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Sample records for coli produces tailor-made

  1. Recombinant Escherichia coli produces tailor-made biopolyester granules for applications in fluorescence activated cell sorting: functional display of the mouse interleukin-2 and myelin oligodendrocyte glycoprotein

    PubMed Central

    Bäckström, B Thomas; Brockelbank, Jane A; Rehm, Bernd HA

    2007-01-01

    Background Fluorescence activated cell sorting (FACS) is a powerful technique for the qualitative and quantitative detection of biomolecules used widely in both basic research and clinical diagnostic applications. Beads displaying a specific antigen are used to bind antibodies which are then fluorescently labelled using secondary antibodies. As the individual suspension bead passes through the sensing region of the FACS machine, fluorescent signals are acquired and analysed. Currently, antigens are tediously purified and chemically cross-linked to preformed beads. Purification and coupling of proteins often renders them inactive and they will not be displayed in its native configuration. As an alternative, we genetically engineered Escherichia coli to produce biopolyester (polyhdroxyalkanoate=PHA) granules displaying diagnostically relevant antigens in their native conformation and suitable for FACS analysis. Results Hybrid genes were constructed, which encode either the mouse interleukin-2 (IL2) or the myelin oligodendrocyte glycoprotein (MOG) fused via an enterokinase site providing linker region to the C terminus of the PHA granule associated protein PhaP, respectively. The hybrid genes were expressed in PHA-accumulating recombinant E. coli. MOG and IL2 fusion proteins were abundantly attached to PHA granules and were identified by MALDI-TOF/MS analysis and N terminal sequencing. A more abundant second fusion protein of either MOG or IL2 resulted from an additional N terminal fusion, which did surprisingly not interfere with attachment to PHA granule. PHA granules displaying either IL2 or MOG were used for FACS using monoclonal anti-IL2 or anti-MOG antibodies conjugated to a fluorescent dye. FACS analysis showed significant and specific binding of respective antibodies. Enterokinase treatment of IL2 displaying PHA granules enabled removal of IL2 as monitored by FACS analysis. Mice were immunized with either MOG or OVA (ovalbumin) and the respective sera were

  2. Overproduction of Thermus sp. Strain T2 β-Galactosidase in Escherichia coli and Preparation by Using Tailor-Made Metal Chelate Supports

    PubMed Central

    Pessela, Benevides C. C.; Vian, Alejandro; Mateo, César; Fernández-Lafuente, Roberto; García, José L.; Guisán, José M.; Carrascosa, Alfonso V.

    2003-01-01

    A novel thermostable chimeric β-galactosidase was constructed by fusing a poly-His tag to the N-terminal region of the β-galactosidase from Thermus sp. strain T2 to facilitate its overexpression in Escherichia coli and its purification by immobilized metal-ion affinity chromatography (IMAC). The poly-His tag fusion did not affect the activation, kinetic parameters, and stability of the β-galactosidase. Copper-iminodiacetic acid (Cu-IDA) supports enabled the most rapid adsorption of the His-tagged enzyme, favoring multisubunit interactions, but caused deleterious effects on the enzyme stability. To improve the enzyme purification a selective one-point adsorption was achieved by designing tailor-made low-activated Co-IDA or Ni-IDA supports. The new enzyme was not only useful for industrial purposes but also has become an excellent model to study the purification of large multimeric proteins via selective adsorption on tailor-made IMAC supports. PMID:12676671

  3. Tailor Made Strategies of Dissemination: The Story and Theory Connection.

    ERIC Educational Resources Information Center

    Bhola, H. S.

    General principles of planning tailor-made change strategies arise directly from the CLER (configurations, linkages, environments, resources) model, a model of planned change, as well as from the value assumptions in which the model is embedded. These principles include the following: change requires commitment, planning and management of change…

  4. Tailor-made force fields for crystal-structure prediction.

    PubMed

    Neumann, Marcus A

    2008-08-14

    A general procedure is presented to derive a complete set of force-field parameters for flexible molecules in the crystalline state on a case-by-case basis. The force-field parameters are fitted to the electrostatic potential as well as to accurate energies and forces generated by means of a hybrid method that combines solid-state density functional theory (DFT) calculations with an empirical van der Waals correction. All DFT calculations are carried out with the VASP program. The mathematical structure of the force field, the generation of reference data, the choice of the figure of merit, the optimization algorithm, and the parameter-refinement strategy are discussed in detail. The approach is applied to cyclohexane-1,4-dione, a small flexible ring. The tailor-made force field obtained for cyclohexane-1,4-dione is used to search for low-energy crystal packings in all 230 space groups with one molecule per asymmetric unit, and the most stable crystal structures are reoptimized in a second step with the hybrid method. The experimental crystal structure is found as the most stable predicted crystal structure both with the tailor-made force field and the hybrid method. The same methodology has also been applied successfully to the four compounds of the fourth CCDC blind test on crystal-structure prediction. For the five aforementioned compounds, the root-mean-square deviations between lattice energies calculated with the tailor-made force fields and the hybrid method range from 0.024 to 0.053 kcal/mol per atom around an average value of 0.034 kcal/mol per atom.

  5. Tailor-made force fields for crystal-structure prediction.

    PubMed

    Neumann, Marcus A

    2008-08-14

    A general procedure is presented to derive a complete set of force-field parameters for flexible molecules in the crystalline state on a case-by-case basis. The force-field parameters are fitted to the electrostatic potential as well as to accurate energies and forces generated by means of a hybrid method that combines solid-state density functional theory (DFT) calculations with an empirical van der Waals correction. All DFT calculations are carried out with the VASP program. The mathematical structure of the force field, the generation of reference data, the choice of the figure of merit, the optimization algorithm, and the parameter-refinement strategy are discussed in detail. The approach is applied to cyclohexane-1,4-dione, a small flexible ring. The tailor-made force field obtained for cyclohexane-1,4-dione is used to search for low-energy crystal packings in all 230 space groups with one molecule per asymmetric unit, and the most stable crystal structures are reoptimized in a second step with the hybrid method. The experimental crystal structure is found as the most stable predicted crystal structure both with the tailor-made force field and the hybrid method. The same methodology has also been applied successfully to the four compounds of the fourth CCDC blind test on crystal-structure prediction. For the five aforementioned compounds, the root-mean-square deviations between lattice energies calculated with the tailor-made force fields and the hybrid method range from 0.024 to 0.053 kcal/mol per atom around an average value of 0.034 kcal/mol per atom. PMID:18642947

  6. Shiga Toxin Producing Escherichia coli.

    PubMed

    Bryan, Allen; Youngster, Ilan; McAdam, Alexander J

    2015-06-01

    Shiga toxin-producing Escherichia coli (STEC) is among the common causes of foodborne gastroenteritis. STEC is defined by the production of specific toxins, but within this pathotype there is a diverse group of organisms. This diversity has important consequences for understanding the pathogenesis of the organism, as well as for selecting the optimum strategy for diagnostic testing in the clinical laboratory. This review includes discussions of the mechanisms of pathogenesis, the range of manifestations of infection, and the several different methods of laboratory detection of Shiga toxin-producing E coli.

