopentenyl diphosphate (IPP) toxicity pose immediate challenges for engineering bacterial strains to overproduce commodities utilizing IPP as an intermediate. To overcome these limitations, we developed an “IPP-bypass� isopentenol pathway using the promiscuous activity...

Glycan-functionalized diamond nanoparticles as potent E. coli anti-adhesives

Science.gov (United States)

Barras, Alexandre; Martin, Fernando Ariel; Bande, Omprakash; Baumann, Jean-Sébastien; Ghigo, Jean-Marc; Boukherroub, Rabah; Beloin, Christophe; Siriwardena, Aloysius; Szunerits, Sabine

2013-02-01

Bacterial attachment and subsequent biofilm formation on biotic surfaces or medical devices is an increasing source of infections in clinical settings. A large proportion of these biofilm-related infections are caused by Escherichia coli, a major nosocomial pathogen, in which the major adhesion factor is the FimH adhesin located at the tip of type 1 fimbriae. Inhibition of FimH-mediated adhesion has been identified as an efficient antibiotic-alternative strategy to potentially reduce E. coli-related infections. In this article we demonstrate that nanodiamond particles, covently modified with mannose moieties by a ``click'' chemistry approach, are able to efficiently inhibit E. coli type 1 fimbriae-mediated adhesion to eukaryotic cells with relative inhibitory potency (RIP) of as high as 9259 (bladder cell adhesion assay), which is unprecedented when compared with RIP values previously reported for alternate multivalent mannose-functionalized nanostructures designed to inhibit E. coli adhesion. Also remarkable is that these novel mannose-modified NDs reduce E. coli biofilm formation, a property previously not observed for multivalent glyco-nanoparticles and rarely demonstrated for other multivalent or monovalent mannose glycans. This work sets the stage for the further evaluation of these novel NDs as an anti-adhesive therapeutic strategy against E. coli-derived infections.Bacterial attachment and subsequent biofilm formation on biotic surfaces or medical devices is an increasing source of infections in clinical settings. A large proportion of these biofilm-related infections are caused by Escherichia coli, a major nosocomial pathogen, in which the major adhesion factor is the FimH adhesin located at the tip of type 1 fimbriae. Inhibition of FimH-mediated adhesion has been identified as an efficient antibiotic-alternative strategy to potentially reduce E. coli-related infections. In this article we demonstrate that nanodiamond particles, covently modified with

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 ...

Captive wild birds as reservoirs of enteropathogenic E. coli (EPEC) and Shiga-toxin producing E. coli (STEC).

Science.gov (United States)

Sanches, Lilian Aparecida; Gomes, Marcelo da Silva; Teixeira, Rodrigo Hidalgo Friciello; Cunha, Marcos Paulo Vieira; Oliveira, Maria Gabriela Xavier de; Vieira, Mônica Aparecida Midolli; Gomes, Tânia Aparecida Tardelli; Knobl, Terezinha

Psittacine birds have been identified as reservoirs of diarrheagenic Escherichia coli, a subset of pathogens associated with mortality of children in tropical countries. The role of other orders of birds as source of infection is unclear. The aim of this study was to perform the molecular diagnosis of infection with diarrheagenic E. coli in 10 different orders of captive wild birds in the state of São Paulo, Brazil. Fecal samples were analyzed from 516 birds belonging to 10 orders: Accipitriformes, Anseriformes, Columbiformes, Falconiformes, Galliformes, Passeriformes, Pelecaniformes, Piciformes, Psittaciformes and Strigiformes. After isolation, 401 E. coli strains were subjected to multiplex PCR system with amplification of genes eae and bfp (EPEC), stx1 and stx2 for STEC. The results of these tests revealed 23/401 (5.74%) positive strains for eae gene, 16/401 positive strains for the bfp gene (3.99%) and 3/401 positive for stx2 gene (0.75%) distributed among the orders of Psittaciformes, Strigiformes and Columbiformes. None of strains were positive for stx1 gene. These data reveal the infection by STEC, typical and atypical EPEC in captive birds. The frequency of these pathotypes is low and restricted to few orders, but the data suggest the potential public health risk that these birds represent as reservoirs of diarrheagenic E. coli. Copyright © 2017 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Genome analysis of E. coli isolated from Crohn's disease patients.

Science.gov (United States)

Rakitina, Daria V; Manolov, Alexander I; Kanygina, Alexandra V; Garushyants, Sofya K; Baikova, Julia P; Alexeev, Dmitry G; Ladygina, Valentina G; Kostryukova, Elena S; Larin, Andrei K; Semashko, Tatiana A; Karpova, Irina Y; Babenko, Vladislav V; Ismagilova, Ruzilya K; Malanin, Sergei Y; Gelfand, Mikhail S; Ilina, Elena N; Gorodnichev, Roman B; Lisitsyna, Eugenia S; Aleshkin, Gennady I; Scherbakov, Petr L; Khalif, Igor L; Shapina, Marina V; Maev, Igor V; Andreev, Dmitry N; Govorun, Vadim M

2017-07-19

Escherichia coli (E. coli) has been increasingly implicated in the pathogenesis of Crohn's disease (CD). The phylogeny of E. coli isolated from Crohn's disease patients (CDEC) was controversial, and while genotyping results suggested heterogeneity, the sequenced strains of E. coli from CD patients were closely related. We performed the shotgun genome sequencing of 28 E. coli isolates from ten CD patients and compared genomes from these isolates with already published genomes of CD strains and other pathogenic and non-pathogenic strains. CDEC was shown to belong to A, B1, B2 and D phylogenetic groups. The plasmid and several operons from the reference CD-associated E. coli strain LF82 were demonstrated to be more often present in CDEC genomes belonging to different phylogenetic groups than in genomes of commensal strains. The operons include carbon-source induced invasion GimA island, prophage I, iron uptake operons I and II, capsular assembly pathogenetic island IV and propanediol and galactitol utilization operons. Our findings suggest that CDEC are phylogenetically diverse. However, some strains isolated from independent sources possess highly similar chromosome or plasmids. Though no CD-specific genes or functional domains were present in all CD-associated strains, some genes and operons are more often found in the genomes of CDEC than in commensal E. coli. They are principally linked to gut colonization and utilization of propanediol and other sugar alcohols.

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

Drug Resistance Patterns of Escherichia coli in Ethiopia: A Meta-Analysis.

Science.gov (United States)

Tuem, Kald Beshir; Gebre, Abadi Kahsu; Atey, Tesfay Mehari; Bitew, Helen; Yimer, Ebrahim M; Berhe, Derbew Fikadu

2018-01-01

Antimicrobial drug resistance is a global threat for treatment of infectious diseases and costs life and money and threatens health delivery system's effectiveness. The resistance of E. coli to frequently utilized antimicrobial drugs is becoming a major challenge in Ethiopia. However, there is no inclusive countrywide study. Therefore, this study intended to assess the prevalence of E. coli resistance and antimicrobial-specific resistance pattern among E. coli clinical isolates in Ethiopia. Articles were retrieved from PubMed, Embase, and grey literature from 2007 to 2017. The main outcome measures were overall E. coli and drug-specific resistance patterns. A random-effects model was used to determine pooled prevalence with 95% confidence interval (CI), using DerSimonian and Laird method. In addition, subgroup analysis was conducted to improve the outcome. The study bias was assessed by Begg's funnel plot. This study was registered in PROSPERO as follows: PROSPERO 2017: CRD42017070106. Of 164 articles retrieved, 35 articles were included. A total of 19,235 study samples participated in the studies and 2,635 E. coli strains were isolated. Overall, E. coli antibacterial resistance was 45.38% (95% confidence interval (CI): 33.50 to 57.27). The resistance pattern ranges from 62.55% in Addis Ababa to 27.51% in Tigray region. The highest resistance of E. coli reported was to ampicillin (83.81%) and amoxicillin (75.79%), whereas only 13.55% of E. coli isolates showed resistance to nitrofurantoin. E. coli antimicrobial resistance remains high with disparities observed among regions. The bacterium was found to be highly resistant to aminopenicillins. The finding implies the need for effective prevention strategies for the E. coli drug resistance and calls for multifaceted approaches with full involvement of all stakeholders.

Highly sensitive immunoassay based on E. coli with autodisplayed Z-domain

International Nuclear Information System (INIS)

Jose, Joachim; Park, Min; Pyun, Jae-Chul

2010-01-01

The Z-domain of protein A has been known to bind specifically to the F c region of antibodies (IgGs). In this work, the Z-domain of protein A was expressed on the outer membrane of Escherichia coli by using 'Autodisplay' technology as a fusion protein of autotransport domain. The E. coli with autodisplayed Z-domain was applied to the sandwich-type immunoassay as a solid-support of detection-antibodies against a target analyte. For the feasibility demonstration of the E. coli based immunoassay, C-reactive protein (CRP) assay was carried out by using E. coli with autodisplayed Z-domain. The limit of detection (LOD) and binding capacity of the E. coli based immunoassay were estimated to be far more sensitive than the conventional ELISA. Such a far higher sensitivity of E. coli based immunoassay than conventional ELISA was explained by the orientation control of immobilized antibodies and the mobility of E. coli in assay matrix. From the test results of 45 rheumatoid arthritis (RA) patients' serum and 15 healthy samples, a cut-off value was established to have optimal sensitivity and selectivity values for RA. The CRP test result of each individual sample was compared with ELISA which is the reference method for RA diagnosis. From this work, the E. coli with Z-domain was proved to be feasible for the medical diagnosis based on sandwich-type immunoassay.

Replication and Transcription of Eukaryotic DNA in Esherichia coli

Science.gov (United States)

Morrow, John F.; Cohen, Stanley N.; Chang, Annie C. Y.; Boyer, Herbert W.; Goodman, Howard M.; Helling, Robert B.

1974-01-01

Fragments of amplified Xenopus laevis DNA, coding for 18S and 28S ribosomal RNA and generated by EcoRI restriction endonuclease, have been linked in vitro to the bacterial plasmid pSC101; and the recombinant molecular species have been introduced into E. coli by transformation. These recombinant plasmids, containing both eukaryotic and prokaryotic DNA, replicate stably in E. coli. RNA isolated from E. coli minicells harboring the plasmids hybridizes to amplified X. laevis rDNA. Images PMID:4600264

Antibiotic Resistance and Virulence Properties in Escherichia coli ...

African Journals Online (AJOL)

This study determined E. coli resistance to commonly used antibiotics together with their virulence properties in Ile-Ife, Nigeria. A total of 137 E. coli isolates from cases of urinary tract infection were tested for their sensitivity to commonly used antibiotics and possession of virulence factors using standard methods.

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

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.

Sucralose Increases Antimicrobial Resistance and Stimulates Recovery of Escherichia coli Mutants.

Science.gov (United States)

Qu, Yilin; Li, Rongyan; Jiang, Mingshan; Wang, Xiuhong

2017-07-01

Because of heavy use of antimicrobials, antimicrobial resistance in bacteria has become of great concern. The effect of some widely used food additives such as sucralose on bacteria in the gut and the environment has also drawn increasing attention. In this study, we investigated the interaction between antimicrobials and sucralose impacting antimicrobial resistance and mutation of Escherichia coli (E. coli). To examine antimicrobial resistance and mutation frequency, different subinhibitory concentrations of sucralose were added to cultures of E.coli BW25113 that were then treated with antimicrobials, oxolinic acid, or moxifloxacin. Then the E.coli were assayed for bacterial survival and recovery of mutants resistant to an unrelated antimicrobial, rifampicin. Pre-treatment of E.coli BW25113 with 1/2 minimal inhibitory concentration (MIC) of sucralose increased the survival rate in oxolinic acid or moxifloxacin. A 1/3 MIC of sucralose increased rifampicin-resistant mutation rate of E.coli BW25113 after 72 h, while rifampicin-resistant mutation rate was increased when co-treated with 1/8 MIC, 1/4 MIC, 1/3 MIC sucralose, and oxolinic acid after 24 h. Sucralose can increase the antimicrobial resistance and mutation frequency of E.coli to some antimicrobials.

Study of the irradiation effects on thorium phosphate diphosphate ({beta}-TPD): consequences on its chemical durability; Etude des effets d'irradiation sur le phosphate diphosphate de thorium ({beta}-PDT): consequences sur la durabilite chimique

Energy Technology Data Exchange (ETDEWEB)

Tamain, C

2005-12-15

Since Thorium Phosphate Diphosphate (beta-TPD) can be considered as a potential host matrix for long-term storage in underground repository, it is necessary to study the irradiation effects on the structure of this ceramics and the consequences on its chemical durability. Sintered samples of beta-TPD and of associated solid solutions of beta-TUPD were irradiated under ion beams and then altered in aqueous solutions. Depending on the electronic LET value, beta-TPD can be completely or partly amorphized. Furthermore, the ability of recrystallization of the amorphous material by thermal annealing was also demonstrated. Some leaching tests, realized on these irradiated samples, have shown a significant effect of the amorphous fraction on the normalized dissolution rate which was increased by a factor of 10 from the crystallized to the fully amorphized material. Correlatively, the amorphous fraction also modified the delay to reach the saturation conditions associated to the thermodynamic equilibria involved. On the other hand, it exhibited no influence neither on other kinetic parameters, such as activation energy of the dissolution process or partial order related to the proton concentration, nor on the nature of the neo-formed phase formed at the saturation of the leachate and identified as Thorium Phosphate Hydrogeno-Phosphate Hydrate (TPHPH). Beta-TUPD samples were also irradiated by gamma and alpha rays during leaching tests to study the effects of radiolysis in the leaching medium on the normalized leaching rate. It appeared that the radiolytic species occurring in the dissolution mechanism were unstable, disappearing quickly when stopping the irradiation. (author)

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

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.

Fibrillin 5 Is Essential for Plastoquinone-9 Biosynthesis by Binding to Solanesyl Diphosphate Synthases in Arabidopsis

Science.gov (United States)

Kim, Eun-Ha; Lee, Yongjik

2015-01-01

Fibrillins are lipid-associated proteins in plastids and are ubiquitous in plants. They accumulate in chromoplasts and sequester carotenoids during the development of flowers and fruits. However, little is known about the functions of fibrillins in leaf tissues. Here, we identified fibrillin 5 (FBN5), which is essential for plastoquinone-9 (PQ-9) biosynthesis in Arabidopsis thaliana. Homozygous fbn5-1 mutations were seedling-lethal, and XVE:FBN5-B transgenic plants expressing low levels of FBN5-B had a slower growth rate and were smaller than wild-type plants. In chloroplasts, FBN5-B specifically interacted with solanesyl diphosphate synthases (SPSs) 1 and 2, which biosynthesize the solanesyl moiety of PQ-9. Plants containing defective FBN5-B accumulated less PQ-9 and its cyclized product, plastochromanol-8, but the levels of tocopherols were not affected. The reduced PQ-9 content of XVE:FBN5-B transgenic plants was consistent with their lower photosynthetic performance and higher levels of hydrogen peroxide under cold stress. These results indicate that FBN5-B is required for PQ-9 biosynthesis through its interaction with SPS. Our study adds FBN5 as a structural component involved in the biosynthesis of PQ-9. FBN5 binding to the hydrophobic solanesyl moiety, which is generated by SPS1 and SPS2, in FBN5-B/SPS homodimeric complexes stimulates the enzyme activity of SPS1 and SPS2. PMID:26432861

Occurrence of Coliform and Escherichia coli Contamination and Absence of Escherichia coli O157:H7 on Romaine Lettuce from Retail Stores in the Upper Midwest.

Science.gov (United States)

Greve, Josephine D; Zietlow, Mark S; Miller, Kevin M; Ellingson, Jay L E

2015-09-01

A total of 720 whole, romaine lettuce heads were purchased from retail locations in the Upper Midwest and assessed for coliform and Escherichia coli contamination and for the presence of E. coli O157:H7. During a 16-month period (August 2010 through December 2011), coliform and E. coli counts were enumerated on Petrifilm, and the presence of E. coli O157:H7 and the virulence gene eae was evaluated by real-time PCR (qPCR). Over half (400 of 720) of the lettuce samples were processed with an immunomagnetic separation step before the qPCR assay. All retail lettuce samples were negative for E. coli O157:H7 when tested with the R.A.P.I.D. LT qPCR targeting a region of the O-antigen, and only two (0.28%) were positive for the eae gene when tested with LightCycler qPCR. On Petrifilm, coliform counts of most lettuce samples (96.4%) were between lettuce samples (98.2%) were lettuce heads. These results contribute to the limited recorded data and understanding of microbial contamination of whole romaine lettuce heads purchased from retail locations, specifically revealing the absence of E. coli O157:H7 and low levels of contamination with coliforms and other E. coli strains.

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.

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.

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.

Diarrheagenic Escherichia coli and acute and persistent diarrhea in returned travelers

NARCIS (Netherlands)

Schultsz, C.; van den Ende, J.; Cobelens, F.; Vervoort, T.; van Gompel, A.; Wetsteyn, J. C.; Dankert, J.

2000-01-01

To determine the role of diarrheagenic Escherichia coli in acute and persistent diarrhea in returned travelers, a case control study was performed. Enterotoxigenic E. coli (ETEC) was detected in stool samples from 18 (10.7%) of 169 patients and 4 (3.7%) of 108 controls. Enteroaggregative E. coli

Recyclable Escherichia coli-Specific-Killing AuNP-Polymer (ESKAP) Nanocomposites.

Science.gov (United States)

Yuan, Yuqi; Liu, Feng; Xue, Lulu; Wang, Hongwei; Pan, Jingjing; Cui, Yuecheng; Chen, Hong; Yuan, Lin

2016-05-11

Escherichia coli plays a crucial role in various inflammatory diseases and infections that pose significant threats to both human health and the global environment. Specifically inhibiting the growth of pathogenic E. coli is of great and urgent concern. By modifying gold nanoparticles (AuNPs) with both poly[2-(methacrylamido)glucopyranose] (pMAG) and poly[2-(methacryloyloxy)ethyl trimethylammonium iodide] (pMETAI), a novel recyclable E. coli-specific-killing AuNP-polymer (ESKAP) nanocomposite is proposed in this study, which based on both the high affinity of glycopolymers toward E. coli pili and the merits of antibacterial quaternized polymers attached to gold nanoparticles. The properties of nanocomposites with different ratios of pMAG to pMETAI grafted onto AuNPs are studied. With a pMAG:pMETAI feed ratio of 1:3, the nanocomposite appeared to specifically adhere to E. coli and highly inhibit the bacterial cells. After addition of mannose, which possesses higher affinity for the lectin on bacterial pili and has a competitive advantage over pMAG for adhesion to pili, the nanocomposite was able to escape from dead E. coli cells, becoming available for repeat use. The recycled nanocomposite retained good antibacterial activity for at least three cycles. Thus, this novel ESKAP nanocomposite is a promising, highly effective, and readily recyclable antibacterial agent that specifically kills E. coli. This nanocomposite has potential applications in biological sensing, biomedical diagnostics, biomedical imaging, drug delivery, and therapeutics.

Human exposure assessment to antibiotic-resistant Escherichia coli through drinking water.

Science.gov (United States)

O'Flaherty, E; Borrego, C M; Balcázar, J L; Cummins, E

2018-03-01

Antibiotic-resistant bacteria (ARB) are a potential threat to human health through drinking water with strong evidence of ARB presence in post treated tap water around the world. This study examines potential human exposure to antibiotic-resistant (AR) Escherichia coli (E. coli) through drinking water, the effect of different drinking water treatments on AR E. coli and the concentration of AR E. coli required in the source water for the EU Drinking Water Directive (DWD) (Council Directive 98/83/EC, 0CFU/100ml of E. coli in drinking water) to be exceeded. A number of scenarios were evaluated to examine different water treatment combinations and to reflect site specific conditions at a study site in Europe. A literature search was carried out to collate data on the effect of environmental conditions on AR E. coli, the effect of different water treatments on AR E. coli and typical human consumption levels of tap water. A human exposure assessment model was developed with probability distributions used to characterise uncertainty and variability in the input data. Overall results show the mean adult human exposure to AR E. coli from tap water consumption ranged between 3.44×10 -7 and 2.95×10 -1 cfu/day for the scenarios tested and varied depending on the water treatments used. The level of AR E. coli required in the source water pre-treatment to exceed the DWD varied between 1 and 5logcfu/ml, depending on the water treatments used. This can be used to set possible monitoring criteria in pre-treated water for potential ARB exposure in drinking water. Copyright © 2017 Elsevier B.V. All rights reserved.

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 ...

Characterization of Climical and Commensal Escherichia coli Isolates from an Integrated Turkey Operation

OpenAIRE

Altekruse, Sean Fitzgerald

2001-01-01

Pathogenic E. coli infections cause approximately one quarter of disease losses in commercial turkey flocks. A small subgroup of E. coli causes most infections. Epidemiologic studies of this disease have been hindered by a lack of reliable markers to discriminate between pathogenic and fecal E. coli and by the diversity of poultry strains. Reliance on antimicrobials to control E. coli infections has caused widespread antimicrobial resistance. One hundred five clinical E. coli were obtaine...

Uncomplicated E Coli Urinary Tract Infection in College Women: A Follow-Up Study of E Coli Sensitivities to Commonly Prescribed Antibiotics

Science.gov (United States)

Ansbach, Robert K.; Dybus, Karen; Bergeson, Rachel

2005-01-01

Treatment of uncomplicated urinary tract infections (UTIs) has changed in the past few years with researchers advocating empiric treatment for shorter periods of time without the use of cultures. Researchers report that antibiotic resistance of Escherichia coli (E coli) to commonly prescribed antibiotics in uncomplicated UTIs has been increasing.…

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.

Temporal variations of Escherichia coli concentrations in a large Midwestern river

Science.gov (United States)

Schilling, K.E.; Zhang, Y.-K.; Hill, D.R.; Jones, C.S.; Wolter, C.F.

2009-01-01

The Raccoon River used by the Des Moines Water Works to serve more than 400,000 people in central Iowa is threatened by contamination from Escherichia coli bacteria from point and nonpoint sources. The 9389 km2 watershed is highly agricultural, with 73% of the land in row crop production and widespread animal production. Results from 2155 grab samples from 1997 to 2005 for E. coli analysis were examined for temporal variations. E. coli concentrations were found to vary across years, seasons, and flow conditions, with a 9-year mean value of 1156 most probable number (MPN)/100 ml. Monthly concentrations exhibited clear seasonality with highest values in May through July. Although E. coli concentrations were higher during periods of greater discharge, the relation of log E. coli to log discharge was not particularly strong (r2 = 0.35). The variogram of E. coli concentrations showed temporal correlation within a span of 4 days suggesting that concentrations measured on 1 day may be related in time to concentrations measured up to 3 days later and beyond 4 days the concentrations vary randomly. The spectral analysis of the time series of E. coli was also carried out and was fitted well with the spectrum of an exponential covariance function. Deciphering temporal patterns and correlation of E. coli bacteria in streams may be useful for developing future monitoring strategies to track concentration patterns and loads. ?? 2008 Elsevier B.V. All rights reserved.

Isolation and evaluation of cocktail phages for the control of multidrug-resistant Escherichia coli serotype O104: H4 and E. coli O157: H7 isolates causing diarrhea.

Science.gov (United States)

Safwat Mohamed, Doaa; Farouk Ahmed, Eman; Mohamed Mahmoud, Abobakr; Abd El-Baky, Rehab Mahmoud; John, James

2018-02-01

Escherichia coli serotype O157: H7 and E. coli O104: H4 are well known foodborne pathogens causing sever enteric illness. Using bacteriophages as biocontrol agents of some foodborne pathogens and multidrug-resistant (MDR) bacteria has a great attention nowadays. This study aims to test the effect of cocktail phages on the growth of some foodborne pathogens and MDR E. coli. Routine conventional PCR was used to confirm the identification of E. coli isolates. Double-layered culture technique was used to isolate phages from sewage water. Morphology of bacteriophage was described using transmission electron microscopy, and spot test was performed to determine host range of the phage cocktail. Phage cocktail of Siphoviridae and Podoviridae family infecting E. coli O157: H7, E. coli O104: H4 and untypeable E. coli (neither O157 nor O104) has been isolated from sewage water. Phage cocktail showed both lytic and lysogenic activity. Lytic activity was observed against E. coli O157: H7, E. coli O104: H4 isolates, Staphylococcus. aureus ATCC6538 and Pseudomonas aeruginosa ATCC 10145, while the lysogenic activity was observed against the untypeable strain. The tested phage cocktail showed a promising inhibitory action on E. coli O157: H7 and O104: H4, S. aureus ATCC6538 and P. aeruginosa ATCC 10145, suggesting the possibility of its use as a biocontrol tool or as natural food preservatives for many food products. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

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)

Stimulation of Escherichia coli F-18Col- Type-1 fimbriae synthesis by leuX

DEFF Research Database (Denmark)

Newman, Joseph V.; Burghoff, Robert L.; Pallesen, Lars

1994-01-01

Escherichia coli F-18, a normal human fecal isolate, is an excellent colonizer of the streptomycin-treated mouse large intestine. E. coli F-18Col-, a derivative of E. coli F-18 which no longer makes the E. coli F-18 colicin, colonizes the large intestine as well as E. coli F-18 when fed to mice...... alone but is eliminated when fed together with E. coli F-18. Recently we randomly cloned E. coli F-18 DNA into E. coli F-18Col- and let the mouse intestine select the best colonizer. In this way, we isolated a 6.5-kb E. coli F-18 DNA sequence that simultaneously stimulated synthesis of type 1 fimbriae...... and enhanced E. coli F-18Col- colonizing ability. In the present investigation we show that the gene responsible for stimulation of type 1 fimbriae synthesis appears to be leuX, which encodes a tRNA specific for the rare leucine codon UUG. Moreover, it appears that expression of leuX may be regulated by two...

Transferability of antimicrobial resistance from multidrug-resistant Escherichia coli isolated from cattle in the USA to E. coli and Salmonella Newport recipients

Science.gov (United States)

The objective of this study was to evaluate conjugative transfer of cephalosporin resistance among (n=100) strains of multi-drug resistant Escherichia coli (MDRE) to Salmonella Newport and E. coli DH5-alpha recipients. To accomplish this, phenotypic and genotypic profiles were determined for MDRE, ...

Reproducible gene targeting in recalcitrant Escherichia coli isolates

Directory of Open Access Journals (Sweden)

De Greve Henri

2011-06-01

Full Text Available Abstract Background A number of allele replacement methods can be used to mutate bacterial genes. For instance, the Red recombinase system of phage Lambda has been used very efficiently to inactivate chromosomal genes in E. coli K-12, through recombination between regions of homology. However, this method does not work reproducibly in some clinical E. coli isolates. Findings The procedure was modified by using longer homologous regions (85 bp and 500-600 bp, to inactivate genes in the uropathogenic E. coli strain UTI89. An lrhA regulator mutant, and deletions of the lac operon as well as the complete type 1 fimbrial gene cluster, were obtained reproducibly. The modified method is also functional in other recalcitrant E. coli, like the avian pathogenic E. coli strain APEC1. The lrhA regulator and lac operon deletion mutants of APEC1 were successfully constructed in the same way as the UTI89 mutants. In other avian pathogenic E. coli strains (APEC3E, APEC11A and APEC16A it was very difficult or impossible to construct these mutants, with the original Red recombinase-based method, with a Red recombinase-based method using longer (85 bp homologous regions or with our modified protocol, using 500 - 600 bp homologous regions. Conclusions The method using 500-600 bp homologous regions can be used reliably in some clinical isolates, to delete single genes or entire operons by homologous recombination. However, it does not invariably show a greater efficiency in obtaining mutants, when compared to the original Red-mediated gene targeting method or to the gene targeting method with 85 bp homologous regions. Therefore the length of the homology regions is not the only limiting factor for the construction of mutants in these recalcitrant strains.

Occurrence and characterization of Shiga toxin-producing Escherichia coli O157:H7 and other non-sorbitol-fermenting E. coli in cattle and humans in urban areas of Morogoro, Tanzania.

Science.gov (United States)

Lupindu, Athumani M; Olsen, John E; Ngowi, Helena A; Msoffe, Peter L M; Mtambo, Madundo M; Scheutz, Flemming; Dalsgaard, Anders

2014-07-01

Escherichia coli strains such as Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli, enterotoxigenic, attaching, and effacing E. coli, and enteroinvasive E. coli cause diarrhea in humans. Although other serotypes exist, the most commonly reported STEC in outbreaks is O157:H7. A cross-sectional study was conducted to isolate and characterize non-sorbitol-fermenting (NSF) E. coli O157:H7 from urban and periurban livestock settings of Morogoro, Tanzania. Human stool, cattle feces, and soil and water samples were collected. Observations and questionnaire interview studies were used to gather information about cattle and manure management practices in the study area. E. coli were isolated on sorbitol MacConkey agar and characterized by conventional biochemical tests. Out of 1049 samples, 143 (13.7%) yielded NSF E. coli. Serological and antimicrobial tests and molecular typing were performed to NSF E. coli isolates. These procedures detected 10 (7%) pathogenic E. coli including STEC (n=7), enteropathogenic E. coli (EPEC) (n=2), and attaching and effacing E. coli (A/EEC) (n=1) strains. The STEC strains had the ability to produce VT1 and different VT2 toxin subtypes that caused cytopathic effects on Vero cells. The prevalence of STEC in cattle was 1.6%, out of which 0.9% was serotype O157:H7 and the overall prevalence of diarrheagenic E. coli in cattle was 2.2%. The serotypes O157:H7, O142:H34, O113:H21, O+:H-, O+:H16, and O25:H4 were identified. One ESBL-producing isolate showed the MLST type ST131. To our knowledge, this is the first finding in Tanzania of this recently emerged worldwide pandemic clonal group, causing widespread antimicrobial-resistant infections, and adds knowledge of the geographical distribution of ST131. Cattle manure was indiscriminately deposited within residential areas, and there was direct contact between humans and cattle feces during manure handling. Cattle and manure management practices expose humans, animals, and the environment

Escherichia coli O104 associated with human diarrhea, South Africa, 2004-2011.

Science.gov (United States)

Tau, Nomsa P; Meidany, Parastu; Smith, Anthony M; Sooka, Arvinda; Keddy, Karen H

2012-08-01

To determine the origin of >4,000 suspected diarrheagenic Escherichia coli strains isolated during 2004-2011 in South Africa, we identified 7 isolates as serotype O104; 5 as enteroaggregative E. coli O104:H4, and 2 as enteropathogenic E. coli O104:non-H4. Pulsed-field gel electrophoresis showed that these isolates were unrelated to the 2011 E. coli O104:H4 outbreak strain from Germany.

Sigma factors in a thousand E. coli genomes

DEFF Research Database (Denmark)

Cook, Helen Victoria; Ussery, David

2013-01-01

, 2013), only less than half (983) are of sufficient quality to use in comparative genomic work. Unfortunately, even some of the ‘complete’ E. coli genomes are in pieces, and a few ‘draft’ genomes are good quality. Six of the seven known sigma factors in E. coli strain K‐12 are extremely well conserved...

Characterization of Arabidopsis FPS isozymes and FPS gene expression analysis provide insight into the biosynthesis of isoprenoid precursors in seeds.

Science.gov (United States)

Keim, Verónica; Manzano, David; Fernández, Francisco J; Closa, Marta; Andrade, Paola; Caudepón, Daniel; Bortolotti, Cristina; Vega, M Cristina; Arró, Montserrat; Ferrer, Albert

2012-01-01

Arabidopsis thaliana contains two genes encoding farnesyl diphosphate (FPP) synthase (FPS), the prenyl diphoshate synthase that catalyzes the synthesis of FPP from isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). In this study, we provide evidence that the two Arabidopsis short FPS isozymes FPS1S and FPS2 localize to the cytosol. Both enzymes were expressed in E. coli, purified and biochemically characterized. Despite FPS1S and FPS2 share more than 90% amino acid sequence identity, FPS2 was found to be more efficient as a catalyst, more sensitive to the inhibitory effect of NaCl, and more resistant to thermal inactivation than FPS1S. Homology modelling for FPS1S and FPS2 and analysis of the amino acid differences between the two enzymes revealed an increase in surface polarity and a greater capacity to form surface salt bridges of FPS2 compared to FPS1S. These factors most likely account for the enhanced thermostability of FPS2. Expression analysis of FPS::GUS genes in seeds showed that FPS1 and FPS2 display complementary patterns of expression particularly at late stages of seed development, which suggests that Arabidopsis seeds have two spatially segregated sources of FPP. Functional complementation studies of the Arabidopsis fps2 knockout mutant seed phenotypes demonstrated that under normal conditions FPS1S and FPS2 are functionally interchangeable. A putative role for FPS2 in maintaining seed germination capacity under adverse environmental conditions is discussed.

The Escherichia coli argW-dsdCXA genetic island is highly variable, and E. coli K1 strains commonly possess two copies of dsdCXA.