  7. Tailor-made polyamide membranes for water desalination.

    PubMed

    Choi, Wansuk; Gu, Joung-Eun; Park, Sang-Hee; Kim, Seyong; Bang, Joona; Baek, Kyung-Youl; Park, Byoungnam; Lee, Jong Suk; Chan, Edwin P; Lee, Jung-Hyun

    2015-01-27

    Independent control of the extrinsic and intrinsic properties of the polyamide (PA) selective layer is essential for designing thin-film composite (TFC) membranes with performance characteristics required for water purification applications besides seawater desalination. Current commercial TFC membranes fabricated via the well-established interfacial polymerization (IP) approach yield materials that are far from ideal because their layer thickness, surface roughness, polymer chemistry, and network structure cannot be separately tailored. In this work, tailor-made PA-based desalination membranes based on molecular layer-by-layer (mLbL) assembly are presented. The mLbL technique enables the construction of an ultrathin and highly cross-linked PA selective layer in a precisely and independently controlled manner. The mLbL-assembled TFC membranes exhibit significant enhancements in performance compared to their IP-assembled counterparts. A maximum sodium chloride rejection of 98.2% is achieved along with over 2.5 times higher water flux than the IP-assembled counterpart. More importantly, this work demonstrates the broad applicability of mLbL in fabricating a variety of PA-based TFC membranes with nanoscale control of the selective layer thickness and roughness independent of the specific polyamide chemistry.

  8. Tailor-made polyamide membranes for water desalination.

    PubMed

    Choi, Wansuk; Gu, Joung-Eun; Park, Sang-Hee; Kim, Seyong; Bang, Joona; Baek, Kyung-Youl; Park, Byoungnam; Lee, Jong Suk; Chan, Edwin P; Lee, Jung-Hyun

    2015-01-27

    Independent control of the extrinsic and intrinsic properties of the polyamide (PA) selective layer is essential for designing thin-film composite (TFC) membranes with performance characteristics required for water purification applications besides seawater desalination. Current commercial TFC membranes fabricated via the well-established interfacial polymerization (IP) approach yield materials that are far from ideal because their layer thickness, surface roughness, polymer chemistry, and network structure cannot be separately tailored. In this work, tailor-made PA-based desalination membranes based on molecular layer-by-layer (mLbL) assembly are presented. The mLbL technique enables the construction of an ultrathin and highly cross-linked PA selective layer in a precisely and independently controlled manner. The mLbL-assembled TFC membranes exhibit significant enhancements in performance compared to their IP-assembled counterparts. A maximum sodium chloride rejection of 98.2% is achieved along with over 2.5 times higher water flux than the IP-assembled counterpart. More importantly, this work demonstrates the broad applicability of mLbL in fabricating a variety of PA-based TFC membranes with nanoscale control of the selective layer thickness and roughness independent of the specific polyamide chemistry. PMID:25548959

  9. Characterization of GaSb thin films from tailor-made single-source precursors

    NASA Astrophysics Data System (ADS)

    Seemayer, Andreas; Hommes, Alexander; Hümann, Sascha; Schulz, Stephan; Wandelt, Klaus

    2008-11-01

    We investigated the growth and surface properties of GaSb thin films on a Si(0 0 1) substrate prepared using a tailor-made fully alkyl-substituted heterocyclic single-source precursor. The precursor properties were monitored during the evaporation process by residual gas analysis (RGA). The initial film growth was monitored by Auger electron spectroscopy (AES). Using a high-vacuum cold wall reactor, dense GaSb films could be produced and were characterized by AES, AFM and synchrotron X-ray photoelectron spectroscopy (S-XPS). The results are discussed in terms of a correlation of the electronic and geometrical properties with the composition and structure of the films.

  10. Gasifier feed - Tailor-made from Illinois coals

    SciTech Connect

    Ehrlinger, H.P. III ); Lytle, J.; Frost, R.R.; Lizzio, A.; Kohlenberger, L.; Brewer, K. DESTEC Energy Williams Technology, Illinois Coal Association )

    1992-01-01

    The main purpose of this project is to produce a feedstock from preparation plant fines from an illinois coal that is ideal for a slurry fed, slagging, entrained-flow coal gasifier. The high sulfur content and high Btu value of Illinois coals are particularly advantageous in such a gasifier; preliminary calculations indicate that the increased cost of removing sulfur from the gas from a high sulfur coal is more than offset by the increased revenue from the sale of the elemental sulfur; additionally the high Btu Illinois coal concentrates more energy into the slurry of a given coal to water ratio. The Btu is higher not only because of the higher Btu value of the coal but also because Illinois coal requires less water to produce a pumpable slurry than western coal, i.e., as little as 30--35% water may be used for Illinois coal as compared to approximately 45% for most western coals.

  11. Gasifier feed - Tailor-made from Illinois coals

    SciTech Connect

    Ehrlinger, H.P. III ); Lytle, J.; Frost, R.R.; Lizzio, A.; Kohlenberger, L.; Brewer, K. DESTEC Energy Williams Technology Illinois Coal Association )

    1992-01-01

    The main purpose of this project is to produce a feedstock from preparation plant fines from an Illinois coal that is ideal for a slurry fed, slagging, entrained-flow coal gasifier. The high sulfur content and high Btu value of Illinois coals are particularly advantageous in such a gasifier; preliminary calculations indicate that the increased cost of removing sulfur from the gas from a high sulfur coal is more than offset by the increased revenue from the sale of the elemental sulfur; additionally the high Btu Illinois coal concentrates more energy into the slurry of a given coal to water ratio. This project will bring the expertise of four organizations together to perform the various tasks. The Illinois Coal Association will help direct the project to be the most beneficial to the Illinois coal industry. DESTEC Energy, a wholly-owned subsidiary of Dow Chemical Company, will provide guidelines and test compatibility of the slurries developed for gasification feedstock. Williams Technology will provide their expertise in long distance slurry pumping, and test selected products for viscosity, pumpability, and handlability. The Illinois State Geological Survey will study methods for producing clean coal/water slurries from preparation plant wastes including the concentration of pyritic sulfur into the coal slurry to increase the revenue from elemental sulfur produced during gasification operations, and decrease the pyritic sulfur content of the waste streams. ISGS will also test the gasification reactivity of the coals. As reported earlier, a variety of possible samples of coal have been analyzed and the gasification performance evaluation reported. Additionally, commercial sized samples of -28 mesh {times} 100 mesh coal -100 {times} 0 coal were subjected to pumpability testing. Neither the coarse product nor the fine product by themselves proved to be good candidates for trouble free pumping, but the mix of the two proved to be a very acceptable product

  12. Advanced bacterial polyhydroxyalkanoates: towards a versatile and sustainable platform for unnatural tailor-made polyesters.

    PubMed

    Park, Si Jae; Kim, Tae Wan; Kim, Min Kyung; Lee, Sang Yup; Lim, Sung-Chul

    2012-01-01

    Polyhydroxyalkanoates (PHAs) are biopolyesters that generally consist of 3-, 4-, 5-, and 6-hydroxycarboxylic acids, which are accumulated as carbon and energy storage materials in many bacteria in limited growth conditions with excess carbon sources. Due to the diverse substrate specificities of PHA synthases, the key enzymes for PHA biosynthesis, PHAs with different material properties have been synthesized by incorporating different monomer components with differing compositions. Also, engineering PHA synthases using in vitro-directed evolution and site-directed mutagenesis facilitates the synthesis of PHA copolymers with novel material properties by broadening the spectrum of monomers available for PHA biosynthesis. Based on the understanding of metabolism of PHA biosynthesis, recombinant bacteria have been engineered to produce different types of PHAs by expressing heterologous PHA biosynthesis genes, and by creating and enhancing the metabolic pathways to efficiently generate precursors for PHA monomers. Recently, the PHA biosynthesis system has been expanded to produce unnatural biopolyesters containing 2-hydroxyacid monomers such as glycolate, lactate, and 2-hydroxybutyrate by employing natural and engineered PHA synthases. Using this system, polylactic acid (PLA), one of the major commercially-available bioplastics, can be synthesized from renewable resources by direct fermentation of recombinant bacteria. In this review, we discuss recent advances in the development of the PHA biosynthesis system as a platform for tailor-made polyesters with novel material properties. PMID:22137963

  13. Candida bombicola as a platform organism for the production of tailor-made biomolecules.

    PubMed

    Roelants, Sophie L K W; Saerens, Karen M J; Derycke, Thibaut; Li, Bing; Lin, Yao-Cheng; Van de Peer, Yves; De Maeseneire, Sofie L; Van Bogaert, Inge N A; Soetaert, Wim

    2013-09-01

    The yeast Candida bombicola is capable of producing high amounts (400 g/L) of the biosurfactant sophorolipids. The genetic makeup of this industrially important yeast has recently been uncovered and molecular manipulation techniques have been developed. Hence, all tools for the development of new bioprocesses with C. bombicola are now available. As a proof of concept, the production of two totally different molecules was aimed for: the bioplastic polyhydroxyalkanoate (PHA) and a new-to-nature cellobioselipid-biosurfactant. Integration of the new functionalities at genomic loci necessary for sophorolipid production safeguards the new biomolecules from sophorolipid contamination, while taking advantage of the regulation of the sophorolipid gene cluster. A maximum yield of 2.0% wt/dwt PHA was obtained; furthermore, this is the first time cellobioselipid synthesis by a non-natural producer is reported. We here provided proof of concept that C. bombicola can be transformed into a platform organism for the production of tailor-made biomolecules. PMID:23475585

  14. Verocytotoxin-producing Escherichia coli (VTEC).

    PubMed

    Karmali, Mohamed A; Gannon, Victor; Sargeant, Jan M

    2010-01-27

    Escherichia coli O157:H7 and other Verocytotoxin-producing E. coli (VTEC) are zoonotic pathogens associated with food and waterborne illness around the world. E. coli O157:H7 has been implicated in large outbreaks as well as in sporadic cases of haemorrhagic colitis and the sometimes fatal haemolytic uremic syndrome. VTs produced by these bacteria are thought to damage host endothelial cells in small vessels of the intestine, kidney and brain resulting in thrombotic microangiopathy. All VTs have the same subunit structure, glycolipid cell receptor and inhibit protein synthesis. During VTEC infection, it is thought one or more bacterial adhesins initiates colonization and establishes intimate attachment and is responsible for the translocation of a variety of effectors which alter the structure and function of host cells. VTEC are widespread in animals but ruminants are thought to be their natural reservoir. E. coli O157:H7 colonizes the terminal colon of cattle and can be shed in very large numbers by specific herdmates known as "supershedders". Faeces containing these organisms act as a source of contamination for a variety of foods and the environment. Many VTEC control efforts have been investigated along the "farm to fork" continuum including, vaccination of cattle with colonization factors, and the use of novel antimicrobials, such as bacteriocins, chloral hydrate, bacteriophage and substances which disrupt quorum sensing. In addition, many barriers have been developed for use in the slaughter and food processing industry such as steam pasteurization and irradiation. Despite these efforts many scientific, technical and regulatory challenges remain in the control and prevention of VTEC-associated human illness.

  15. Development of Thermophilic Tailor-Made Enzyme Mixtures for the Bioconversion of Agricultural and Forest Residues

    PubMed Central

    Karnaouri, Anthi; Matsakas, Leonidas; Topakas, Evangelos; Rova, Ulrika; Christakopoulos, Paul

    2016-01-01

    Even though the main components of all lignocellulosic feedstocks include cellulose, hemicellulose, as well as the protective lignin matrix, there are some differences in structure, such as in hardwoods and softwoods, which may influence the degradability of the materials. Under this view, various types of biomass might require a minimal set of enzymes that has to be tailor-made. Partially defined complex mixtures that are currently commercially used are not adapted to efficiently degrade different materials, so novel enzyme mixtures have to be customized. Development of these cocktails requires better knowledge about the specific activities involved, in order to optimize hydrolysis. The role of filamentous fungus Myceliophthora thermophila and its complete enzymatic repertoire for the bioconversion of complex carbohydrates has been widely proven. In this study, four core cellulases (MtCBH7, MtCBH6, MtEG5, and MtEG7), in the presence of other four “accessory” enzymes (mannanase, lytic polyssacharide monooxygenase MtGH61, xylanase, MtFae1a) and β-glucosidase MtBGL3, were tested as a nine-component cocktail against one model substrate (phosphoric acid swollen cellulose) and four hydrothermally pretreated natural substrates (wheat straw as an agricultural waste, birch, and spruce biomass, as forest residues). Synergistic interactions among different enzymes were determined using a suitable design of experiments methodology. The results suggest that for the hydrolysis of the pure substrate (PASC), high proportions of MtEG7 are needed for efficient yields. MtCBH7 and MtEG7 are enzymes of major importance during the hydrolysis of pretreated wheat straw, while MtCBH7 plays a crucial role in case of spruce. Cellobiohydrolases MtCBH6 and MtCBH7 act in combination and are key enzymes for the hydrolysis of the hardwood (birch). Optimum combinations were predicted from suitable statistical models which were able to further increase hydrolysis yields, suggesting that

  16. Exploitation of desilylation chemistry in tailor-made functionalization on diverse surfaces

    PubMed Central

    Fu, Yongchun; Chen, Songjie; Kuzume, Akiyoshi; Rudnev, Alexander; Huang, Cancan; Kaliginedi, Veerabhadrarao; Baghernejad, Masoud; Hong, Wenjing; Wandlowski, Thomas; Decurtins, Silvio; Liu, Shi-Xia

    2015-01-01

    Interface engineering to attain a uniform and compact self-assembled monolayer at atomically flat surfaces plays a crucial role in the bottom-up fabrication of organic molecular devices. Here we report a promising and operationally simple approach for modification/functionalization not only at ultraflat single-crystal metal surfaces, M(111) (M=Au, Pt, Pd, Rh and Ir) but also at the highly oriented pyrolytic graphite surface, upon efficient in situ cleavage of trimethylsilyl end groups of the molecules. The obtained self-assembled monolayers are ultrastable within a wide potential window. The carbon–surface bonding on various substrates is confirmed by shell-isolated nanoparticle-enhanced Raman spectroscopy. Application of this strategy in tuning surface wettability is also demonstrated. The most valuable finding is that a combination of the desilylation with the click chemistry represents an efficient method for covalent and tailor-made functionalization of diverse surfaces. PMID:25758661

  17. myADS-arXiv -- a Tailor-made, Open Access, Virtual Journal

    NASA Astrophysics Data System (ADS)

    Henneken, E.; Kurtz, M. J.; Eichhorn, G.; Accomazzi, A.; Grant, C. S.; Thompson, D.; Bohlen, E.; Murray, S. S.

    2007-10-01

    The myADS-arXiv service provides the scientific community with a one stop shop for staying up-to-date with a researcher's field of interest. The service provides a powerful and unique filter on the enormous amount of bibliographic information added to the ADS on a daily basis. It also provides a complete view of the most relevant papers available in the subscriber's field of interest. With this service, the subscriber will get to know the latest developments, popular trends and the most important papers. This makes the service not only unique from a technical point of view, but also from a content point of view. On this poster we will argue why myADS-arXiv is a tailor-made, open access, virtual journal and we will illustrate its unique character.