Science.gov (United States)

Moritz, Rebecca L; Welch, Rodney A

2006-11-01

The genome sequences of Escherichia coli pathotypes reveal extensive genetic variability in the argW-dsdCXA island. Interestingly, the archetype E. coli K1 neonatal meningitis strain, strain RS218, has two copies of the dsdCXA genes for d-serine utilization at the argW and leuX islands. Because the human brain contains d-serine, an epidemiological study emphasizing K1 isolates surveyed the dsdCXA copy number and function. Forty of 41 (97.5%) independent E. coli K1 isolates could utilize d-serine. Southern blot hybridization revealed physical variability within the argW-dsdC region, even among 22 E. coli O18:K1:H7 isolates. In addition, 30 of 41 K1 strains, including 21 of 22 O18:K1:H7 isolates, had two dsdCXA loci. Mutational analysis indicated that each of the dsdA genes is functional in a rifampin-resistant mutant of RS218, mutant E44. The high percentage of K1 strains that can use d-serine is in striking contrast to our previous observation that only 4 of 74 (5%) isolates in the diarrheagenic E. coli (DEC) collection have this activity. The genome sequence of diarrheagenic E. coli isolates indicates that the csrRAKB genes for sucrose utilization are often substituted for dsdC and a portion of dsdX present at the argW-dsdCXA island of extraintestinal isolates. Among DEC isolates there is a reciprocal pattern of sucrose fermentation versus d-serine utilization. The ability to use d-serine is a trait strongly selected for among E. coli K1 strains, which have the ability to infect a wide range of extraintestinal sites. Conversely, diarrheagenic E. coli pathotypes appear to have substituted sucrose for d-serine as a potential nutrient.

Emergence and mechanism of carbapenem-resistant Escherichia coli in Henan, China, 2014

Directory of Open Access Journals (Sweden)

Wen-juan Liang

2018-05-01

Full Text Available The emergence and dissemination of carbapenem-resistant Escherichia coli (E. coli strains is a main risk for global public health, but little is known of carbapenemase producing E. coli in Henan, China. The study was undertaken to investigate the prevalence and mechanism of carbapenem-resistant E. coli strains in a hospital in Xinxiang, Henan, China, 2014. A total of 5 carbapenemase-producing E. coli strains were screened from 1014 isolates. We found that they were all resistant to meropenem and imipenem. Amikacin showed the best sensitivity, with gentamicin coming up next. The positive rate of blaNDM was 80% (4/5. The sequencing results showed that two isolates belonged to blaNDM-1 whereas other 2 isolates carried the blaNDM-5. Other carbapenemase genes including blaIMP, blaVIM, blaKPC and blaOXA-48 were not detected. The blaCTX-M-15, blaTEM-1, sul2, aad, and aac(6”–Ib–cr were also detected. MLST analysis showed that NDM-producing E. coli were sporadic. Conjugation test indicated blaNDM could be transferred. In conclusion, the blaNDM was the principal resistance mechanism of carbapenem-resistant E. coli in the hospital, Henan, China. Keywords: blaNDM, Carbapenem-resistant, Escherichia coli

Occurrence and characterization of Shiga toxin-producing Escherichia coli O157:H7 and other non-sorbitol-fermenting E. coli in cattle and humans in urban areas of Morogoro, Tanzania

DEFF Research Database (Denmark)

Lupindu, Athumani M; Olsen, John Elmerdahl; Ngowi, Helena A

2014-01-01

Escherichia coli strains such as Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli, enterotoxigenic, attaching, and effacing E. coli, and enteroinvasive E. coli cause diarrhea in humans. Although other serotypes exist, the most commonly reported STEC in outbreaks is O157:H7. A cross-...

Escherichia coli Contamination of Lettuce Grown in Soils Amended with Animal Slurry

DEFF Research Database (Denmark)

Jensen, Annette Nygaard; Storm, Christina; Forslund, Anita

2013-01-01

A pilot study was conducted to assess the transfer of Escherichia coli from animal slurry fertilizer to lettuce, with E. coli serving as an indicator of fecal contamination and as an indicator for potential bacterial enteric pathogens. Animal slurry was applied as fertilizer to three Danish agric...... types between slurry, soil, and lettuce. The frequent finding of fecal-contaminated lettuce indicates that human pathogens such as Salmonella and Campylobacter can be present and represent food safety hazards.......A pilot study was conducted to assess the transfer of Escherichia coli from animal slurry fertilizer to lettuce, with E. coli serving as an indicator of fecal contamination and as an indicator for potential bacterial enteric pathogens. Animal slurry was applied as fertilizer to three Danish....... coli. A relatively higher frequency of E. coli in lettuce compared with the soil samples at harvest suggests environmental sources of fecal contamination, e.g., wildlife. The higher frequency was supported by the finding of 21 different PFGE types among the E. coli isolates, with only a few common PFGE...

Longitudinal study of transmission of Escherichia coli from broiler breeders to broilers

DEFF Research Database (Denmark)

Poulsen, Louise Ladefoged; Thoefner, Ida; Bisgaard, Magne

2017-01-01

of E. coli, isolated from broiler breeders with salpingitis, to the progeny and the possibility of subsequent first week mortality. Four parent flocks were followed during the whole production period (20-60 weeks) by post mortem and bacteriological examination of randomly selected dead birds. Newly...... selected for Pulsed-Field-Gel-Electrophoresis (PFGE) and Multi-Locus-Sequence-Typing (MLST) to determine their clonal relationships. E. coli was the main cause of both salpingitis in parents and first week mortality in broilers, and E. coli dominated the bacterial flora of the cloaca of newly hatched...... chickens. PFGE of E. coli showed identical band patterns in isolates from the three different sources indicating a transmission of E. coli from parent birds to chickens. In conclusion, E. coli isolated from salpingitis in broiler parents were found to be transmitted to broilers in which some sequence types...

Protein (Cyanobacteria): 653003198 [PGDBj - Ortholog DB

Lifescience Database Archive (English)

Full Text Available phosphoribosyl)-5-amino-4-imidazole-carboxylate carboxylase Planktothrix agardhii MNPEALQQLLESVASGQITPTDALDK...IKYFDFEPVGDFARIDHHRKLRTGFPEVIWGLNKTPEQIIKIIEVMRQRNPVVMATRIEPHVYQQLQAQIPDLRYYEIAKICAIHPDEIPRSNSTGIITILTAGTADL

Protein (Cyanobacteria): 652996447 [PGDBj - Ortholog DB

Lifescience Database Archive (English)

Full Text Available phosphoribosyl)-5-amino-4-imidazole-carboxylate carboxylase Planktothrix agardhii MNPEALQQLLESVASGQITPTDALDK...IKYFDFEPVGDFARIDHHRKLRTGFPEVIWGLNKTPEQIIKIIEVMRQRNPVVMATRIEPHVYQQLQAQIPDLRYYEIAKICAIHPDEIPRSNSTGIITILTAGTADL

Determination of the prevalence of subclinical endometritis and evaluation of molecular characterization of Escherichia coli (E-coli separated of them in mares repeat breeder in Yazd province

Directory of Open Access Journals (Sweden)

Taktaz Hafshejani Taghi

2016-11-01

Full Text Available Escherichia coli are known as the most common cause of reproductive tract infection in mare. Due to the progressive process of antibiotics use and increasing prevalence of antibiotic resistance, the aim of this study is evaluate the prevalence of subclinical endometritis and antibiotic resistance genes in Escherichia coli isolated. In this study, 60 mares were used with infertility background. Diagnosis of endometritis was performed using history and ultrasonography. Cytology, culture, Antibiogram were done of samples and PCR test was used to examine the gene virulence and antibiotic resistance. E-coli bacteria was isolated 48/33 % from sample culture. In PCR test 66/21 % of bacteria had virulence gene. It was determined, the lowest resistance to chloramphenicol about 38/15% and greatest resistance into ampicillin, tetracycline and streptomycin with 23/69 percent, respectively. 93% samples cytology had neutrophil more than two and the agent of 50% showed E. coli. The cause of half of subclinical endometritis in infertile maresis E-coli bacteriaEscherichia coli are known as the most common cause of reproductive tract infection in mare. Due to the progressive process of antibiotics use and increasing prevalence of antibiotic resistance, the aim of this study is evaluate the prevalence of subclinical endometritis and antibiotic resistance genes in Escherichia coli isolated. In this study, 60 mares were used with infertility background. Diagnosis of endometritis was performed using history and ultrasonography. Cytology, culture, Antibiogram were done of samples and PCR test was used to examine the gene virulence and antibiotic resistance. E-coli bacteria was isolated 48/33 % from sample culture. In PCR test 66/21 % of bacteria had virulence gene. It was determined, the lowest resistance to chloramphenicol about 38/15% and greatest resistance into ampicillin, tetracycline and streptomycin with 23/69 percent, respectively. 93% samples cytology had

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 ...

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.

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...

Evaluation of the efficacy of an autogenous Escherichia coli vaccine in broiler breeders

DEFF Research Database (Denmark)

Li, Lili; Thøfner, Ida; Christensen, Jens Peter

2017-01-01

vaccine in broiler breeders. Three groups of 28 weeks old broiler breeders (unvaccinated, vaccinated once and twice, respectively) were challenged with a homologous E. coli strain (same strain as included in the vaccine) or a heterologous challenge strain in an experimental ascending model. The clinical...... infection, significant protection of an autogenous E. coli vaccine against neither a homologous nor a heterologous E. coli challenge could not be documented.......In poultry production Escherichia coli autogenous vaccines are often used. However, the efficacy of autogenous E. coli vaccinations has not been evaluated experimentally in chickens after start of lay. The aim of the present study was to evaluate the protective effect of an autogenous E. coli...

Transcriptome of E. coli K1 bound to human brain microvascular endothelial cells

OpenAIRE

Xie, Yi; Parthasarathy, Geetha; Di Cello, Francescopaolo; Teng, Ching-Hao; Paul-Satyaseela, Maneesh; Kim, Kwang Sik

2007-01-01

Escherichia coli K1 is the most common Gram-negative organism causing neonatal meningitis. Binding to human brain microvascdular endothelial cells (HBMEC) is an essential step for E. coli K1 traversal of the blood-brain barrier. In this study, we examined expression profiles of E. coli K1 strain RS218 during its binding to HBMEC. Comparison of HBMEC-bound E. coli K1 with collagen-bound E. coli revealed more than one hundred genes whose expression patterns were significantly changed in HBMEC-b...

Seroprevalence of Escherichia coli in traditional cheeses manufactured in Maragheh rural

Directory of Open Access Journals (Sweden)

S Mahdavi

2014-11-01

Full Text Available Coliforms and Escherichia coli are major microbial indicators in the accessing the quality of foodstuffs. The presence of these bacteria in foods is considered as an indication of fecal contamination. E. coli O157:H7 is the most pathogenic strain that is transmitted to human through animal-foods. This study was performed on 100 traditional cheese samples manufactured in Maragheh rural to determine the seroprevalence of E. coli. The samples were analyzed with standard microbiological methods followed by biochemical confirmatory tests. Afterwards, the isolates were assayed for the detection of O-serotypes using direct agglutination method. Among the 100 cheese samples, E. coli O157serotypewas not detected in any sample. However, other E. coli serotypes including 32 isolates of non-O157 serotypes were detected. Among the isolates, enteropathogenic, enterotoxigenic and enterohaemorhhagic serogruops was also detected.

Dialogue between E. coli free radical pathways and the mitochondria of C. elegans.

Science.gov (United States)

Govindan, J Amaranath; Jayamani, Elamparithi; Zhang, Xinrui; Mylonakis, Eleftherios; Ruvkun, Gary

2015-10-06

The microbial world presents a complex palette of opportunities and dangers to animals, which have developed surveillance and response strategies to hints of microbial intent. We show here that the mitochondrial homeostatic response pathway of the nematode Caenorhabditis elegans responds to Escherichia coli mutations that activate free radical detoxification pathways. Activation of C. elegans mitochondrial responses could be suppressed by additional mutations in E. coli, suggesting that C. elegans responds to products of E. coli to anticipate challenges to its mitochondrion. Out of 50 C. elegans gene inactivations known to mediate mitochondrial defense, we found that 7 genes were required for C. elegans response to a free radical producing E. coli mutant, including the bZip transcription factor atfs-1 (activating transcription factor associated with stress). An atfs-1 loss-of-function mutant was partially resistant to the effects of free radical-producing E. coli mutant, but a constitutively active atfs-1 mutant growing on wild-type E. coli inappropriately activated the pattern of mitochondrial responses normally induced by an E. coli free radical pathway mutant. Carbonylated proteins from free radical-producing E. coli mutant may directly activate the ATFS-1/bZIP transcription factor to induce mitochondrial stress response: feeding C. elegans with H2O2-treated E. coli induces the mitochondrial unfolded protein response, and inhibition of a gut peptide transporter partially suppressed C. elegans response to free radical damaged E. coli.

The population structure of Escherichia coli isolated from subtropical and temperate soils

Science.gov (United States)

Byappanahalli, Muruleedhara N.; Yan, Tao; Hamilton, Matthew J.; Ishii, Satoshi; Fujioka, Roger S.; Whitman, Richard L.; Sadowsky, Michael J.

2012-01-01

While genotypically-distinct naturalized Escherichia coli strains have been shown to occur in riparian soils of Lake Michigan and Lake Superior watersheds, comparative analyses of E. coli populations in diverse soils across a range of geographic and climatic conditions have not been investigated. The main objectives of this study were to: (a) examine the population structure and genetic relatedness of E. coli isolates collected from different soil types on a tropical island (Hawaii), and (b) determine if E. coli populations from Hawaii and temperate soils (Indiana, Minnesota) shared similar genotypes that may be reflective of biome-related soil conditions. DNA fingerprint and multivariate statistical analyses were used to examine the population structure and genotypic characteristics of the E. coli isolates. About 33% (98 of 293) of the E. coli from different soil types and locations on the island of Oahu, Hawaii, had unique DNA fingerprints, indicating that these bacteria were relatively diverse; the Shannon diversity index for the population was 4.03. Nearly 60% (171 of 293) of the E. coli isolates from Hawaii clustered into two major groups and the rest, with two or more isolates, fell into one of 22 smaller groups, or individual lineages. Multivariate analysis of variance of 89, 21, and 106 unique E. coli DNA fingerprints for Hawaii, Indiana, and Minnesota soils, respectively, showed that isolates formed tight cohesive groups, clustering mainly by location. However, there were several instances of clonal isolates being shared between geographically different locations. Thus, while nearly identical E. coli strains were shared between disparate climatologically- and geographically-distinct locations, a vast majority of the soil E. coli strains were genotypically diverse and were likely derived from separate lineages. This supports the hypothesis that these bacteria are not unique and multiple genotypes can readily adapt to become part of the soil autochthonous

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.

Protein (Cyanobacteria): 652402338 [PGDBj - Ortholog DB

Lifescience Database Archive (English)

Full Text Available 5-phosphoribosyl)-5-amino-4-imidazole-carboxylate carboxylase Planktothrix prolifica MNPEALQQLLESVASGQITPTDA...LDKIKYFDFEPVGDFARIDHHRKLRTGFPEVIWGLNKTPEQIIKIIEVMRQRNPVVMATRIEPHVYQQLQAQIPDLRYYEIAKICAIHPDEIPRSNSTGIITILTAGT

Molecular detection and antimicrobial resistance of diarrheagenic Escherichia coli strains isolated from diarrheal cases

International Nuclear Information System (INIS)

Aslani, Mehdi M.; Salmanzadeh-Ahrabi, S.; Jafari, F.; Zali, Reza M.; Mani, M.; Alikhani, Yousef M.

2008-01-01

Objective was to identify and classify Iranian isolates of diarrheagenic Escherichia coli (E. coli) on the basis of presence of virulence genes and to determine antibiotic susceptibility of isolated strains. The current cross-sectional study was conducted in 2005 at the Pasteur Institute, Tehran, Iran. One hundred and ninety-three diarrheagenic E. coli isolated from diarrheal patients in different regions of Iran were included in current study. Virulence factors genees for diarrheagenic E. coli were detected by polymerase chain reaction. Of the 193 diarrheagenic E. coli detected by PCR, 86(44.5%) were Shiga toxin-producing E. coli (STEC), 74 (38.4%) enteropathogenic E. coli (EPEC), 19 (9.8%) enteroaggregative E. coli and 14 (7.3%) enterotoxigenic E. coli isolates. Susceptibility to 12 clinically important antimicrobial agents was determined for 193 strains of diarrhheagenic E. coli. A high incidence of resistance to tetracycline (63%), ampicillin (62%), streptomycin (56%), amoxicillin/clavulanic acid (44.5%), trimetoprim/sulphamethoxazole (39.5%) and cephalothin (37%) was observed. The STEC and EPEC strains with high resistance to tetracycline and ampicillin but highly susceptible to quinolones are among the most important causative agent of diarrhea in Iran. This study suggests that antimicrobial resistance is wide spread among E. coli strains colonizing Iranian patients. Guidelines for appropriate use of antibiotics in developing countries require updating. (author)

Thermal inactivation of Escherichia coli 0157:H7 (ECOH) and non-0157 Shiga toxin-producing E.coli (STEC)in mechanically tenderized veal

Science.gov (United States)

We quantified thermal destruction of Shiga toxin-producing Escherichia coli O157:H7 (ECOH) and Shiga toxin-producing non-O157 E. coli (STEC) cells within mechanically tenderized veal cutlets following cooking on an electric skillet. For each of five trials, flattened veal cutlets (ca. 71.6 g; ca. 1/...

prevalence of escherichia coli 0157:h7 in fresh and roasted beef

African Journals Online (AJOL)

DR. AMINU

The prevalence of Enterohemorrhagic Escherichia coli 0157:H7 in 300 fresh beef and 150 roasted beef samples from ... likely cause of E. coli O157:H7 infection is undercooked ground beef. ..... coli O157:H7 in a sheep model. Appl. Environ.

Evaluation of mericon E. coli O157 Screen Plus and mericon E. coli STEC O-Type Pathogen Detection Assays in Select Foods: Collaborative Study, First Action 2017.05.

Science.gov (United States)

Bird, Patrick; Benzinger, M Joseph; Bastin, Benjamin; Crowley, Erin; Agin, James; Goins, David; Armstrong, Marcia

2018-05-01

QIAGEN mericon Escherichia coli O157 Screen Plus and mericon E. coli Shiga toxin-producing E. coli (STEC) O-Type Pathogen Detection Assays use Real-Time PCR technology for the rapid, accurate detection of E. coli O157 and the "big six" (O26, O45, O103, O111, O121, O145) (non-O157 STEC) in select food types. Using a paired study design, the assays were compared with the U.S. Department of Agriculture, Food Safety Inspection Service Microbiology Laboratory Guidebook Chapter 5.09 reference method for the detection of E. coli O157:H7 in raw ground beef. Both mericon assays were evaluated using the manual and an automated DNA extraction method. Thirteen technicians from five laboratories located within the continental United States participated in the collaborative study. Three levels of contamination were evaluated. Statistical analysis was conducted according to the probability of detection (POD) statistical model. Results obtained for the low-inoculum level test portions produced a difference between laboratories POD (dLPOD) value with a 95% confidence interval of 0.00 (-0.12, 0.12) for the mericon E. coli O157 Screen Plus with manual and automated extraction and mericon E. coli STEC O-Type with manual extraction and -0.01 (-0.13, 0.10) for the mericon E. coli STEC O-Type with automated extraction. The dLPOD results indicate equivalence between the candidate methods and the reference method.

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.

Plasmid Conjugation in E. coli and Drug Resistance | Igwe ...

African Journals Online (AJOL)

This study aimed at determining the antibiotics susceptibility pattern of E. coli isolates claimed to be multidrug resistance using disc diffusion method. It also determined the presence of transferable resistance plasmids through conjugation and evaluated the medical significance of plasmid encoding E. coli and drug ...

Anthranoid self-medication causing rapid development of melanosis coli

NARCIS (Netherlands)

M. Willems; H.R. van Buuren (Henk); R.R. de Krijger (Ronald)

2003-01-01

textabstractIt is widely known that long-term use of anthranoid-containing laxatives is the cause of melanosis coli. We describe a case of melanosis coli, which occurred in a 39-year-old liver transplant patient who took an over-the-counter product containing aloe, rheum and

Molecular typing of uropathogenic E. coli strains by the ERIC-PCR method.

Science.gov (United States)

Ardakani, Maryam Afkhami; Ranjbar, Reza

2016-04-01

Escherichia coli (E. coli) is the most common cause of urinary infections in hospitals. The aim of this study was to evaluate the ERIC-PCR method for molecular typing of uropathogenic E. coli strains isolated from hospitalized patients. In a cross sectional study, 98 E. coli samples were collected from urine samples taken from patients admitted to Baqiyatallah Hospital from June 2014 to January 2015. The disk agar diffusion method was used to determine antibiotic sensitivity. DNA proliferation based on repetitive intergenic consensus was used to classify the E. coli strains. The products of proliferation were electrophoresed on 1.5% agarose gel, and their dendrograms were drawn. The data were analyzed by online Insillico software. The method used in this research proliferated numerous bands (4-17 bands), ranging from 100 to 3000 base pairs. The detected strains were classified into six clusters (E1-E6) with 70% similarity between them. In this study, uropathogenic E. coli strains belonged to different genotypic clusters. It was found that ERIC-PCR had good differentiation power for molecular typing of uropathogenic E. coli strains isolated from the patients in the study.

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.

The significance of E. coli treatment in perinatal period

Directory of Open Access Journals (Sweden)

Ljubić Aleksandar D.

2016-01-01

Full Text Available Introduction: Bacteriuria of pregnancy is a common condition. Case report: Patient, 30-years, pregnant woman. During pregnancy, E. coli infection recurred in 4 times, applied Cephalexin and Ceftriaxone. The delivery was terminated by CS, GW 38; girl infant, AS 9. After the period of lactation: secretory status - the patient was a secretor of A and H blood type substance; ultrasonography and contrast radiography - presence of the third kidney. The therapy was added by vaccine UroVaxom, and there was no E. coli infection during 2 years follow up period. The Child is now 7 years old girl, having brilliant psychomotorical development. Possible child brain damage, lung damage, mental diseases are the reason for necessity E. coli infection treatment during pregnancy. Conclusion: All pregnant women should be screened for bacteriuria. E. coli is most commonly sensitive to group B antibiotics (cephalexin and amoxicillin, safe to be included in pregnancy. Long-term follow up of infants born from mothers having bacterial infection during pregnancy is necessary.

Treatment for bovine Escherichia coli mastitis - an evidence-based approach.

Science.gov (United States)

Suojala, L; Kaartinen, L; Pyörälä, S

2013-12-01

Bovine mastitis caused by Escherichia coli can range from being a subclinical infection of the mammary gland to a severe systemic disease. Cow-dependent factors such as lactation stage and age affect the severity of coliform mastitis. Evidence for the efficacy of antimicrobial treatment for E. coli mastitis is very limited. Antimicrobial resistance is generally not a limiting factor for treatment, but it should be monitored to detect changes in resistance profiles. The only antimicrobials for which there is some scientific evidence of beneficial effects in the treatment for E. coli mastitis are fluoroquinolones and cephalosporins. Both are critically important drugs, the use of which in animals destined for food should be limited to specific indications and should be based on bacteriological diagnosis. The suggested routine protocol in dairy herds could target the primary antimicrobial treatment for mastitis, specifically infections caused by gram-positive bacteria. In E. coli mastitis with mild to moderate clinical signs, a non-antimicrobial approach (anti-inflammatory treatment, frequent milking and fluid therapy) should be the first option. In cases of severe E. coli mastitis, parenteral administration of fluoroquinolones, or third- or fourth-generation cephalosporins, is recommended due to the risk of unlimited growth of bacteria in the mammary gland and ensuing bacteremia. Evidence for the efficacy of intramammary-administered antimicrobial treatment for E. coli mastitis is so limited that it cannot be recommended. Nonsteroidal anti-inflammatory drugs have documented the efficacy in the treatment for E. coli mastitis and are recommended for supportive treatment for clinical mastitis. © 2013 John Wiley & Sons Ltd.

Mechanisms of antibiotic resistance to enrofloxacin in uropathogenic Escherichia coli in dog

Science.gov (United States)

Escherichia coli (E. coli) urinary tract infections (UTIs) are becoming a serious problem both for pets and humans (zoonosis) due to the close contact and to the increasing resistance to antibiotics. Canine E. coli represents a good experimental model useful to study this pathology. Moreover, as des...

Immune responses to recombinants of the South African vaccine strain of lumpy skin disease virus generated by using thymidine kinase gene insertion.

Science.gov (United States)

Wallace, David B; Viljoen, Gerrit J

2005-04-27

The South African vaccine strain of lumpy skin disease virus (type SA-Neethling) is currently being developed as a vector for recombinant vaccines of economically important livestock diseases throughout Africa. In this study, the feasibility of using the viral thymidine kinase gene as the site of insertion was investigated and recombinant viruses were evaluated in animal trials. Two separate recombinants were generated and selected for homogeneity expressing either the structural glycoprotein gene of bovine ephemeral fever virus (BEFV) or the two structural glycoprotein genes of Rift Valley fever virus (RVFV). Both recombinants incorporate the enhanced green fluorescent protein (EGFP) as a visual marker and the Escherichia coli guanine phosphoribosyl transferase (gpt) gene for dominant positive selection. The LSDV-RVFV recombinant construct (rLSDV-RVFV) protected mice against virulent RVFV challenge. In a small-scale BEFV-challenge cattle trial the rLSDV-BEFV construct failed to fully protect the cattle against virulent challenge, although both a humoral and cellular BEFV-specific immune response was elicited.

Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract

DEFF Research Database (Denmark)

Connell, Hugh; Agace, William; Klemm, Per

1996-01-01

of Escherichia coli for the urinary tract by promoting bacterial persistence and enhancing the inflammatory responce to infection. In a clinical study, we observed that disease severity was greater in children infected with E. coli O1:K1:H7 isolates expressing type 1 fimbriae than in those infected with type 1...... negative isolates of the same serotype. The E. coli O1:K1:H7 isolates had the same electrophoretic type, were hemolysin-negative, expressed P fimbriae, and carried the fim DNA sequences. When tested in a mouse urinary tract infection model, the type 1-positive E. coli O1:K1:H7 isolates survived inhigher...... urinary tract infection model. E. coli CN1016 reconstituted with type 1 fimbriae had restored virulence similar to that of the wild-type parent strain. These results show that type 1 fimbriae in the genetic background of a uropathogenic strain contribute to the pathogenesis of E. coli in the urinary tract....

Comparison of agrobacterium mediated wheat and barley transformation with nucleoside diphosphate kinase 2 (NDPK2) gene

International Nuclear Information System (INIS)

Waheed, U.; Shah, M.M.; Smedley, M.; Harwood, W.

2016-01-01

An efficient and reliable transformation system is imperative for improvement of important crop species like barley and wheat. Wheat transformation is complex due to larger genome size and polyploidy while barley has a limitation of genotypic dependency. The objective of current study was to compare the relative transformation efficiency of wheat and barley using specific expression vector pBRACT 214-NDPK2 constructed through gateway cloning carrying Nucleoside Diphosphate Kinase 2 (NDPK2) gene. The vector was used to compare the transformation response in both crops using immature embryos through Agrobacterium mediated transformation. Both wheat and barley showed different responses towards callus induction and regeneration. Immature embryos of 1.5 to 2 mm in diameter was found optimum for wheat callus induction while 1 to 1.5 mm for barley. Both embryogenic and non-embryogenic calli were found in wheat with significantly greater tendency for embryogenecity in barley. The overall regeneration response was found different for all transformed wheat and barley cultivars. Wheat cultivars showed good response initially that drastically slowed down in later stages with the exception of Fielder that reached to the green shoots with good roots. The barley transformed lines showed good regeneration response as compared to wheat. PCR analysis of putative transformants using genomic DNA showed a maximum of 27% transformation efficiency in barely. No true transformation response was obtained in all cultivars of wheat used in this study. The protocol developed for wheat and barley transformation will greatly be helpful in crop improvement programme through genetic engineering especially in diploid relatives of cereals. (author)

Study of the irradiation effects on thorium phosphate diphosphate (β-TPD): consequences on its chemical durability

International Nuclear Information System (INIS)

Tamain, C.

2005-12-01

Since Thorium Phosphate Diphosphate (beta-TPD) can be considered as a potential host matrix for long-term storage in underground repository, it is necessary to study the irradiation effects on the structure of this ceramics and the consequences on its chemical durability. Sintered samples of beta-TPD and of associated solid solutions of beta-TUPD were irradiated under ion beams and then altered in aqueous solutions. Depending on the electronic LET value, beta-TPD can be completely or partly amorphized. Furthermore, the ability of recrystallization of the amorphous material by thermal annealing was also demonstrated. Some leaching tests, realized on these irradiated samples, have shown a significant effect of the amorphous fraction on the normalized dissolution rate which was increased by a factor of 10 from the crystallized to the fully amorphized material. Correlatively, the amorphous fraction also modified the delay to reach the saturation conditions associated to the thermodynamic equilibria involved. On the other hand, it exhibited no influence neither on other kinetic parameters, such as activation energy of the dissolution process or partial order related to the proton concentration, nor on the nature of the neo-formed phase formed at the saturation of the leachate and identified as Thorium Phosphate Hydrogeno-Phosphate Hydrate (TPHPH). Beta-TUPD samples were also irradiated by gamma and alpha rays during leaching tests to study the effects of radiolysis in the leaching medium on the normalized leaching rate. It appeared that the radiolytic species occurring in the dissolution mechanism were unstable, disappearing quickly when stopping the irradiation. (author)

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....

Removal of Escherichia coli via low frequency electromagnetic field in riverbank filtration system.

Science.gov (United States)

Selamat, Rossitah; Abustan, Ismail; Rizal Arshad, Mohd; Mokhtar Kamal, Nurul Hana

2018-04-01

The removal of Escherichia coli (E. coli) via low frequency of electromagnetic field (LF-EMF) with different magnetic field was studied. LF-EMF is known as a high magnetic susceptibility method, which could affect E. coli growth without the usage of chemicals. The aim of this study was to investigate the removal of E. coli by using LF-EMF in water abstraction for the riverbank filtration (RBF) application. The effect of LF-EMF with the intensity of 2 to 10mT and 50Hz on coiled column of 1mm copper wire at 1 to 6 hours was assessed. The removal of E. coli after exposing to LF-EMF on the column model was measured using most probable number (MPN/100mL) and colonies forming unit (CFU/100mL) methods. Water flows into the column were varied up to 6 hours and with flowrate of 100 mL/min. Experimental results demonstrate that 100% of E. coli was removed at 8mT after 6 hours exposure. The magnetic field at 10mT removed 100% of E. coli after 4 hours exposure. The results obtained in this study proved that the LF-EMF was efficient in E. coli removal from RBF system. These finding indicated that the LF-EMF intensities and time of exposure can affect the removal of E. coli.

Analysis of early-onset bloodstream infection due to Escherichia coli infection in premature babies

OpenAIRE

Chen, I-Lun; Huang, Hsin-Chun; Wu, Chih-Te; Ou-Yang, Mei-Chen; Chung, Mei-Yung; Chen, Chih-Cheng; Suen, Jau-Ling; Hung, Chih-Hsing

2017-01-01

Abstract In early-onset bacteremia among preterm neonates, Escherichia coli (E. coli) is the main pathogen and can cause a high mortality rate. Thus, the predictive factors of mortality and extended-spectrum ?-lactamase (ESBL)-producing E. coli in preterm babies with E. coli early-onset bacteremia were reported. We retrospectively reviewed preterm neonates who had E. coli bacteremia occurring within 3 days after birth between 2004 and 2015. Maternal and perinatal information were collected fr...

75 FR 14607 - Small Entity Compliance Guide: Bottled Water: Total Coliform and E. coli

Science.gov (United States)

2010-03-26

...] Small Entity Compliance Guide: Bottled Water: Total Coliform and E. coli; Availability AGENCY: Food and... the availability of a guidance for industry entitled ``Bottled Water: Total Coliform and E. coli... determine whether any of the coliform organisms are Escherichia coli (E. coli), an indicator of fecal...

Identification of genes important for growth of asymptomatic bacteriuria Escherichia coli in urine.

Science.gov (United States)

Vejborg, Rebecca M; de Evgrafov, Mari R; Phan, Minh Duy; Totsika, Makrina; Schembri, Mark A; Hancock, Viktoria

2012-09-01

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 of symptoms. While ABU E. coli can persist in the bladder for long periods of time, little is known about the genetic determinants required for its growth and fitness in urine. To identify such genes, we have employed a transposon mutagenesis approach using the prototypic ABU E. coli strain 83972 and the clinical ABU E. coli strain VR89. Six genes involved in the biosynthesis of various amino acids and nucleobases were identified (carB, argE, argC, purA, metE, and ilvC), and site-specific mutants were subsequently constructed in E. coli 83972 and E. coli VR89 for each of these genes. In all cases, these mutants exhibited reduced growth rates and final cell densities in human urine. The growth defects could be complemented in trans as well as by supplementation with the appropriate amino acid or nucleobase. When assessed in vivo in a mouse model, E. coli 83972carAB and 83972argC showed a significantly reduced competitive advantage in the bladder and/or kidney during coinoculation experiments with the parent strain, whereas 83972metE and 83972ilvC did not. Taken together, our data have identified several biosynthesis pathways as new important fitness factors associated with the growth of ABU E. coli in human urine.