  18. Tailor-made TALEN system for highly efficient targeted gene replacement in the rice blast fungus.

    PubMed

    Arazoe, Takayuki; Ogawa, Tetsuo; Miyoshi, Kennosuke; Yamato, Tohru; Ohsato, Shuichi; Sakuma, Tetsushi; Yamamoto, Takashi; Arie, Tsutomu; Kuwata, Shigeru

    2015-07-01

    Genetic manipulation is key to unraveling gene functions and creating genetically modified strains of microbial organisms. Recently, engineered nucleases that can generate DNA double-strand breaks (DSBs) at a specific site in the desired locus within genome are utilized in a rapidly developing genome editing technology via DSBs repair. However, the use of engineered nucleases in filamentous fungi has not been validated. In this study, we demonstrated that tailor-made transcriptional activator-like effector nucleases (TALENs) system, Platinum-Fungal TALENs (PtFg TALENs), could improve the efficiency of homologous recombination-mediated targeted gene replacement by up to 100% in the rice blast fungus Pyricularia oryzae. This high-efficiency PtFg TALEN has great potential for basic and applied biological applications in filamentous fungi. PMID:25683503

  19. Achieving population dispersal through tailor-made community planning: an Israeli experiment in the Galilee region.

    PubMed

    Carmon, N

    1994-04-01

    The author describes a program designed to encourage population dispersal that was carried out in the central Galilee region of Israel during the 1980s. The program involved setting up 52 small communities with the appropriate infrastructures to attract young, well-educated settlers. "The plan succeeded in attracting the desired type of population...to the region, and the newcomers viewed the new communities as their permanent homes. Based on this experience and on the analysis of relevant literature, a development strategy of tailor-made community planning is hereby recommended for future projects. It is especially appropriate in the context of developed countries with a slow to zero population growth and with spreading social norms of the postindustrial society."

  20. Shiga toxin-producing Escherichia coli

    PubMed Central

    Etcheverría, Analía Inés; Padola, Nora Lía

    2013-01-01

    Shiga toxin-producing Escherichia coli (STEC) cause hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS) in humans. Outbreaks are linked to bovine food sources. STEC O157:H7 has been responsible for the most severe outbreaks worldwide. However, non-O157 serotypes have emerged as important enteric pathogens in several countries. The main virulence factor of STEC is the production of Shiga toxins 1 and 2. Additional virulence markers are a plasmid-encoded enterohemolysin (ehxA), an autoagglutinating adhesin (Saa), a catalase-peroxidase (katP), an extracellular serine protease (espP), a zinc metalloprotease (stcE), a subtilase cytotoxin (subAB), among others. Other virulence factors are intimin and adhesins that had a roll in the adherence of STEC to bovine colon. This review focuses on the virulence traits of STEC and especially on those related to the adhesion to bovine colon. The known of the interaction between STEC and the bovine host is crucial to develop strategies to control cattle colonization. PMID:23624795

  1. Shiga toxin-producing Escherichia coli.

    PubMed

    Smith, James L; Fratamico, Pina M; Gunther, Nereus W

    2014-01-01

    In the United States, it is estimated that non-O157 Shiga toxin-producing Escherichia coli (STEC) cause more illnesses than STEC O157:H7, and the majority of cases of non-O157 STEC infections are due to serogroups O26, O45, O103, O111, O121, and O145, referred to as the top six non-O157 STEC. The diseases caused by non-O157 STEC are generally milder than those induced by O157 STEC; nonetheless, non-O157 STEC strains have also been associated with serious illnesses such as hemorrhagic colitis and hemolytic uremic syndrome, as well as death. Ruminants, particularly cattle, are reservoirs for both O157 and non-O157 STEC, which are transmitted to humans by person-to-person or animal contact and by ingestion of food or water contaminated with animal feces. Improved strategies to control STEC colonization and shedding in cattle and contamination of meat and produce are needed. In general, non-O157 STEC respond to stresses such as acid, heat, and other stresses induced during food preparation similar to O157 STEC. Similar to O157:H7, the top six non-O157 STEC are classified as adulterants in beef by the USDA Food Safety and Inspection Service, and regulatory testing for these pathogens began in June 2012. Due to the genetic and phenotypic variability of non-O157 STEC strains, the development of accurate and reliable methods for detection and isolation of these pathogens has been challenging. Since the non-O157 STEC are responsible for a large portion of STEC-related illnesses, more extensive studies on their physiology, genetics, pathogenicity, and evolution are needed in order to develop more effective control strategies.

  2. Dry particle coating of polymer particles for tailor-made product properties

    SciTech Connect

    Blümel, C. Schmidt, J. Dielesen, A. Sachs, M. Winzer, B. Peukert, W. Wirth, K.-E.

    2014-05-15

    Disperse polymer powders with tailor-made particle properties are of increasing interest in industrial applications such as Selective Laser Beam Melting processes (SLM). This study focuses on dry particle coating processes to improve the conductivity of the insulating polymer powder in order to assemble conductive devices. Therefore PP particles were coated with Carbon Black nanoparticles in a dry particle coating process. This process was investigated in dependence of process time and mass fraction of Carbon Black. The conductivity of the functionalized powders was measured by impedance spectroscopy. It was found that there is a dependence of process time, respectively coating ratio and conductivity. The powder shows higher conductivities with increasing number of guest particles per host particle surface area, i.e. there is a correlation between surface functionalization density and conductivity. The assembled composite particles open new possibilities for processing distinct polymers such as PP in SLM process. The fundamentals of the dry particle coating process of PP host particles with Carbon Black guest particles as well as the influence on the electrical conductivity will be discussed.

  3. Short and Intense Tailor-Made Notched Music Training against Tinnitus: The Tinnitus Frequency Matters

    PubMed Central

    Teismann, Henning; Okamoto, Hidehiko; Pantev, Christo

    2011-01-01

    Tinnitus is one of the most common diseases in industrialized countries. Here, we developed and evaluated a short-term (5 subsequent days) and intensive (6 hours/day) tailor-made notched music training (TMNMT) for patients suffering from chronic, tonal tinnitus. We evaluated (i) the TMNMT efficacy in terms of behavioral and magnetoencephalographic outcome measures for two matched patient groups with either low (≤8 kHz, N = 10) or high (>8 kHz, N = 10) tinnitus frequencies, and the (ii) persistency of the TMNMT effects over the course of a four weeks post-training phase. The results indicated that the short-term intensive TMNMT took effect in patients with tinnitus frequencies ≤8 kHz: subjective tinnitus loudness, tinnitus-related distress, and tinnitus-related auditory cortex evoked activity were significantly reduced after TMNMT completion. However, in the patients with tinnitus frequencies >8 kHz, significant changes were not observed. Interpreted in their entirety, the results also indicated that the induced changes in auditory cortex evoked neuronal activity and tinnitus loudness were not persistent, encouraging the application of the TMNMT as a longer-term training. The findings are essential in guiding the intended transfer of this neuro-scientific treatment approach into routine clinical practice. PMID:21935438

  4. Tailor-made asymmetric PVDF hollow fibers for soluble gas removal

    SciTech Connect

    Li, K.; Kong, J.F.; Wang, D.; Teo, W.K.

    1999-06-01

    Tailor-made polyvinylidene fluoride (PVDF) asymmetric hollow-fiber membranes and their membrane modules were employed for soluble gas removal, such as H{sub 2}S from waste gas streams. This study focused on the techniques of fabricating and characterizing the PVDF asymmetric hollow-fiber membranes and their membrane modules for removal of H{sub 2}S using an aqueous solution containing 10% NaOH. A laminar parabolic velocity profile was used to characterize the flow of the H{sub 2}S gas mixture in the hollow-fiber lumen. Effects of operating conditions and the morphological structures of the membranes on the membrane`s coefficient, k{sub AM}, were examined both theoretically and experimentally. The capabilities of the hollow-fiber membranes developed for removal of H{sub 2}S from waste gas streams were evaluated and compared with conventional symmetric hydrophobic hollow-fiber membranes, such as polypropylene. An analysis of H{sub 2}S transfer across the more developed PVDF membranes reveals that the membrane`s coefficient, k{sub AM}, evaluated from its structure parameters, such as the effective surface porosity and mean radius, agreed well with the experimental data obtained from absorption experiments.