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 ...

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 ...

Dietary N-Carbamylglutamate Supplementation Boosts Intestinal Mucosal Immunity in Escherichia coli Challenged Piglets.

Directory of Open Access Journals (Sweden)

Fengrui Zhang

Full Text Available N-carbamylglutamate (NCG has been shown to enhance performance in neonatal piglets. However, few studies have demonstrated the effect of NCG on the intestinal mucosal barrier. This study was conducted to determine the effects of dietary NCG supplementation on intestinal mucosal immunity in neonatal piglets after an Escherichia coli (E. coli challenge. New-born piglets (4 d old were assigned randomly to one of four treatments (n = 7, including (I sham challenge, (II sham challenge +50 mg/kg NCG, (III E. coli challenge, and (IV E. coli challenge +50 mg/kg NCG. On d 8, pigs in the E. coli challenge groups (III and IV were orally challenged with 5 mL of E. coli K88 (10(8 CFU/mL, whereas pigs in the sham challenge groups (I and II were orally dosed with an equal volume of water. On d 13, all piglets were sacrificed, and samples were collected and examined. The results show that average daily gain in the E. coli challenged piglets (III and IV was decreased (PE.coli<0.05. However, it tended to be higher in the NCG treated piglets (II and IV. Ileum secretory IgA, as well as IFN-γ, IL-2, IL-4 and IL-10 in ileal homogenates, were increased in E. coli challenged piglets (III and IV. Similarly, ileum SIgA and IL-10 levels, and CD4(+ percentage in NCG treated piglets (II and IV were higher than no-NCG treated piglets (PNCG<0.05. However, the IL-2 level was only decreased in the piglets of E. coli challenge + NCG group (IV compared with E. coli challenge group (III (P<0.05. No change in the IL-2 level of the sham challenged piglets (III was observed. In conclusion, dietary NCG supplementation has some beneficial effects on intestinal mucosal immunity in E. coli challenged piglets, which might be associated with stimulated lymphocyte proliferation and cytokine synthesis. Our findings have an important implication that NCG may be used to reduce diarrhea in neonatal piglets.

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.

Involvement of UV-inducible repair in pyrimidine dimer excision in Escherichia coli

International Nuclear Information System (INIS)

Masek, F.; Sedliakova, M.

1978-01-01

The influence of UV radiation on pyrimidine dimer excision in the cells of three excision-proficient E.coli strains was studied. For this purpose cells were irradiated with a first fluence of 300 ergs/mm 2 and at different time intervals with a second fluence of 500 ergs/mm 2 . After the second fluence dimer excision was found to be partly inhibited in E.coli B/r Hcr + and E.coli 15 555-7, but not in E.coli K12 SR20. (author)

Involvement of UV-inducible repair in pyrimidine dimer excision in Escherichia coli

Energy Technology Data Exchange (ETDEWEB)

Masek, F; Sedliakova, M [Slovenska Akademia Vied, Bratislava (Czechoslovakia)

1978-11-15

The influence of UV radiation on pyrimidine dimer excision in the cells of three excision-proficient E.coli strains was studied. For this purpose cells were irradiated with a first fluence of 300 ergs/mm/sup 2/ and at different time intervals with a second fluence of 500 ergs/mm/sup 2/. After the second fluence dimer excision was found to be partly inhibited in E.coli B/r Hcr/sup +/ and E.coli 15 555-7, but not in E.coli K12 SR20.

Factors affecting Escherichia coli concentrations at Lake Erie public bathing beaches

Science.gov (United States)

Francy, Donna S.; Darner, Robert A.

1998-01-01

The environmental and water-quality factors that affect concentrations of Escherichia coli (E. coli) in water and sediment were investigated at three public bathing beachesEdgewater Park, Villa Angela, and Sims Parkin the Cleveland, Ohio metropolitan area. This study was done to aid in the determination of safe recreational use and to help water- resource managers assess more quickly and accurately the degradation of recreational water quality. Water and lake-bottom sediments were collected and ancillary environmental data were compiled for 41 days from May through September 1997. Water samples were analyzed for E. coli concentrations, suspended sediment concentrations, and turbidity. Lake- bottom sediment samples from the beach area were analyzed for E. coli concentrations and percent dry weight. Concentrations of E. coli were higher and more variable at Sims Park than at Villa Angela or Edgewater Park; concentrations were lowest at Edgewater Park. Time-series plots showed that short-term storage (less than one week) of E. coli in lake-bottom sediments may have occurred, although no evidence for long-term storage was found during the sampling period. E. coli concentrations in water were found to increase with increasing wave height, but the resuspension of E. coli from lake-bottom sediments by wave action could not be adequately assessed; higherwave heights were often associated with the discharge of sewage containing E. coli during or after a rainfall and wastewater-treatment plant overflow. Multiple linear regression (MLR) was used to develop models to predict recreational water quality at the in water. The related variables included turbidity, antecedent rainfall, antecedent weighted rainfall, volumes of wastewater-treatment plant overflows and metered outfalls (composed of storm-water runoff and combined-sewer overflows), a resuspension index, and wave heights. For the beaches in this study, wind speed, wind direction, water temperature, and the prswimmers

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.

[Current antibiotic resistance profile of uropathogenic Escherichia coli strains and therapeutic consequences].

Science.gov (United States)

El Bouamri, M C; Arsalane, L; Kamouni, Y; Yahyaoui, H; Bennouar, N; Berraha, M; Zouhair, S

2014-12-01

Urinary tract infections (UTI) are a very common reason for consultation and prescription in current practice. Excessive or inappropriate use of antibiotics in treating urinary tract infections is responsible for the emergence and spread of multiresistant uropathogenic bacteria. To evaluate the isolation frequency and antibiotic resistance of uropathogenic Escherichia coli strains isolated at the Marrakech region. We conducted a retrospective study over a period of three years (from 1st January 2010 to 31 December 2012). It included all non-redundant uropathogenic E. coli strains isolated in the microbiology laboratory of the Avicenne hospital of Marrakech, Morocco. During this study, 1472 uropathogenic enterobacteriaceae were isolated including 924 non-repetitive E. coli strains, an overall isolation frequency of 63%. Antibiotic resistance of isolated E. coli strains showed resistance rates to amoxicillin (65%), sulfamethoxazole-triméthropime (55%), amoxicillin-clavulanic acid (43%), ciprofloxacin (22%), gentamicin (14%), nitrofurans (11%), amikacin (8%) and fosfomycin (7%). The number of E. coli strains resistant to C3G by ESBL production was 67, an average frequency of 4.5% of all isolated uropathogenic enterobacteria. The associated antibiotic resistance in the case of ESBL-producing E. coli were 82% for ciprofloxacin, 76% for sulfamethozole trimethoprim, 66% for gentamicin and 56% for amikacin. No resistance to imipenem was recorded for the isolated E. coli strains, which represents an imipenem sensitivity of 100%. Antibiotic resistance of uropathogenic E. coli strains limits treatment options and therefore constitutes a real public health problem. The regular updating of antibiotic susceptibility statistics of E. coli strains allows a better adaptation of the probabilistic antibiotic therapy to local epidemiological data. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Escherichia coli Sequence Type 131 H30 Is the Main Driver of Emerging Extended-Spectrum-β-Lactamase-Producing E. coli at a Tertiary Care Center.

Science.gov (United States)

Johnson, James R; Johnston, Brian; Thuras, Paul; Launer, Bryn; Sokurenko, Evgeni V; Miller, Loren G

2016-01-01

The H 30 strain of Escherichia coli sequence type 131 (ST131- H 30) is a recently emerged, globally disseminated lineage associated with fluoroquinolone resistance and, via its H 30Rx subclone, the CTX-M-15 extended-spectrum beta-lactamase (ESBL). Here, we studied the clonal background and resistance characteristics of 109 consecutive recent E. coli clinical isolates (2015) and 41 historical ESBL-producing E. coli blood isolates (2004 to 2011) from a public tertiary care center in California with a rising prevalence of ESBL-producing E. coli isolates. Among the 2015 isolates, ST131, which was represented mainly by ST131- H 30, was the most common clonal lineage (23% overall). ST131- H 30 accounted for 47% (8/17) of ESBL-producing, 47% (14/30) of fluoroquinolone-resistant, and 33% (11/33) of multidrug-resistant isolates. ST131- H 30 also accounted for 53% (8/14) of dually fluoroquinolone-resistant, ESBL-producing isolates, with the remaining 47% comprised of diverse clonal groups that contributed a single isolate each. ST131- H 30Rx, with CTX-M-15, was the major ESBL producer (6/8) among ST131- H 30 isolates. ST131- H 30 and H 30Rx also dominated (46% and 37%, respectively) among the historical ESBL-producing isolates (2004 to 2011), without significant temporal shifts in relative prevalence. Thus, this medical center's recently emerging ESBL-producing E. coli strains, although multiclonal, are dominated by ST131- H 30 and H 30Rx, which are the only clonally expanded fluoroquinolone-resistant, ESBL-producing lineages. Measures to rapidly and effectively detect, treat, and control these highly successful lineages are needed. IMPORTANCE The ever-rising prevalence of resistance to first-line antibiotics among clinical Escherichia coli isolates leads to worse clinical outcomes and higher health care costs, thereby creating a need to discover its basis so that effective interventions can be developed. We found that the H 30 subset within E. coli sequence type 131

Discovery and Characterization of Novel Nonsubstrate and Substrate NAMPT Inhibitors

Energy Technology Data Exchange (ETDEWEB)

Wilsbacher, Julie L.; Cheng, Min; Cheng, Dong; Trammell, Samuel A.J.; Shi, Yan; Guo, Jun; Koeniger, Stormy L.; Kovar, Peter J.; He, Yupeng; Selvaraju, Sujatha; Heyman, H. Robin; Sorensen, Bryan K.; Clark, Richard F.; Hansen, T. Matthew; Longenecker, Kenton L.; Raich, Diana; Korepanova, Alla V.; Cepa, Steven; Towne, Danli L.; Abraham, Vivek C.; Tang, Hua; Richardson, Paul L.; McLoughlin, Shaun M.; Badagnani, Ilaria; Curtin, Michael L.; Michaelides, Michael R.; Maag, David; Buchanan, F. Gregory; Chiang, Gary G.; Gao, Wenqing; Rosenberg, Saul H.; Brenner, Charles; Tse, Chris (AbbVie)

2017-05-03

Cancer cells are highly reliant on NAD+-dependent processes, including glucose metabolism, calcium signaling, DNA repair, and regulation of gene expression. Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme for NAD+ salvage from nicotinamide, has been investigated as a target for anticancer therapy. Known NAMPT inhibitors with potent cell activity are composed of a nitrogen-containing aromatic group, which is phosphoribosylated by the enzyme. Here, we identified two novel types of NAM-competitive NAMPT inhibitors, only one of which contains a modifiable, aromatic nitrogen that could be a phosphoribosyl acceptor. Both types of compound effectively deplete cellular NAD+, and subsequently ATP, and produce cell death when NAMPT is inhibited in cultured cells for more than 48 hours. Careful characterization of the kinetics of NAMPT inhibition in vivo allowed us to optimize dosing to produce sufficient NAD+ depletion over time that resulted in efficacy in an HCT116 xenograft model. Our data demonstrate that direct phosphoribosylation of competitive inhibitors by the NAMPT enzyme is not required for potent in vitro cellular activity or in vivo antitumor efficacy. Mol Cancer Ther; 16(7); 1236–45.

Synthesis and processing of escherichia-coli tem-beta-lactamase and bacillus-licheniformis alpha-amylase in escherichia-coli : The role of signal peptidase-i

NARCIS (Netherlands)

van Dijl, J M; Smith, H; Bron, Sierd; Venema, Gerard

A mutant of Escherichia coli, in which signal peptidase I synthesis can be regulated, was constructed. The mutant was used to study the effects of signal peptidase I limitation on the synthesis and efficiency of processing of two proteins: the periplasmic E. coli TEM-beta-lactamase and Bacillus

SYNTHESIS AND PROCESSING OF ESCHERICHIA-COLI TEM-BETA-LACTAMASE AND BACILLUS-LICHENIFORMIS ALPHA-AMYLASE IN ESCHERICHIA-COLI : THE ROLE OF SIGNAL PEPTIDASE-I

NARCIS (Netherlands)

van Dijl, J M; SMITH, H; BRON, S; VENEMA, G

A mutant of Escherichia coli, in which signal peptidase I synthesis can be regulated, was constructed. The mutant was used to study the effects of signal peptidase I limitation on the synthesis and efficiency of processing of two proteins: the periplasmic E. coli TEM-beta-lactamase and Bacillus

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 ...

Antibiotic Resistance of Escherichia coli Isolated from Healthy Food ...

African Journals Online (AJOL)

It was concluded from the study that healthy food animals form a reservoir of multiple resistant E. coli which may be transmitted to humans through the food chain. Thus, continued surveillance of E. coli obtained from food production continuum is merited to identify emerging antimicrobial-resistant phenotypes. The Kenya ...

The thorium phosphate diphosphate as matrix for radioactive waste conditioning: radionuclide immobilization and behavior under irradiation; Le phosphate diphosphate de thorium, matrice pour le conditionnement des dechets radioactifs: immobilisation de radionucleides, comportement sous irradiation

Energy Technology Data Exchange (ETDEWEB)

Pichot, Erwan [Inst. de Physique Nucleaire, Paris-11 Univ., 91 - Orsay (France)

1999-04-13

The aim of this work was to perform successively the decontamination of liquid solutions and the final immobilization of radionuclide storage using the same matrix. For this, thorium phosphate-diphosphate (TPD) of the formula Th{sub 4}P{sub 6}O{sub 23}, is proposed as a very resistant to water corrosion matrix. A new compound, thorium phosphate hydrogeno-phosphate (TPHP) of the formula Th{sub 2}(PO{sub 4}){sub 2}(HPO{sub 4}), nH{sub 2}O with n=3-7 was synthesized and characterized. Heated at 1100 deg.C it is transformed into the TDP. Ion exchange properties of TPHP were investigated. The exchange yields of imponderable caesium, strontium and americium ion onto TPHP (NaNO{sub 3} 0.1 M media at pH=6) are equal to 60% for the first one and 100% for the two others. The results interpreted in terms of ion-exchange led to determine selectivity coefficient values for each cation and suggested that only hydrated ions are exchanged. While the TPD is proposed for the high level nuclear waste storage, the irradiation effects, particularly structural modifications were studied using both {gamma} irradiation and charged particle irradiation. ESR and TL methods were carried out in order to identify radicals created during gamma radiation exposure. Correlation between ESR and TL experiments performed at room temperature clearly show three of PO{sub 3}{sup 2-} species and one POO{center_dot} species of free radicals. We have shown that Au-ion irradiation in the range of MeV energy involved TPD structure and chemical modifications. Important sputtering was interpreted in terms of local thermal chemical decomposition. We have shown, at room temperature, that the amorphization dose for heavy ion irradiation is between 0.1 to 0.4 dpa. (author) 146 refs., 46 figs., 21 tabs.

Explaining and modeling the concentration and loading of Escherichia coli in a stream-A case study.

Science.gov (United States)

Wang, Chaozi; Schneider, Rebecca L; Parlange, Jean-Yves; Dahlke, Helen E; Walter, M Todd

2018-09-01

Escherichia coli (E. coli) level in streams is a public health indicator. Therefore, being able to explain why E. coli levels are sometimes high and sometimes low is important. Using citizen science data from Fall Creek in central NY we found that complementarily using principal component analysis (PCA) and partial least squares (PLS) regression provided insights into the drivers of E. coli and a mechanism for predicting E. coli levels, respectively. We found that stormwater, temperature/season and shallow subsurface flow are the three dominant processes driving the fate and transport of E. coli. PLS regression modeling provided very good predictions under stormwater conditions (R 2  = 0.85 for log (E. coli concentration) and R 2  = 0.90 for log (E. coli loading)); predictions under baseflow conditions were less robust. But, in our case, both E. coli concentration and E. coli loading were significantly higher under stormwater condition, so it is probably more important to predict high-flow E. coli hazards than low-flow conditions. Besides previously reported good indicators of in-stream E. coli level, nitrate-/nitrite-nitrogen and soluble reactive phosphorus were also found to be good indicators of in-stream E. coli levels. These findings suggest management practices to reduce E. coli concentrations and loads in-streams and, eventually, reduce the risk of waterborne disease outbreak. Copyright © 2018. Published by Elsevier B.V.

The Impact of Media, Phylogenetic Classification, and E. coli Pathotypes on Biofilm Formation in Extraintestinal and Commensal E. coli From Humans and Animals.

Science.gov (United States)

Nielsen, Daniel W; Klimavicz, James S; Cavender, Tia; Wannemuehler, Yvonne; Barbieri, Nicolle L; Nolan, Lisa K; Logue, Catherine M

2018-01-01

Extraintestinal pathogenic Escherichia coli (ExPEC) include avian pathogenic E. coli (APEC), neonatal meningitis E. coli (NMEC), and uropathogenic E. coli (UPEC) and are responsible for significant animal and human morbidity and mortality. This study sought to investigate if biofilm formation by ExPEC likely contributes to these losses since biofilms are associated with recurrent urinary tract infections, antibiotic resistance, and bacterial exchange of genetic material. Therefore, the goal of this study was to examine differences in biofilm formation among a collection of ExPEC and to ascertain if there is a relationship between their ability to produce biofilms and their assignment to phylogenetic groups in three media types - M63, diluted TSB, and BHI. Our results suggest that ExPEC produce relatively different levels of biofilm formation in the media tested as APEC (70.4%, p = 0.0064) and NMEC (84.4%, p = 0.0093) isolates were poor biofilm formers in minimal medium M63 while UPEC isolates produced significantly higher ODs under nutrient-limited conditions with 25% of strains producing strong biofilms in diluted TSB ( p = 0.0204). Additionally, E. coli phylogenetic assignment using Clermont's original and revised typing scheme demonstrated significant differences among the phylogenetic groups in the different media. When the original phylogenetic group isolates previously typed as group D were phylogenetically typed under the revised scheme and examined, they showed substantial variation in their ability to form biofilms, which may explain the significant values of revised phylogenetic groups E and F in M63 ( p = 0.0291, p = 0.0024). Our data indicates that biofilm formation is correlated with phylogenetic classification and subpathotype or commensal grouping of E. coli strains.

The Detection Method of Escherichia coli in Water Resources: A Review

Science.gov (United States)

Nurliyana, M. R.; Sahdan, M. Z.; Wibowo, K. M.; Muslihati, A.; Saim, H.; Ahmad, S. A.; Sari, Y.; Mansor, Z.

2018-04-01

This article reviews several approaches for Escherichia coli (E. coli) bacteria detection from conventional methods, emerging method and goes to biosensor-based techniques. Detection and enumeration of E. coli bacteria usually required long duration of time in obtaining the result since laboratory-based approach is normally used in its assessment. It requires 24 hours to 72 hours after sampling to process the culturing samples before results are available. Although faster technique for detecting E. coli in water such as Polymerase Chain Reaction (PCR) and Enzyme-Linked Immunosorbent Assay (ELISA) have been developed, it still required transporting the samples from water resources to the laboratory, high-cost, complicated equipment usage, complex procedures, as well as the requirement of skilled specialist to cope with the complexity which limit their wide spread practice in water quality detection. Recently, development of biosensor device that is easy to perform, portable, highly sensitive and selective becomes indispensable in detecting extremely lower consolidation of pathogenic E. coli bacteria in water samples.

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

An evolutionary analysis of genome expansion and pathogenicity in Escherichia coli.

Science.gov (United States)

Bohlin, Jon; Brynildsrud, Ola B; Sekse, Camilla; Snipen, Lars

2014-10-09

There are several studies describing loss of genes through reductive evolution in microbes, but how selective forces are associated with genome expansion due to horizontal gene transfer (HGT) has not received similar attention. The aim of this study was therefore to examine how selective pressures influence genome expansion in 53 fully sequenced and assembled Escherichia coli strains. We also explored potential connections between genome expansion and the attainment of virulence factors. This was performed using estimations of several genomic parameters such as AT content, genomic drift (measured using relative entropy), genome size and estimated HGT size, which were subsequently compared to analogous parameters computed from the core genome consisting of 1729 genes common to the 53 E. coli strains. Moreover, we analyzed how selective pressures (quantified using relative entropy and dN/dS), acting on the E. coli core genome, influenced lineage and phylogroup formation. Hierarchical clustering of dS and dN estimations from the E. coli core genome resulted in phylogenetic trees with topologies in agreement with known E. coli taxonomy and phylogroups. High values of dS, compared to dN, indicate that the E. coli core genome has been subjected to substantial purifying selection over time; significantly more than the non-core part of the genome (pcoli genome size correlated with estimated HGT size (pcoli are largely attained through HGT. No associations were found between selective pressures operating on the E. coli core genome, as estimated using relative entropy, and genome size (p~0.98). On a larger time frame, genome expansion in E. coli, which is significantly associated with the acquisition of virulence factors, appears to be independent of selective forces operating on the core genome.

Virulence Factors and Antibiotic Resistance in Uropathogenic and Commensal Escherichia coli Isolates

Directory of Open Access Journals (Sweden)

Iraj Sedighi

2016-10-01

Full Text Available Background: Urinary Tract Infections (UTIs, including cystitis and pyelonephritis, are the most common infectious diseases in childhood. Aim and Objectives: Escherichia coli (E. coli account for as much as 90% of the community-acquired and also 50% of nosocomial UTIs. Therefore, the identification of E. coli strains and antibiotic resistance patterns is important for both clinical and epidemiological implications. Material and Methods: To characterize uropathogenic strains E. coli, we studied 100 strains recovered from both urine samples of children aged less than 7 years with community-acquired UTIs and stool samples of healthy children, respectively. Results: We assessed Virulence Factors (VFs and drug sensitivities of E. coli isolates. Drug sensitivities of the isolates were 94% (amikacin, 90% (nitrofurantoin, 66% (gentamicin, 56% (cefixime, 40% (nalidixic acid and 28% (cotrimoxazol. Laboratory tests showed that the prevalence of virulence factors ranged from 18% for hemolysin and P-fimbriae to 2% for type1-fimbriae. Most drug resistance was cotrimoxazole and amikacin was the lowest. P-fimbriae and hemolysin in uropathogenic E. coli were more frequent than non-pathogen type of E. coli. Conclusion: Although amikacin appeared to be the first choice for UTI in children, but nitrofurantoin seems to be practical and could be considered as the selective choice for uncomplicated lower UTIs.

Distribution of Escherichia Coli as Soil Pollutant around Antang Landfills

Science.gov (United States)

Artiningsih, Andi; Zubair, Hazairin; Imran, A. M.; Widodo, Sri

2018-03-01

Tamangapa Antang Landfill locates around the residential area and faces an air and water pollution due to an open dumping system in its operation. The system arises a potential pollution in air, water and soil. Sampling was done surround the landfill in two parts, parallel and perpendicular to the ground water flow. This study shows the abundance of E. coli bacteria in soil around the Antang Landfills at depth of 10 to 20 cm (93x105 cfu/gr of soil) in the direction of groundwater flow. While in other locations the E. coli bacteria is not detected. The abundance of E. coli bacteria is a conjunction factor from landfill and human activities surround the area. The absence of E. coli bacteria in other location highly interpreted that the landfill is the major contributor of pollutant.

Kinetic characterization of recombinant Bacillus coagulans FDP-activated l-lactate dehydrogenase expressed in Escherichia coli and its substrate specificity.

Science.gov (United States)

Jiang, Ting; Xu, Yanbing; Sun, Xiucheng; Zheng, Zhaojuan; Ouyang, Jia

2014-03-01

Bacillus coagulans is a homofermentative, acid-tolerant and thermophilic sporogenic lactic acid bacterium, which is capable of producing high yields of optically pure lactic acid. The l-(+)-lactate dehydrogenase (l-LDH) from B. coagulans is considered as an ideal biocatalyst for industrial production. In this study, the gene ldhL encoding a thermostable l-LDH was amplified from B. coagulans NL01 genomic DNA and successfully expressed in Escherichia coli BL21 (DE3). The recombinant enzyme was partially purified and its enzymatic properties were characterized. Sequence analysis demonstrated that the l-LDH was a fructose 1,6-diphosphate-activated NAD-dependent lactate dehydrogenase (l-nLDH). Its molecular weight was approximately 34-36kDa. The Km and Vmax values of the purified l-nLDH for pyruvate were 1.91±0.28mM and 2613.57±6.43μmol(minmg)(-1), respectively. The biochemical properties of l-nLDH showed that the specific activity were up to 2323.29U/mg with optimum temperature of 55°C and pH of 6.5 in the pyruvate reduction and 351.01U/mg with temperature of 55°C and pH of 11.5 in the lactate oxidation. The enzyme also showed some activity in the absence of FDP, with a pH optimum of 4.0. Compared to other lactic acid bacterial l-nLDHs, the enzyme was found to be relatively stable at 50°C. Ca(2+), Ba(2+), Mg(2+) and Mn(2+) ions had activated effects on the enzyme activity, and the enzyme was greatly inhibited by Ni(2+) ion. Besides these, l-nLDH showed the higher specificity towards pyruvate esters, such as methyl pyruvate and ethyl pyruvate. Copyright © 2014 Elsevier Inc. All rights reserved.

DNA fingerprinting approaches to trace Escherichia coli sharing between dogs and owners.

Science.gov (United States)

Naziri, Z; Derakhshandeh, A; Firouzi, R; Motamedifar, M; Shojaee Tabrizi, A

2016-02-01

To determine the prevalence of cross-species sharing of Escherichia coli between healthy dogs and humans living in the same household. Two faecal E. coli isolates from 25 healthy dog-owner pairs and 16 healthy control humans were tested using three fingerprinting methods. The prevalence of within-household sharing of E. coli was 4, 8 and 8% using pulsed-field gel electrophoresis, randomly amplified polymorphic DNA and enterobacterial repetitive intergenic consensus-PCR analyses respectively. Within-household bacterial sharing was more prevalent than across-household sharing (P fingerprinting techniques will help to find ways for reducing the economic impact of E. coli infections. This study support claims that public health concerns regarding the cross-species sharing of E. coli are warranted but this risk is minimal. © 2015 The Society for Applied Microbiology.

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...

Prevalence of Salmonella and E. coli in neonatal diarrheic calves

Directory of Open Access Journals (Sweden)

F.R. El-Seedy

2016-03-01

Full Text Available Neonatal calf diarrhea remains one of the most important problems faced by livestock, causing great economic losses. This study investigated the prevalence of Salmonella and Escherichia coli, especially enterotoxigenic E. coli (ETEC, in diarrheic calves. Fecal samples were collected from 127 diarrheic calves up to 3 months of age at 12 farms from different governorates in Egypt. 119 bacterial isolates (93.7% were recovered and the prevalences of Salmonella and E. coli in diarrheic calves were 18.1% and 75.6%, respectively. Serotyping of Salmonella isolates revealed that S. Enteritidis and S. Typhimurium were the most prevalent serotypes, representing 60.9% and 30.4%, respectively, while S. Dublin was 8.7%. Serogrouping of E. coli isolates showed that 10 O-serogroups were obtained where O26 and O103 were the most prevalent (17.7% of each. Salmonella serotypes showed positive results with PCR test using oligonucleotide primer amplifying 521 bp fragment of invA gene of Salmonella while 70% of E. coli serogroups possessed ETEC virulent gene (K99. The in-vitro antibiotic sensitivity test indicated that Salmonella serotypes showed high sensitivity against enrofloxacin, spectinomycin and neomycin while E. coli isolates showed high sensitivities against marbofloxacin, spectinomycin and neomycin only.

FREQUENCY AND DISTRIBUTION OF DIARRHOEAGENIC ESCHERICHIA COLI STRAINS ISOLATED FROM PEDIATRIC PATIENTS WITH DIARRHOEA IN BOSNIA AND HERZEGOVINA

OpenAIRE

Dedeić-Ljubović, AmeLa; Hukić, Mirsada; Bekić, DaRia; Zvizdić, AmrA

2009-01-01

Diarrhoeal disease is a major cause of illness and death among infants and young children worldwide. Among the Escherichia coli (E. coli) causing intestinal diseases, there are six well-described categories: enteroaggregative E. coli (EAEC), diffusely adherent E. coli (DAEC), enteroinvasive E. coli (EIEC), entero-pathogenic E. coli (EPEC), enterohaemorrhagic E. coli (EHEC) and enterotoxigenic E. coli (ETEC).

Probiotic Mixture Golden Bifido Prevents Neonatal Escherichia coli K1 Translocation via Enhancing Intestinal Defense

Directory of Open Access Journals (Sweden)

Qing Zeng

2017-09-01

Full Text Available Escherichia coli (E. coli K1 sepsis and meningitis is a severe infection characterized by high mortality in neonates. Successful colonization and translocation across the intestinal mucosa have been regarded as the critical steps for E. coli K1 sepsis and meningitis. We recently reported that the probiotic mixture, Golden Bifido (containing live Lactobacillus bulgaricus, Bifidobacterium, and Streptococcus thermophilus, LBS has a preventive role against neonatal E. coli K1 bacteremia and meningitis. However, the interaction between the neonatal gut barrier, probiotics and E. coli K1 is still not elucidated. The present study aims to investigate how LBS exerts its protective effects on neonatal gut barrier during E. coli K1 infection. The beneficial effects of LBS were explored in vitro and in vivo using human colon carcinoma cell lines HT-29 and rat model of neonatal E. coli K1 infection, respectively. Our results showed that stimulation with E. coli K1 was able to cause intestinal barrier dysfunction, which were reflected by E. coli K1-induced intestinal damage and apoptosis of intestinal epithelial cells, reduction of mucin, immunoglobulin A (IgA and tight junction proteins expression, as well as increase in intestinal permeability, all these changes facilitate E. coli K1 intestinal translocation. However, these changes were alleviated when HT-29 cells were treated with LBS before E. coli K1 infection. Furthermore, we found that LBS-treated neonatal rats (without E. coli K1 infection have showed higher production of mucin, ZO-1, IgA, Ki67 in intestinal mucosa as well as lower intestinal permeability than that of non-treated rats, indicating that LBS could accelerate the development of neonatal intestinal defense. Taken together, our results suggest that enhancement of the neonatal intestinal defense to fight against E. coli K1 translocation could be the potential mechanism to elucidate how LBS confers a protective effect against neonatal E

Pathogenic Escherichia coli producing Extended-Spectrum β-Lactamases isolated from surface water and wastewater.

Science.gov (United States)

Franz, Eelco; Veenman, Christiaan; van Hoek, Angela H A M; de Roda Husman, Ana; Blaak, Hetty

2015-09-24

To assess public health risks from environmental exposure to Extended-Spectrum β-Lactamases (ESBL)-producing bacteria, it is necessary to have insight in the proportion of relative harmless commensal variants and potentially pathogenic ones (which may directly cause disease). In the current study, 170 ESBL-producing E. coli from Dutch wastewater (n = 82) and surface water (n = 88) were characterized with respect to ESBL-genotype, phylogenetic group, resistance phenotype and virulence markers associated with enteroaggregative E. coli (EAEC), enteroinvasive E. coli (EIEC), enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), extraintesinal E. coli (ExPEC), and Shiga toxin-producing E. coli (STEC). Overall, 17.1% of all ESBL-producing E. coli were suspected pathogenic variants. Suspected ExPECs constituted 8.8% of all ESBL-producing variants and 8.3% were potential gastrointestinal pathogens (4.1% EAEC, 1.8% EPEC, 1.2% EIEC, 1.2% ETEC, no STEC). Suspected pathogens were significantly associated with ESBL-genotype CTX-M-15 (X(2) = 14.7, P antibiotics. In conclusion, this study demonstrates that the aquatic environment is a potential reservoir of E. coli variants that combine ESBL-genes, a high level of multi-drug resistance and virulence factors, and therewith pose a health risk to humans upon exposure.

Chemical interaction of potassium diphosphate with cadmium nitrate in aqueous solution. Khimicheskie vzaimodejstviya difosfatov kaliya s azotnokislym kadmiem v vodnom rastvore

Energy Technology Data Exchange (ETDEWEB)

Kokhanovskij, V V [AN Belorusskoj SSR, Minsk (Belarus). Inst. Obshchej i Neorganicheskoj Khimii

1993-06-01

Formation of low-soluble compounds in 1.5 mol/l isomolar cross section of K[sub 4]P[sub 2]O[sub 7]-Cd(NO[sub 3])[sub 2]-H[sub 2]O system was studied. Liquid phases are studied by the methods of refractometry and pH value measuring, an solid ones - by the methods of chemical and X-ray phase analysis, IR spectroscopy, chromatography and microscopy. Three individual chemical compounds K[sub 2]CdP[sub 2]O[sub 7] x 4H[sub 2]O, K[sub 2]Cd[sub 3](P[sub 2]O[sub 7])[sub 2] x 3H[sub 2]O and Cd[sub 2]P[sub 2]O[sub 7] x 3.5H[sub 2]O and some their mixtures were isolated and investigated. It is shown that doulble diphosphate K[sub 6]Cd(P[sub 2]O[sub 7])[sub 2] x 6H[sub 2]O does not precipitate spontanously, but instead of it in wide region of system K[sub 2]CdP[sub 2]O[sub 7] x 4H[sub 2]O crystallizes as elongated acicular crystals or as thin plates of improper form.