  5. Dry particle coating of polymer particles for tailor-made product properties

    NASA Astrophysics Data System (ADS)

    Blümel, C.; Schmidt, J.; Dielesen, A.; Sachs, M.; Winzer, B.; Peukert, W.; Wirth, K.-E.

    2014-05-01

    Disperse polymer powders with tailor-made particle properties are of increasing interest in industrial applications such as Selective Laser Beam Melting processes (SLM). This study focuses on dry particle coating processes to improve the conductivity of the insulating polymer powder in order to assemble conductive devices. Therefore PP particles were coated with Carbon Black nanoparticles in a dry particle coating process. This process was investigated in dependence of process time and mass fraction of Carbon Black. The conductivity of the functionalized powders was measured by impedance spectroscopy. It was found that there is a dependence of process time, respectively coating ratio and conductivity. The powder shows higher conductivities with increasing number of guest particles per host particle surface area, i.e. there is a correlation between surface functionalization density and conductivity. The assembled composite particles open new possibilities for processing distinct polymers such as PP in SLM process. The fundamentals of the dry particle coating process of PP host particles with Carbon Black guest particles as well as the influence on the electrical conductivity will be discussed.

  6. Infection by verocytotoxin-producing Escherichia coli.

    PubMed Central

    Karmali, M A

    1989-01-01

    Verocytotoxin (VT)-producing Escherichia coli (VTEC) are a newly recognized group of enteric pathogens which are increasingly being recognized as common causes of diarrhea in some geographic settings. Outbreak studies indicate that most patients with VTEC infection develop mild uncomplicated diarrhea. However, a significant risk of two serious and potentially life-threatening complications, hemorrhagic colitis and the hemolytic uremic syndrome, makes VTEC infection a public health problem of serious concern. The main reservoirs of VTEC appear to be the intestinal tracts of animals, and foods of animal (especially bovine) origin are probably the principal sources for human infection. The term VT refers to a family of subunit exotoxins with high biological activity. Individual VTEC strains elaborate one or both of at least two serologically distinct, bacteriophage-mediated VTs (VT1 and VT2) which are closely related to Shiga toxin and are thus also referred to as Shiga-like toxins. The holotoxins bind to cells, via their B subunits, to a specific receptor which is probably the glycolipid, globotriosyl ceramide (Gb3). Binding is followed by internalization of the A subunit, which, after it is proteolytically nicked and reduced to the A1 fragment, inhibits protein synthesis in mammalian cells by inactivating 60S ribosomal subunits through selective structural modification of 28S ribosomal ribonucleic acid. The mechanism of VTEC diarrhea is still controversial, and the relative roles of locally acting VT and "attaching and effacing adherence" of VTEC to the mucosa have yet to be resolved. There is increasing evidence that hemolytic uremic syndrome and possibly hemorrhagic colitis result from the systemic action of VT on vascular endothelial cells. The role of antitoxic immunity in preventing the systemic complications of VTEC infection is being explored. Antibiotics appear to be contraindicated in the treatment of VTEC infection. The most common VTEC serotype associated

  7. Insights into heterogeneous atmospheric oxidation chemistry through vibrational sum frequency generation studies of tailor-made model systems

    NASA Astrophysics Data System (ADS)

    Voges, Andrea Beth

    2006-04-01

    As surfaces are known to have profound implications for chemical transport, reactivity, and energy budgets in atmospheric environments, we have designed a nonlinear optical sum frequency generation (SFG) system that can be used to study atmospherically relevant heterogeneous chemistry. The aims of this project were two-fold: first, to develop and characterize tailor-made organically coated substrates relevant to tropospheric chemistry and second, carry out kinetic studies that model naturally occurring heterogeneous atmospheric reactions. After construction of the broadband SFG system, we studied siloxane substrates functionalized with organic adlayers. The organic adlayers were specifically chosen to contain environmentally relevant functional groups, namely, an acid-terminated alkyl chain, several ester functionalized alkyl chains, and a non-functionalized alkyl chain. Hydrolysis of methyl ester functionalized surfaces was carried out to produce carboxylic acid functionalized surfaces and monitored using SFG. In order to access more complicated atmospherically relevant substrates, we then focused on the synthesis and characterization of a derivative of limonene, a biogenically emitted compound, chemically bound to a glass surface. We employed both electrophilic, and nucleophilic linker chemistries to increase the versatility of our approach. SFG spectra indicated that while orientation of the surface-bound terpenes depended on the linker strategy we employed, the C=C double bond was accessible to gas phase ozone regardless of the strategy applied. We then used SFG to track the interaction of a terpene-linked species with ozone and calculate reaction probabilities. Exposure of the amide-linked terpene substrate to ppm levels of ozone at a total pressure of 1 atm and 300 K yielded an initial reaction probability of approximately 1 x 10 -5, which was significantly higher than the corresponding gas phase reaction involving 1-methyl-1-cyclohexene. The chemical

  8. Tailor-made biopolymers porous scaffold fabrication for tissue engineering: application of radiant energy in the form of microwave under vacuum.

    PubMed

    Jaya, S; Durance, T D

    2008-01-01

    Many methods are available for developing three-dimensional porous scaffolds using various polymeric materials for tissue-engineering applications. Each has its own advantages and disadvantages. Some of the available methods and their limitations were discussed briefly. This paper focuses on the scope of novel technology called radiant energy application under vacuum for the fabrication of three-dimensional porous scaffolds for tissue engineering applications. Radiant energy application in the form of microwave under vacuum has been shown to develop and maintain the porous structure in fruits and vegetables after dehydration, which produced the microstructure similar to the freeze dried materials. Same principle of applying radiant energy under vacuum was used on the biopolymeric gels to create tailor-made, porous scaffolds for biomedical purposes. It has many advantages over the other existing methods of scaffold fabrication. This paper also reviews the scaffolds design recently fabricated by the authors using radiant energy under vacuum.

  9. Gasifier feed - Tailor-made from Illinois coals. Technical report, December 1, 1991--February 29, 1992

    SciTech Connect

    Ehrlinger, H.P. III; Lytle, J.; Frost, R.R.; Lizzio, A.; Kohlenberger, L.; Brewer, K. |||

    1992-08-01

    The main purpose of this project is to produce a feedstock from preparation plant fines from an illinois coal that is ideal for a slurry fed, slagging, entrained-flow coal gasifier. The high sulfur content and high Btu value of Illinois coals are particularly advantageous in such a gasifier; preliminary calculations indicate that the increased cost of removing sulfur from the gas from a high sulfur coal is more than offset by the increased revenue from the sale of the elemental sulfur; additionally the high Btu Illinois coal concentrates more energy into the slurry of a given coal to water ratio. The Btu is higher not only because of the higher Btu value of the coal but also because Illinois coal requires less water to produce a pumpable slurry than western coal, i.e., as little as 30--35% water may be used for Illinois coal as compared to approximately 45% for most western coals.

  10. Playing and listening to tailor-made notched music: cortical plasticity induced by unimodal and multimodal training in tinnitus patients.

    PubMed

    Pape, Janna; Paraskevopoulos, Evangelos; Bruchmann, Maximilian; Wollbrink, Andreas; Rudack, Claudia; Pantev, Christo

    2014-01-01

    BACKGROUND. The generation and maintenance of tinnitus are assumed to be based on maladaptive functional cortical reorganization. Listening to modified music, which contains no energy in the range of the individual tinnitus frequency, can inhibit the corresponding neuronal activity in the auditory cortex. Music making has been shown to be a powerful stimulator for brain plasticity, inducing changes in multiple sensory systems. Using magnetoencephalographic (MEG) and behavioral measurements we evaluated the cortical plasticity effects of two months of (a) active listening to (unisensory) versus (b) learning to play (multisensory) tailor-made notched music in nonmusician tinnitus patients. Taking into account the fact that uni- and multisensory trainings induce different patterns of cortical plasticity we hypothesized that these two protocols will have different affects. RESULTS. Only the active listening (unisensory) group showed significant reduction of tinnitus related activity of the middle temporal cortex and an increase in the activity of a tinnitus-coping related posterior parietal area. CONCLUSIONS. These findings indicate that active listening to tailor-made notched music induces greater neuroplastic changes in the maladaptively reorganized cortical network of tinnitus patients while additional integration of other sensory modalities during training reduces these neuroplastic effects.