Chemical synthesis of guanosine diphosphate mannuronic acid (GDP-ManA) and its C-4-O-methyl and C-4-deoxy congeners.

Science.gov (United States)

Zhang, Qingju; Howell, P Lynne; Overkleeft, Herman S; Filippov, Dmitri V; van der Marel, Gijsbert A; Codée, Jeroen D C

2017-10-10

Described is the first synthesis of guanosine diphosphate mannuronic acid (GDP-ManA), the sugar donor used by algae and bacteria for the production of alginate, an anionic polysaccharide composed of β-d-mannuronic acid (ManA) and α-l-guluronic acid (GulA). Understanding the biosynthesis of these polyanionic polysaccharides on the molecular level, opens up avenues to use and modulate the biosynthesis machinery for biotechnological and therapeutic applications. The synthesis reported here delivers multi-milligram amounts of the GDP-ManA donor that can be used to study the polymerase (Alg8 in Pseudomonas aeruginosa) that generates the poly-ManA chain. Also reported is the assembly of two close analogues of GDP-ManA: the first bears a C-4-O-methyl group, while the second has been deoxygenated at this position. Both molecules may be used as "chain stoppers" in future enzymatic ManA polymerisation reactions. The crucial pyrophosphate linkage of the GDP-mannuronic acids has been constructed by the phosphorylation of the appropriate ManA-1-phosphates with a guanosine phosphoramidite. Copyright © 2017 Elsevier Ltd. All rights reserved.

Mathematical model of rhamnolipid production using E.coli bacteria

Science.gov (United States)

Adham, Muhammad Fariduddin; Apri, Mochamad; Moeis, Maelita Ramdani

2018-03-01

Rhamnolipid is one of biosurfactants that is widely used in many industries. Despite its wide use, production of rhamnolipid usually involves a pathogen that may endanger our health. To tackle this issue, in iGEM (International Genetically Engineered Machine) competition 2015, our team engineered Escherichia coli (E.coli) to produce rhamnolipid. The bacteria were then put into medium containing glucose and lactose. It turned out that bacteria E. coli produced lower rhamnolipid than that by pseudomonas, therefore a good strategy is required to improve their productivity. We present a mathematical model to describe the production of rhamnolipid by the engineered E coli. Using bifurcation analysis, the equilibrium points of the model and their stabilities were analyzed as the amount of lactose was varied. We show that the system produces bistability behavior for some interval values of lactose. From this analysis we found that to guarantee a high production of rhamnolipid, a high level of lactose is required. To maintain the productivity, however, it is sufficient to maintain the lactose level above a certain threshold value.

Efficacy of a Blend of Sulfuric Acid and Sodium Sulfate against Shiga Toxin-Producing Escherichia coli, Salmonella, and Nonpathogenic Escherichia coli Biotype I on Inoculated Prerigor Beef Surface Tissue.

Science.gov (United States)

Scott-Bullard, Britteny R; Geornaras, Ifigenia; Delmore, Robert J; Woerner, Dale R; Reagan, James O; Morgan, J Bred; Belk, Keith E

2017-12-01

A study was conducted to investigate the efficacy of a sulfuric acid-sodium sulfate blend (SSS) against Escherichia coli O157:H7, non-O157 Shiga toxin-producing E. coli (STEC), Salmonella, and nonpathogenic E. coli biotype I on prerigor beef surface tissue. The suitability of using the nonpathogenic E. coli as a surrogate for in-plant validation studies was also determined by comparing the data obtained for the nonpathogenic inoculum with those for the pathogenic inocula. Prerigor beef tissue samples (10 by 10 cm) were inoculated (ca. 6 log CFU/cm 2 ) on the adipose side in a laboratory-scale spray cabinet with multistrain mixtures of E. coli O157:H7 (5 strains), non-O157 STEC (12 strains), Salmonella (6 strains), or E. coli biotype I (5 strains). Treatment parameters evaluated were two SSS pH values (1.5 and 1.0) and two spray application pressures (13 and 22 lb/in 2 ). Untreated inoculated beef tissue samples served as controls for initial bacterial populations. Overall, the SSS treatments lowered inoculated (6.1 to 6.4 log CFU/cm 2 ) bacterial populations by 0.6 to 1.5 log CFU/cm 2 (P SSS was applied to samples inoculated with any of the tested E. coli inocula; however, solution pH did have a significant effect (P SSS was applied to samples inoculated with Salmonella. Results indicated that the response of the nonpathogenic E. coli inoculum to the SSS treatments was similar (P ≥ 0.05) to that of the pathogenic inocula tested, making the E. coli biotype I strains viable surrogate organisms for in-plant validation of SSS efficacy on beef. The application of SSS at the tested parameters to prerigor beef surface tissue may be an effective intervention for controlling pathogens in a commercial beef harvest process.

Colisão com o 'efeito estilingue'

OpenAIRE

Silveira,Fernando Lang da; Braun,Luci F.M; Braun,Thomas

2010-01-01

Abordamos teoricamente a colisão com o 'efeito estilingue' onde um corpo transfere momento linear e energia cinética para um segundo corpo de massa menor, fazendo com que a energia mecânica desse segundo corpo cresça de forma surpreendente. Mostramos que, mesmo quando as colisões são inelásticas, o ganho de energia mecânica pode ser grande. Apresentamos um estudo experimental do interessante efeito realizado a partir de um vídeo. Finalmente discutimos o 'efeito estilingue gravitacional' utili...

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...

Transmission of F4+ E. coli in groups of early weaned piglets

NARCIS (Netherlands)

Geenen, P.L.; Döpfer, D.; Meulen, van der J.; Jong, de M.C.M.

2005-01-01

The aim of this study was to estimate transmission parameters of enterotoxigenic F4+ Escherichia coli F4 (F4+ E. coli) in groups of early weaned piglets with F4-receptor-positive (F4R+) and F4-receptor-negative piglets (F4R[minus sign]). Transmission of F4+ E. coli was quantified in four

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...

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.

Prognostic value of low blood glucose at the presentation of E. coli bacteremia.

Science.gov (United States)

Alamgir, Shamsuddin; Volkova, Natalia B; Peterson, Michael W

2006-11-01

Septicemia is the tenth leading cause of death in the United States, and Escherichia coli is the most common isolate in blood cultures. Low blood glucose is a known complication of sepsis. The prognostic role of low blood glucose in E. coli bacteremia is unknown. The study's objective was to identify the incidence of low blood glucose at the presentation of E. coli bacteremia and determine its influence on prognosis and outcome. A retrospective cohort study was conducted in university-affiliated community hospitals. Subjects were consecutive patients diagnosed with E. coli bacteremia between 1997 and 2003. We identified 1060 patients with documented E. coli bacteremia. We excluded 105 patients who were younger than 18 years old or pregnant. We recorded demographic characteristics, discharge diagnosis, and outcome. Among the 955 patients with E. coli bacteremia, the average age was 64+/-19.4 years. Overall, 4.6% had documented low blood glucose (blood glucose <70 mg/dL) at presentation. The incidence of low blood glucose was the same in diabetic and nondiabetic patients. Patients with low blood glucose had a 4.7 times higher risk of death compared to patients with non-low blood glucose. Race, age, sex, and diabetes had no influence on survival. Gastrointestinal and genitourinary sources for E. coli bacteremia were more commonly associated with low blood glucose (P <.001). The study was limited to E. coli-positive blood cultures and to the one hospital system. Low blood glucose is present at the onset of E. coli bacteremia in 4.6% of patients. This represents a potentially large number of patients because E. coli is the most common blood culture isolate. Low blood glucose predicts poor outcome, especially in patients with abnormal hepatic and renal function. Low blood glucose should be considered an early clinical sign of E. coli bacteremia and aggressive therapy should be instituted to potentially save lives.

Genotypic characterization of ESBL-producing E. coli from imported meat in South Korea.

Science.gov (United States)

Kim, Young-Jo; Moon, Jin-San; Oh, Deog-Hwan; Chon, Jung-Whan; Song, Bo-Ra; Lim, Jong-Su; Heo, Eun-Jeong; Park, Hyun-Jung; Wee, Sung-Hwan; Sung, Kidon

2018-05-01

Twenty extended-spectrum β-lactamase (ESBL)-producing E. coli strains were isolated from imported meat in South Korea. ESBL strains of E. coli were detected in chicken (14/20) more often than in pork (6/20) and beef (0/20); the highest number (12/20) was detected in Brazilian meats. The bla CTX-M genes were predominant in meats from many countries. E. coli from pork imported from France produced the bla CTX-M-58 enzyme, which has never been documented previously in ESBL-producing bacteria from clinical or environmental sources. Additionally, the coexistence of the bla CTX-M-2 and bla OXA-1 enzymes in EC12-5 isolate was found for the first time in an ESBL E. coli isolate. A rare bla CTX-M type, bla CTX-M-25 , was found in 40% of ESBL E. coli isolates. Phenotypic susceptibility testing showed that E. coli isolates were resistant to up to eleven antibiotics, including ciprofloxacin. For the first time, a new combination in an integron gene cassette, aacA4-cmlA6-qacEΔ1, was found in an E. coli isolate from poultry imported from Brazil. Three E. coli ST117 isolates, from an avian pathogenic lineage producing CTX-M-94, harbored fimH, fyuA, iutA, papC, rfc, and traT virulence genes and were not susceptible to quinolones. For the first time, rfc and papG virulence factors were detected in ESBL E. coli strains isolated from meat products. Even though E. coli CC21 and CC22 were obtained from meats from the USA and Brazil, respectively, they had a similarity coefficient higher than 99% in rep-PCR and the same MLST type (ST117), phenotypic antibiotic resistance pattern, integron gene (qacEΔ1), and plasmid DNA profile. This study indicates that imported meat products may be a source of ESBL-producing E. coli strains in South Korea. Published by Elsevier Ltd.

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...

Resistência a antimicrobianos dependente do sistema de efluxo multidrogas em Escherichia coli isoladas de leite mastítico Antimicrobial resistance dependent on multidrugs efflux in Escherichia coli isolated from the mastitic milk

Directory of Open Access Journals (Sweden)

M.A.S. Moreira

2008-12-01

Full Text Available Identificaram-se e caracterizaram-se a resistência e a multirresistência aos principais antimicrobianos usados no tratamento de mastite bovina causada por Escherichia coli. A concentração inibitória mínima (MIC e o sistema de efluxo foram detectados pelas curvas de crescimento, com base na densidade óptica, em diferentes concentrações da droga e na presença e na ausência do desacoplador da força próton-motora (PMF. E. coli 1 foi resistente à neomicina e à gentamicina; E. coli 3 e 4, à tetraciclina e à estreptomicina; e E. coli 2 e 6 à gentamicina. E. coli 5 apresentou modelo de sensibilidade. Observou-se que MICs de todos os antimicrobianos dos multirresistentes (E. coli 1, 3 e 4 diminuíram na presença do desacoplador, o que sugere sistema de efluxo multidrogas. Após cura, apenas E. coli 1 apresentou modelo de sensibilidade, porém não houve alterações das MICs, antes e após adição do desacoplador. Os resultados indicam possível presença de mecanismo de resistência dependente da PMF codificado, ou parte dele, em plasmídeo.Resistance and multiresistance to main antimicrobials used for treating bovine mastitis caused by Escherichia coli were identified and characterized. The minimal inhibitory concentration (MIC and efflux systems were detected by the use of growth curves based on optical density at different drug concentrations and both presence and absence of uncoupler of the proton-motive force (PMF. E. coli 1 was resistant to neomycin and gentamycin, E. coli 3 and 4 were resistant to tetracycline and streptomycin, whereas E. coli 2 and 6 were resistant to gentamycin. E. coli 5 showed sensibility model. MICs of all antimicrobials of the multiresistant samples (E. coli 1, 3, and 4 were decreased in presence of the uncoupler, therefore suggesting the presence of the multidrug efflux system. After healing, only E. coli 1 showed sensibility model, however no alteration occurred in MIC(s before and after adding the

Radiosensitization of Escherichia coli and Salmonella typhi in presence of active compounds

International Nuclear Information System (INIS)

Lacroix, M.; Chiasson, F.; Borsa, J.; Ouattara, B.

2004-01-01

The radiosensitization of Escherichia coli and Salmonella typhi in ground beef was evaluated in the presence of 18 active compounds. Medium fat ground beef (23% fat) was inoculated with E. coli or S. typhi and each active compound was added separately at various concentrations. For E. coli, the most efficient compounds were trans-cinnamaldehyde, thymol and thyme. For S. typhi, the most efficient compounds was trans-cinnamaldehyde, carvacrol and thymol. The addition of tetrasodium pyrophosphate, carvacrol and ascorbic acid had no effect on the irradiation sensitivity of E. coli. For S. typhi, only ascorbic acid had no effect

Radiosensitization of Escherichia coli and Salmonella typhi in presence of active compounds

Energy Technology Data Exchange (ETDEWEB)

Lacroix, M. E-mail: monique.lacroix@inrs-iaf.uquebec.ca; Chiasson, F.; Borsa, J.; Ouattara, B

2004-10-01

The radiosensitization of Escherichia coli and Salmonella typhi in ground beef was evaluated in the presence of 18 active compounds. Medium fat ground beef (23% fat) was inoculated with E. coli or S. typhi and each active compound was added separately at various concentrations. For E. coli, the most efficient compounds were trans-cinnamaldehyde, thymol and thyme. For S. typhi, the most efficient compounds was trans-cinnamaldehyde, carvacrol and thymol. The addition of tetrasodium pyrophosphate, carvacrol and ascorbic acid had no effect on the irradiation sensitivity of E. coli. For S. typhi, only ascorbic acid had no effect.

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 coli pathogenic bacteria. Keyword : Leaves starfruit, E. Coli

Sex-related effects of nutritional supplementation of Escherichia coli: relevance to eating disorders.

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Tennoune, Naouel; Legrand, Romain; Ouelaa, Wassila; Breton, Jonathan; Lucas, Nicolas; Bole-Feysot, Christine; do Rego, Jean-Claude; Déchelotte, Pierre; Fetissov, Sergueï O

2015-03-01

The biological background of sex-related differences in the development of eating disorders (EDs) is unknown. Recent data showed that gut bacteria Escherichia coli induce autoantibodies against anorexigenic α-melanocyte-stimulating hormone (α-MSH) associated with psychopathology in ED. The aim of this study was to compare the effects of E. coli on feeding and autoantibodies against α-MSH and adrenocorticotropic hormone (ACTH), between female and male rats. Commensal E. coli K12 were given in a culture medium daily to adult Wistar rats by intragastric gavage over a 3-wk period; control rats received culture medium only. Before gavage, E. coli K12 DNA was detected in feces of female but not male rats. E. coli provision was accompanied by an increase in body weight gain in females, but a decrease in body weight gain and food intake in males. Independent of E. coli treatment, plasma levels of anti-α-MSH and ACTH immunoglobulin (Ig)G were higher in female than male rats. Females responded to E. coli by increasing α-MSH IgG levels and affinity, but males by increasing α-MSH IgM levels. Affinity of IgG for ACTH was increased in both E. coli-treated females and males, although with different kinetics. IgG from females stimulated more efficiently α-MSH-induced cyclic adenosine monophosphate production by melanocortin 4 receptor-expressing cells compared with IgG from males. Sex-related response to how E. coli affects feeding and anti-melanocortin hormone antibody production may depend on the presence of these bacteria in the gut before E. coli supplementation. These data suggest that sex-related presence of certain gut bacteria may represent a risk factor for ED development. Copyright © 2015 Elsevier Inc. All rights reserved.

The Impact of Media, Phylogenetic Classification, and E. coli Pathotypes on Biofilm Formation in Extraintestinal and Commensal E. coli From Humans and Animals

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Daniel W. Nielsen

2018-05-01

Full Text Available Extraintestinal pathogenic Escherichia coli (ExPEC include avian pathogenic E. coli (APEC, neonatal meningitis E. coli (NMEC, and uropathogenic E. coli (UPEC and are responsible for significant animal and human morbidity and mortality. This study sought to investigate if biofilm formation by ExPEC likely contributes to these losses since biofilms are associated with recurrent urinary tract infections, antibiotic resistance, and bacterial exchange of genetic material. Therefore, the goal of this study was to examine differences in biofilm formation among a collection of ExPEC and to ascertain if there is a relationship between their ability to produce biofilms and their assignment to phylogenetic groups in three media types – M63, diluted TSB, and BHI. Our results suggest that ExPEC produce relatively different levels of biofilm formation in the media tested as APEC (70.4%, p = 0.0064 and NMEC (84.4%, p = 0.0093 isolates were poor biofilm formers in minimal medium M63 while UPEC isolates produced significantly higher ODs under nutrient-limited conditions with 25% of strains producing strong biofilms in diluted TSB (p = 0.0204. Additionally, E. coli phylogenetic assignment using Clermont’s original and revised typing scheme demonstrated significant differences among the phylogenetic groups in the different media. When the original phylogenetic group isolates previously typed as group D were phylogenetically typed under the revised scheme and examined, they showed substantial variation in their ability to form biofilms, which may explain the significant values of revised phylogenetic groups E and F in M63 (p = 0.0291, p = 0.0024. Our data indicates that biofilm formation is correlated with phylogenetic classification and subpathotype or commensal grouping of E. coli strains.

Growth modeling of uropathogenic Escherichia coli in ground chicken meat

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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...

Synthesis and structural characterisation of iron(II) and copper(II) diphosphates containing flattened metal oxotetrahedra

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Keates, Adam C. [School of Chemistry, University of Southampton, Southampton, Hampshire SO17 1B,. UK (United Kingdom); Wang, Qianlong [Department of Chemistry, University of Bath, Claverton Down, Bath, BA2 7AY (United Kingdom); Weller, Mark T., E-mail: m.t.weller@bath.ac.uk [Department of Chemistry, University of Bath, Claverton Down, Bath, BA2 7AY (United Kingdom)

2014-02-15

Single crystal and bulk polycrystalline forms of K{sub 2}MP{sub 2}O{sub 7} (M=Fe(II), Cu(II)) have been synthesised and their structures determined from single crystal X-ray diffraction data. Both compounds crystallize in the tetragonal system, space group P-42{sub 1}m. Their structures are formed from infinite sheets of linked oxopolyhedra of the stoichiometry [MP{sub 2}O{sub 7}]{sup 2−} with potassium cations situated between the layers. The MO{sub 4} tetrahedra share oxygen atoms with [P{sub 2}O{sub 7}]{sup 4−} diphosphate groups and the potassium ions have KO{sub 8} square prismatic geometry. In both compounds the M(II) centre has an unusual strongly flattened, tetrahedral coordination to oxygen, as a result of the Jahn–Teller (JT) effect for the high spin d{sup 6} Fe(II) and p-orbital mixing or a second order JT effect for d{sup 9} Cu(II) centres in four fold coordination. The uncommon transition metal ion environments found in these materials are reflected in their optical absorption spectra and magnetism data. - Graphical abstract: The structures of the tetragonal polymorphs of K{sub 2}MP{sub 2}O{sub 7}, M=Cu(II), Fe(II), consist of infinite sheets of stoichiometry [MP{sub 2}O{sub 7}]{sup 2−}, formed from linked pyrophosphate groups and MO{sub 4} tetrahedra, separated by potassium ions. In both compounds the unusual tetrahedral coordination of the M(II) centre is strongly flattened as a result of Jahn–Teller (JT) effects for high spin, d{sup 6} Fe(II) and p-orbital mixing and second-order JT effects for d{sup 9} Cu(II). Display Omitted - Highlights: • Tetrahedral copper and iron(II) coordinated by oxygen. • New layered phosphate structure. • Jahn–Teller and d{sup 10} distorted coordinations.

Synthesis and structural characterisation of iron(II) and copper(II) diphosphates containing flattened metal oxotetrahedra

International Nuclear Information System (INIS)

Keates, Adam C.; Wang, Qianlong; Weller, Mark T.

2014-01-01

Single crystal and bulk polycrystalline forms of K 2 MP 2 O 7 (M=Fe(II), Cu(II)) have been synthesised and their structures determined from single crystal X-ray diffraction data. Both compounds crystallize in the tetragonal system, space group P-42 1 m. Their structures are formed from infinite sheets of linked oxopolyhedra of the stoichiometry [MP 2 O 7 ] 2− with potassium cations situated between the layers. The MO 4 tetrahedra share oxygen atoms with [P 2 O 7 ] 4− diphosphate groups and the potassium ions have KO 8 square prismatic geometry. In both compounds the M(II) centre has an unusual strongly flattened, tetrahedral coordination to oxygen, as a result of the Jahn–Teller (JT) effect for the high spin d 6 Fe(II) and p-orbital mixing or a second order JT effect for d 9 Cu(II) centres in four fold coordination. The uncommon transition metal ion environments found in these materials are reflected in their optical absorption spectra and magnetism data. - Graphical abstract: The structures of the tetragonal polymorphs of K 2 MP 2 O 7 , M=Cu(II), Fe(II), consist of infinite sheets of stoichiometry [MP 2 O 7 ] 2− , formed from linked pyrophosphate groups and MO 4 tetrahedra, separated by potassium ions. In both compounds the unusual tetrahedral coordination of the M(II) centre is strongly flattened as a result of Jahn–Teller (JT) effects for high spin, d 6 Fe(II) and p-orbital mixing and second-order JT effects for d 9 Cu(II). Display Omitted - Highlights: • Tetrahedral copper and iron(II) coordinated by oxygen. • New layered phosphate structure. • Jahn–Teller and d 10 distorted coordinations

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

African Journals Online (AJOL)

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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 ...

Neonatal Escherichia coli Bloodstream Infections: Clinical Outcomes and Impact of Initial Antibiotic Therapy.

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Bergin, Stephen P; Thaden, Joshua T; Ericson, Jessica E; Cross, Heather; Messina, Julia; Clark, Reese H; Fowler, Vance G; Benjamin, Daniel K; Hornik, Christoph P; Smith, P Brian

2015-09-01

Escherichia coli is a common cause of bloodstream infections (BSIs) in infants and is associated with high mortality and morbidity among survivors. The clinical significance of antibiotic resistance and timing of appropriate antimicrobial therapy in this population is poorly understood. We identified all infants with E. coli BSIs discharged from 77 neonatal intensive care units managed by the Pediatrix Medical Group in 2012. We used multivariable logistic regression to evaluate the association between 30-day mortality and ampicillin-resistant E. coli BSI, as well as the number of active empiric antimicrobial agents administered, controlling for gestational age, small-for-gestational age status, early-onset versus late-onset BSI, oxygen requirement, ventilator support and inotropic support on the day of the first positive blood culture. We identified 258 episodes of E. coli BSI, including 123 (48%) ampicillin-resistant isolates. Unadjusted 30-day mortality did not significantly differ between infants with ampicillin-resistant versus ampicillin-susceptible E. coli BSI [11 of 123 (9%) vs. 7 of 135 (5%); P = 0.33; adjusted odds ratio = 1.37 (95% confidence interval: 0.39, 4.77)]. Among ampicillin-resistant E. coli BSIs, 30-day mortality was not significantly lower for infants treated with at least one empiric antimicrobial active against ampicillin-resistant E. coli versus infants receiving no active empiric agent [adjusted odds ratio = 1.50 (0.07, 33.6)]. In this population of infants with E. coli BSI, ampicillin resistance was not associated with significantly increased mortality. Among the subset of infants with ampicillin-resistant E. coli, appropriate empirical antibiotic therapy was not associated with lower mortality.

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.

Detection of Enterohemorrhagic Escherichia coli Related Genes in E. coli Strains Belonging to B2 Phylogroup Isolated from Urinary Tract Infections in Combination with Antimicrobial Resistance Phenotypes

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Hamid Staji

2017-07-01

Full Text Available Background:  This study was conducted to detect the prevalence of EHEC virulence genes and antimicrobial resistance profile of Escherichia coli strains belonging to B2 phylogroup implicated in Urinary tract infections in Semnan, Iran.Methods:   From 240 urine samples 160 E. coli strains were isolated, biochemically. Then, E. coli isolates were examined by Multiplex-PCR for phylogenetic typing and detection of virulence genes (hly, stx1, stx2, eae associated with Enterohemorrhagic E. coli. Finally, Antimicrobial resistance of E. coli isolates were characterized using Disk Diffusion method.  Results:  From 160 E. coli isolates, 75 strains (47% were assigned to B2 phylogenetic group and prevalence of virulence genes were as follow: hly (21.3%, stx1 (16%, stx2 (10.6% and eae (6.7%, subsequently.  Phenotypic antimicrobial resistance of B2 isolates showed that all isolates were sensitive to Meropenem and Furazolidone and then highest frequency of resistance was observed to Streptomycin, Oxytetracycline, Neomycin, Nalidixic acid and Ampicillin (98.7% to 49.3%. Also low resistance prevalence was observed in case of Ceftizoxime, Lincospectin, Imipenem, Chloramphenicol and flurefenicole (16% to 1.3%.Conclusion:   The data suggest a high prevalence of antibiotic resistance in UPEC strains belonging to B2 phylogroup even for the antimicrobials using in pet and farm animals and their potential to cause EHEC specific clinical symptoms which may represent a serious health risk since these strains can be transmitted to GI tract and act as a reservoir for other uropathogenic E. coli and commensal strains.

High carriage of adherent invasive E. coli in wildlife and healthy individuals.

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Rahmouni, Oumaïra; Vignal, Cécile; Titécat, Marie; Foligné, Benoît; Pariente, Benjamin; Dubuquoy, Laurent; Desreumaux, Pierre; Neut, Christel

2018-01-01

Adherent invasive Escherichia coli (AIEC) are suspected to be involved in the pathogenesis of inflammatory bowel diseases. Since AIEC was first described in 1999, despite important progress on its genomic and immune characterizations, some crucial questions remain unanswered, such as whether there exists a natural reservoir, or whether there is asymptomatic carriage. The ECOR collection, including E. coli strains isolated mainly from the gut of healthy humans and animals, constitutes an ideal tool to investigate AIEC prevalence in healthy condition. A total of 61 E. coli strains were examined for characteristics of AIEC. The adhesion, invasion and intramacrophage replication capabilities (AIEC phenotype) of 61 intestinal E. coli strains were determined. The absence of virulence-associated diarrheagenic E. coli pathotypes (EPEC, ETEC, EIEC, EHEC, DAEC, EAEC), and uropathogenic E. coli was checked. Out of 61 intestinal strains, 13 (21%) exhibit the AIEC phenotype, 7 are from human origin and 6 are from animal origin. Prevalence of AIEC strains is about 24 and 19% in healthy humans and animals respectively. These strains are highly genetically diverse as they are distributed among the main described phylogroups. Among E. coli strains from the ECOR collection, we also detected strains able to detach I-407 cells. Our study described for the first time AIEC strains isolated from the feces of healthy humans and animals.

Cranberry extract inhibits in vitro adhesion of F4 and F18+Escherichia coli to pig intestinal epithelium and reduces in vivo excretion of pigs orally challenged with F18+ verotoxigenic E. coli.

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Coddens, Annelies; Loos, Michaela; Vanrompay, Daisy; Remon, Jean Paul; Cox, Eric

2017-04-01

F4 + E. coli and F18 + E. coli infections are an important threat for pig industry worldwide. Antibiotics are commonly used to treat infected piglets, but the emerging development of resistance against antibiotics raises major concerns. Hence, alternative therapies to prevent pigs from F4 + E. coli and F18 + E. coli infections need to be developed. Since cranberry previously showed anti-adhesive activity against uropathogenic E. coli, we aimed to investigate whether cranberry extract could also inhibit binding of F4 + E. coli and F18 + E. coli to pig intestinal epithelium. Using the in vitro villus adhesion assay, we found that low concentrations of cranberry extract (20μg or 100μg/ml) have strong inhibitory activity on F4 + E. coli (75.3%, S.D.=9.31 or 95.8%, S.D.=2.56, respectively) and F18 + E. coli adherence (100% inhibition). This effect was not due to antimicrobial activity. Moreover, cranberry extract (10mg or 100mg) could also abolish in vivo binding of F4 and F18 fimbriae to the pig intestinal epithelium in ligated loop experiments. Finally, two challenge experiments with F18 + E. coli were performed to address the efficacy of in-feed or water supplemented cranberry extract. No effect could be observed in piglets that received cranberry extract only in feed (1g/kg or 10g/kg). However, supplementation of feed (10g/kg) and drinking water (1g/L) significantly decreased excretion and diarrhea. The decreased infection resulted in a decreased serum antibody response indicating reduced exposure to F18 + E. coli. Copyright © 2017 Elsevier B.V. All rights reserved.

Pneumatosis Coli Mimicking Colorectal Cancer

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Teresa Jacob

2014-01-01

Full Text Available Pneumatosis coli (PC is a rare condition of the gastrointestinal tract involving extraluminal gas confined within the bowel wall. We report the case of a 40-year-old gentleman presenting clinically and endoscopically with suspected colorectal cancer. In light of the patient’s red flag symptoms, and carpet of polyps seen endoscopically, surgical management by an anterior resection was performed with the patient making a successful recovery. Histological analysis of the resected specimen confirmed pneumatosis coli with no evidence of colonic neoplasia. Although PC can be an incidental finding in asymptomatic patients and considered a benign condition, it can also present as a life-threatening emergency with bowel necrosis and obstruction requiring emergency surgical intervention. Also, when PC mimics malignancy, surgical management is the most appropriate step to ensure that the diagnosis of cancer is not missed.

Effectiveness of sanitizing agents in inactivating Escherichia coli (ATCC 25922 in food cutting board surfaces. Removal E. coli using different sanitizers

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CEZAR AUGUSTO BELTRAME

2016-03-01

Full Text Available The objective of this study was to investigate Escherichia coli adhesion on new and used polyethylene cutting board surface and evaluate it’s removal using different sanitizer (peracetic acid,chlorhexidine, sodium hypochlorite and organic acids. Results indicated that the number of adherent cells increased with time in both surfaces evaluated. Evaluating the sanitizer action, 0.5%peracetic acid was more effective in removal E. coli than chlorhexidine and organic acids at same concentration in both surfaces. Peracetic acid and sodium hypochlorite also showed effectiveness at concentrations of 0.2% and 0.5% on new surfaces, respectively. 0.8% of chlorhexidine and 2.0% of organic acids showed similar effectiveness in the removal E. coli on new and used surfaces, respectively.These results suggest that peracetic acid is considerable promise sanitizer for application in surfaces of the food processing industry.

Escherichia coli O26 IN RAW BUFFALO MILK: PRELIMINARY RESULTS

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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.

Secretion of clostridium cellulase by E. coli

Science.gov (United States)

Yu, Ida Kuo

1998-01-01

A gene, encoding an endocellulase from a newly isolated mesophilic Clostridium strain IY-2 which can digest bamboo fibers, cellulose, rice straw, and sawdust, was isolated by shotgun cloning in an E. coli expression plasmid pLC2833. E. coli positive clones were selected based on their ability to hydrolyze milled bamboo fibers and cellulose present in agar plates. One clone contained a 2.8 kb DNA fragment that was responsible for cellulase activity. Western blot analyses indicated that the positive clone produced a secreted cellulase with a mass of about 58,000 daltons that was identical in size to the subunit of one of the three major Clostridium cellulases. The products of cellulose digestion by this cloned cellulase were cellotetraose and soluble higher polymers. The cloned DNA contained signal sequences capable of directing the secretion of heterologous proteins from an E. coli host. The invention describes a bioprocess for the treatment of cellulosic plant materials to produce cellular growth substrates and fermentation end products suitable for production of liquid fuels, solvents, and acids.

Patotipos de Escherichia coli causadores de diarreia em bezerros: uma atualização

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Fernanda M. Coura

2014-09-01

Full Text Available A diarreia é uma das doenças mais frequentes de bezerros com até 30 dias de idade e é uma importante causa de perdas econômicas. Sua etiologia é complexa e envolve a interação de diversos fatores infecciosos, nutricionais, imunológicos, gerenciais e ambientais. Os principais sinais clínicos são a diarreia, desidratação progressiva, acidose metabólica, desequilíbrio de eletrólitos e balanço energético negativo com ou sem hipoglicemia, que se não tratados, levam à morte do animal. Escherichia coli se destaca como um importante enteropatógeno envolvido na síndrome diarreica. Cepas de E. coli patogênicas são classificadas em grupos ou patotipos, de acordo com a produção de fatores de virulência e mecanismos pelos quais causam doença. Já foram identificados cinco patotipos de E. coli associados à diarreia em bezerros: E. coli enterotoxigênica (ETEC, E. coli enteropatogênica (EPEC, E. coli enterohemorrágica (EHEC, E. coli produtora de toxina Shiga (STEC e E. coli necrotoxigênica (NTEC. Nesse artigo apresentamos as principais características e os atuais conhecimentos sobre os patotipos de E. coli causadores de diarreia em bezerros.

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

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Jennifer A. Pienaar

2016-05-01

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

Antimicrobial Resistant Pattern of Escherichia Coli Strains Isolated from Pediatric Patients in Jordan

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