  11. Playing and Listening to Tailor-Made Notched Music: Cortical Plasticity Induced by Unimodal and Multimodal Training in Tinnitus Patients

    PubMed Central

    Rudack, Claudia

    2014-01-01

    Background. The generation and maintenance of tinnitus are assumed to be based on maladaptive functional cortical reorganization. Listening to modified music, which contains no energy in the range of the individual tinnitus frequency, can inhibit the corresponding neuronal activity in the auditory cortex. Music making has been shown to be a powerful stimulator for brain plasticity, inducing changes in multiple sensory systems. Using magnetoencephalographic (MEG) and behavioral measurements we evaluated the cortical plasticity effects of two months of (a) active listening to (unisensory) versus (b) learning to play (multisensory) tailor-made notched music in nonmusician tinnitus patients. Taking into account the fact that uni- and multisensory trainings induce different patterns of cortical plasticity we hypothesized that these two protocols will have different affects. Results. Only the active listening (unisensory) group showed significant reduction of tinnitus related activity of the middle temporal cortex and an increase in the activity of a tinnitus-coping related posterior parietal area. Conclusions. These findings indicate that active listening to tailor-made notched music induces greater neuroplastic changes in the maladaptively reorganized cortical network of tinnitus patients while additional integration of other sensory modalities during training reduces these neuroplastic effects. PMID:24895541

  12. Tailor-made rehabilitation approach using multiple types of hybrid assistive limb robots for acute stroke patients: A pilot study.

    PubMed

    Fukuda, Hiroyuki; Morishita, Takashi; Ogata, Toshiyasu; Saita, Kazuya; Hyakutake, Koichi; Watanabe, Junko; Shiota, Etsuji; Inoue, Tooru

    2016-01-01

    This article investigated the feasibility of a tailor-made neurorehabilitation approach using multiple types of hybrid assistive limb (HAL) robots for acute stroke patients. We investigated the clinical outcomes of patients who underwent rehabilitation using the HAL robots. The Brunnstrom stage, Barthel index (BI), and functional independence measure (FIM) were evaluated at baseline and when patients were transferred to a rehabilitation facility. Scores were compared between the multiple-robot rehabilitation and single-robot rehabilitation groups. Nine hemiplegic acute stroke patients (five men and four women; mean age 59.4 ± 12.5 years; four hemorrhagic stroke and five ischemic stroke) underwent rehabilitation using multiple types of HAL robots for 19.4 ± 12.5 days, and 14 patients (six men and eight women; mean age 63.2 ± 13.9 years; nine hemorrhagic stroke and five ischemic stroke) underwent rehabilitation using a single type of HAL robot for 14.9 ± 8.9 days. The multiple-robot rehabilitation group showed significantly better outcomes in the Brunnstrom stage of the upper extremity, BI, and FIM scores. To the best of the authors' knowledge, this is the first pilot study demonstrating the feasibility of rehabilitation using multiple exoskeleton robots. The tailor-made rehabilitation approach may be useful for the treatment of acute stroke.

  13. Gasifier feed - Tailor-made from Illinois coals. [Quarterly] report, March 1, 1992--May 31, 1992

    SciTech Connect

    Ehrlinger, H.P. III; Lytle, J.; Frost, R.R.; Lizzio, A.; Kohlenberger, L.; Brewer, K. |||

    1992-10-01

    The main purpose of this project is to produce a feedstock from preparation plant fines from an Illinois coal that is ideal for a slurry fed, slagging, entrained-flow coal gasifier. The high sulfur content and high Btu value of Illinois coals are particularly advantageous in such a gasifier; preliminary calculations indicate that the increased cost of removing sulfur from the gas from a high sulfur coal is more than offset by the increased revenue from the sale of the elemental sulfur; additionally the high Btu Illinois coal concentrates more energy into the slurry of a given coal to water ratio. This project will bring the expertise of four organizations together to perform the various tasks. The Illinois Coal Association will help direct the project to be the most beneficial to the Illinois coal industry. DESTEC Energy, a wholly-owned subsidiary of Dow Chemical Company, will provide guidelines and test compatibility of the slurries developed for gasification feedstock. Williams Technology will provide their expertise in long distance slurry pumping, and test selected products for viscosity, pumpability, and handlability. The Illinois State Geological Survey will study methods for producing clean coal/water slurries from preparation plant wastes including the concentration of pyritic sulfur into the coal slurry to increase the revenue from elemental sulfur produced during gasification operations, and decrease the pyritic sulfur content of the waste streams. ISGS will also test the gasification reactivity of the coals. As reported earlier, a variety of possible samples of coal have been analyzed and the gasification performance evaluation reported. Additionally, commercial sized samples of -28 mesh {times} 100 mesh coal -100 {times} 0 coal were subjected to pumpability testing. Neither the coarse product nor the fine product by themselves proved to be good candidates for trouble free pumping, but the mix of the two proved to be a very acceptable product

  14. Non-O157 Shiga toxin-producing Escherichia coli: detection and characterization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Escherichia coli strains that produce Shiga toxins, referred to as Shiga toxin-producing E. coli (STEC) or verotoxigenic E. coli (VTEC) are important food-borne pathogens that cause hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). E. coli O157:H7 is a common cause of STEC infection; ho...

  15. Non-O157 Shiga Toxin-Producing E. coli Associated with Muscle Foods

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Escherichia coli strains that produce Shiga toxins, referred to as Shiga toxin-producing E. coli (STEC) or verotoxigenic E. coli (VTEC), cause hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). E. coli O157:H7 is the most common cause of STEC infection; however, numerous non-O157 STECs b...

  16. Shiga Toxin (Verotoxin)-Producing Escherichia coli in Japan.

    PubMed

    Terajima, Jun; Iyoda, Sunao; Ohnishi, Makoto; Watanabe, Haruo

    2014-10-01

    A series of outbreaks of infection with Shiga toxin (verocytotoxin)-producing Escherichia coli or enterohemorrhagic E. coli (EHEC) O157:H7 occurred in Japan in 1996, the largest outbreak occurring in primary schools in Sakai City, Osaka Prefecture, where more than 7,500 cases were reported. Although the reason for the sudden increase in the number of reports of EHEC isolates in 1996 is not known, the number of reports has grown to more than 3,000 cases per year since 1996, from an average of 105 reports each year during the previous 5-year period (1991-1995). Despite control measures instituted since 1996, including designating Shiga toxin-producing E. coli infection as a notifiable disease, and nationwide surveillance effectively monitoring the disease, the number of reports remains high, around 3,800 cases per year. Serogroup O157 predominates over other EHEC serogroups, but isolation frequency of non-O157 EHEC has gone up slightly over the past few years. Non-O157 EHEC has recently caused outbreaks where consumption of a raw beef dish was the source of the infection, and some fatal cases occurred. Laboratory surveillance comprised prefectural and municipal public health institutes, and the National Institute of Infectious Diseases has contributed to finding not only multiprefectural outbreaks but recognizing sporadic cases that could have been missed as an outbreak without the aid of molecular subtyping of EHEC isolates. This short overview presents recent information on the surveillance of EHEC infections in Japan. PMID:26104366

  17. Metabolic engineering of Escherichia coli to produce zeaxanthin.

    PubMed

    Li, Xi-Ran; Tian, Gui-Qiao; Shen, Hong-Jie; Liu, Jian-Zhong

    2015-04-01

    Zeaxanthin is a high-value carotenoid that is used in nutraceuticals, cosmetics, food, and animal feed industries. Zeaxanthin is chemically synthesized or purified from microorganisms as a natural product; however, increasing demand requires development of alternative sources such as heterologous biosynthesis by recombinant bacteria. For this purpose, we molecularly engineered Escherichia coli to optimize the synthesis of zeaxanthin from lycopene using fusion protein-mediated substrate channeling as well as by the introduction of tunable intergenic regions. The tunable intergenic regions approach was more efficient compared with protein fusion for coordinating expression of lycopene β-cyclase gene crtY and β-carotene 3-hydroxylase gene crtZ. The influence of the substrate channeling effect suggests that the reaction catalyzed by CrtZ is the rate-limiting step in zeaxanthin biosynthesis. Then Pantoea ananatis, Pantoea agglomerans and Haematococcus pluvialis crtZ were compared. Because P. ananatis crtZ is superior to that of P. agglomerans or H. pluvialis for zeaxanthin production, we used it to generate a recombinant strain of E. coli BETA-1 containing pZSPBA-2(P37-crtZPAN) that produced higher amounts of zeaxanthin (11.95 ± 0.21 mg/g dry cell weight) than other engineered E. coli strains described in the literature.

  18. Metabolic engineering of Escherichia coli to produce zeaxanthin.

    PubMed

    Li, Xi-Ran; Tian, Gui-Qiao; Shen, Hong-Jie; Liu, Jian-Zhong

    2015-04-01

    Zeaxanthin is a high-value carotenoid that is used in nutraceuticals, cosmetics, food, and animal feed industries. Zeaxanthin is chemically synthesized or purified from microorganisms as a natural product; however, increasing demand requires development of alternative sources such as heterologous biosynthesis by recombinant bacteria. For this purpose, we molecularly engineered Escherichia coli to optimize the synthesis of zeaxanthin from lycopene using fusion protein-mediated substrate channeling as well as by the introduction of tunable intergenic regions. The tunable intergenic regions approach was more efficient compared with protein fusion for coordinating expression of lycopene β-cyclase gene crtY and β-carotene 3-hydroxylase gene crtZ. The influence of the substrate channeling effect suggests that the reaction catalyzed by CrtZ is the rate-limiting step in zeaxanthin biosynthesis. Then Pantoea ananatis, Pantoea agglomerans and Haematococcus pluvialis crtZ were compared. Because P. ananatis crtZ is superior to that of P. agglomerans or H. pluvialis for zeaxanthin production, we used it to generate a recombinant strain of E. coli BETA-1 containing pZSPBA-2(P37-crtZPAN) that produced higher amounts of zeaxanthin (11.95 ± 0.21 mg/g dry cell weight) than other engineered E. coli strains described in the literature. PMID:25533633

  19. Characterisation of the thermoluminescence (TL) properties of tailor-made Ge-doped silica glass fibre for applications in medical radiation therapy dosimetry

    NASA Astrophysics Data System (ADS)

    Zahaimi, N. A.; Zin, H.; Mahdiraji, G. A.; Rahman, A. L. Abdul; Bradley, D. A.; Rahman, A. T. Abdul

    2014-11-01

    We have investigated the characterisation of new fabricated material Ge doped silica glass thermoluminescence TL dosimeter (Photonic Research Centre, University of Malaya) for medical radiation dosimetry at therapy energy. Previously, the dosimeter has been studied to provide ideal dosimetry system, suitable to ensure an accurate delivery of radiation doses to tumour tissue while minimising the amount of radiation administrated to healthy tissue. Both energies of photon and electron were used in this experiment for a dose range of 1 to 5 Gy. The various sizes of core diameter Ge doped silica glass (120, 241, 362, 483 and 604 μm) were exposed by using linear accelerator at Pantai Medical Centre. For both energies, the optical fibres were found to produce a flat response to a fixed photon and electron doses to within 4% (S.D) of the mean of the TL distribution. In terms of dose response, the fibres provide linear response over the range investigated, from a fraction of 1-5 Gy. The finding shows 120 μm fibres have 1.82 greater dose response than 604 pm fibres irradiated at 6 MV photon with a fixed dose of 3 Gy. While for electron energy 12 MeV, the response shows 120 μm fibres have 1.58 greater dose response compared to 604 μm fibres. The good responses are suitable to make these tailor-made doped silica fibres a promising TL material for use as a dosimetric system in medical radiation therapy.

  20. Beta-Lactamase Producing Escherichia coli Isolates in Imported and Locally Produced Chicken Meat from Ghana.

    PubMed

    Rasmussen, Mette Marie; Opintan, Japheth A; Frimodt-Møller, Niels; Styrishave, Bjarne

    2015-01-01

    The use of antibiotics in food animals is of public health concern, because resistant zoonotic pathogens can be transmitted to humans. Furthermore, global trade with food may rapidly spread multi-resistant pathogens between countries and even continents. The purpose of the study was to investigate whether imported chicken meat and meat from locally reared chicken are potential sources for human exposure to multi resistant Escherichia coli isolates. 188 samples from imported and locally produced chicken meat were sampled and analyzed. 153 bacteria isolates were successfully cultured and identified as E. coli using MALDI-ToF. Of these 109 isolates were from meat whereas the remaining 44 were isolated from the cloaca of locally reared live chickens. Antimicrobial susceptibility test was done on the identified E. coli isolates. Additionally, beta-lactamases production (ESBL and/or AmpC) were phenotypically confirmed on all isolates showing resistance to cefpodoxime. Beta-lactamase producing (BLP) E. coli meat isolates were further genotyped. Antimicrobial resistance to four antibiotic markers with highest resistance was detected more frequently in isolates from local chickens compared to imported chickens (tetracycline 88.9% vs. 57.5%, sulphonamide 75.0% vs. 46.6%, ampicillin 69.4% vs. 61.6% and trimethoprim 66.7% vs. 38.4%). Beta-lactamase production was found in 29 E. coli meat isolates, with 56.9% of them being multiple drug resistant (≥ 3). The predominant phylogroup identified was B1 followed by A and D, with similar distribution among the isolates from meat of locally reared chickens and imported chickens. Beta-lactamase producing genotype blaCTX-M-15 (50%; 10/20) was the most frequently drug resistant gene detected. More BLP E. coli isolates were found in imported chicken meat compared to locally reared chickens, demonstrating that these isolates may be spreading through food trade. In conclusion, both imported and locally produced chicken meats are potential

  1. Beta-Lactamase Producing Escherichia coli Isolates in Imported and Locally Produced Chicken Meat from Ghana

    PubMed Central

    Rasmussen, Mette Marie; Opintan, Japheth A.; Frimodt-Møller, Niels; Styrishave, Bjarne

    2015-01-01

    The use of antibiotics in food animals is of public health concern, because resistant zoonotic pathogens can be transmitted to humans. Furthermore, global trade with food may rapidly spread multi-resistant pathogens between countries and even continents. The purpose of the study was to investigate whether imported chicken meat and meat from locally reared chicken are potential sources for human exposure to multi resistant Escherichia coli isolates. 188 samples from imported and locally produced chicken meat were sampled and analyzed. 153 bacteria isolates were successfully cultured and identified as E. coli using MALDI-ToF. Of these 109 isolates were from meat whereas the remaining 44 were isolated from the cloaca of locally reared live chickens. Antimicrobial susceptibility test was done on the identified E. coli isolates. Additionally, beta-lactamases production (ESBL and/or AmpC) were phenotypically confirmed on all isolates showing resistance to cefpodoxime. Beta-lactamase producing (BLP) E. coli meat isolates were further genotyped. Antimicrobial resistance to four antibiotic markers with highest resistance was detected more frequently in isolates from local chickens compared to imported chickens (tetracycline 88.9% vs. 57.5%, sulphonamide 75.0% vs. 46.6%, ampicillin 69.4% vs. 61.6% and trimethoprim 66.7% vs. 38.4%). Beta-lactamase production was found in 29 E. coli meat isolates, with 56.9% of them being multiple drug resistant (≥ 3). The predominant phylogroup identified was B1 followed by A and D, with similar distribution among the isolates from meat of locally reared chickens and imported chickens. Beta-lactamase producing genotype blaCTX-M-15 (50%; 10/20) was the most frequently drug resistant gene detected. More BLP E. coli isolates were found in imported chicken meat compared to locally reared chickens, demonstrating that these isolates may be spreading through food trade. In conclusion, both imported and locally produced chicken meats are potential

  2. Beneficial knockouts in Escherichia coli for producing hydrogen from glycerol.

    PubMed

    Tran, Kien Trung; Maeda, Toshinari; Sanchez-Torres, Viviana; Wood, Thomas K

    2015-03-01

    Glycerol is an inexpensive and abundant source for biofuel production on a large scale. Escherichia coli is a robust bacterium for producing hydrogen; however, its hydrogen productivity from glycerol is low. In this study, we conducted random transposon mutagenesis to identify uncharacterized genes whose inactivation is beneficial for hydrogen production from glycerol. Through screening, four mutant strains were found that are able to have from 1.3- to 1.6-fold higher hydrogen productivity (μmol H2/mg protein) than that of their parent strain (p < 0.05). These mutations were identified as aroM, gatZ, ycgR, and yfgI. The hydrogen yield (mol H2/mol glycerol consumed) of the aroM, gatZ, ycgR, and yfgI strains was 1.7-, 1.4-, 2.4-, and 2.1-fold higher than that of their parent strain, respectively. Moreover, a single disruption in these genes resulted in a faster cell growth and glycerol consumption under anaerobic conditions. In E. coli, AroM is predicted to be involved in the shikimate pathway, GatZ is tagatose-1,6-bisphosphate aldolase 2 which converts dihydroxyacetone phosphate to 1,6-biphosphate, and YcgR acts as a molecular brake limiting the swimming speed and ATP consumption. So far, the function of YfgI in general and in hydrogen production in particular remains unknown.

  3. Impact of Spectral Notch Width on Neurophysiological Plasticity and Clinical Effectiveness of the Tailor-Made Notched Music Training.

    PubMed

    Wunderlich, Robert; Lau, Pia; Stein, Alwina; Engell, Alva; Wollbrink, Andreas; Rudack, Claudia; Pantev, Christo

    2015-01-01

    Tinnitus, the ringing in the ears that is unrelated to any external source, causes a significant loss in quality of life, involving sleep disturbance and depression for 1 to 3% of the general population. While in the first place tinnitus may be triggered by damage to the inner ear cells, the neural generators of subjective tinnitus are located in central regions of the nervous system. A loss of lateral inhibition, tonotopical reorganization and a gain-increase in response to the sensory deprivation result in hypersensitivity and hyperactivity in certain regions of the auditory cortex. In the tailor-made notched music training (TMNMT) patients listen to music from which the frequency spectrum of the tinnitus has been removed. This evokes strong lateral inhibition from neurons tuned to adjacent frequencies onto the neurons involved in the tinnitus percept. A reduction of tinnitus loudness and tinnitus-related neural activity was achieved with TMNMT in previous studies. As the effect of lateral inhibition depends on the bandwidth of the notch, in the current study we altered the notch width to find the most effective notch width for TMNMT. We compared 1-octave notch width with ½-octave and ¼-octave. Participants chose their favorite music for the training that included three month of two hours daily listening. The outcome was measured by means of standardized questionnaires and magnetoencephalography. We found a general reduction of tinnitus distress in all administered tinnitus questionnaires after the training. Additionally, tinnitus-related neural activity was reduced after the training. Nevertheless, notch width did not have an influence on the behavioral or neural effects of TMNMT. This could be due to a non-linear resolution of lateral inhibition in high frequencies. PMID:26406446

  4. Enhancing Inhibition-Induced Plasticity in Tinnitus – Spectral Energy Contrasts in Tailor-Made Notched Music Matter

    PubMed Central

    Stein, Alwina; Engell, Alva; Lau, Pia; Wunderlich, Robert; Junghoefer, Markus; Wollbrink, Andreas; Bruchmann, Maximilian; Rudack, Claudia; Pantev, Christo

    2015-01-01

    Chronic tinnitus seems to be caused by reduced inhibition among frequency selective neurons in the auditory cortex. One possibility to reduce tinnitus perception is to induce inhibition onto over-activated neurons representing the tinnitus frequency via tailor-made notched music (TMNM). Since lateral inhibition is modifiable by spectral energy contrasts, the question arises if the effects of inhibition-induced plasticity can be enhanced by introducing increased spectral energy contrasts (ISEC) in TMNM. Eighteen participants suffering from chronic tonal tinnitus, pseudo randomly assigned to either a classical TMNM or an ISEC-TMNM group, listened to notched music for three hours on three consecutive days. The music was filtered for both groups by introducing a notch filter centered at the individual tinnitus frequency. For the ISEC-TMNM group a frequency bandwidth of 3/8 octaves on each side of the notch was amplified, additionally, by about 20 dB. Before and after each music exposure, participants rated their subjectively perceived tinnitus loudness on a visual analog scale. During the magnetoencephalographic recordings, participants were stimulated with either a reference tone of 500 Hz or a test tone with a carrier frequency representing the individual tinnitus pitch. Perceived tinnitus loudness was significantly reduced after TMNM exposure, though TMNM type did not influence the loudness ratings. Tinnitus related neural activity in the N1m time window and in the so called tinnitus network comprising temporal, parietal and frontal regions was reduced after TMNM exposure. The ISEC-TMNM group revealed even enhanced inhibition-induced plasticity in a temporal and a frontal cortical area. Overall, inhibition of tinnitus related neural activity could be strengthened in people affected with tinnitus by increasing spectral energy contrast in TMNM, confirming the concepts of inhibition-induced plasticity via TMNM and spectral energy contrasts. PMID:25951605

  5. Impact of Spectral Notch Width on Neurophysiological Plasticity and Clinical Effectiveness of the Tailor-Made Notched Music Training

    PubMed Central

    Wunderlich, Robert; Lau, Pia; Stein, Alwina; Engell, Alva; Wollbrink, Andreas; Rudack, Claudia; Pantev, Christo

    2015-01-01

    Tinnitus, the ringing in the ears that is unrelated to any external source, causes a significant loss in quality of life, involving sleep disturbance and depression for 1 to 3% of the general population. While in the first place tinnitus may be triggered by damage to the inner ear cells, the neural generators of subjective tinnitus are located in central regions of the nervous system. A loss of lateral inhibition, tonotopical reorganization and a gain-increase in response to the sensory deprivation result in hypersensitivity and hyperactivity in certain regions of the auditory cortex. In the tailor-made notched music training (TMNMT) patients listen to music from which the frequency spectrum of the tinnitus has been removed. This evokes strong lateral inhibition from neurons tuned to adjacent frequencies onto the neurons involved in the tinnitus percept. A reduction of tinnitus loudness and tinnitus-related neural activity was achieved with TMNMT in previous studies. As the effect of lateral inhibition depends on the bandwidth of the notch, in the current study we altered the notch width to find the most effective notch width for TMNMT. We compared 1-octave notch width with ½-octave and ¼-octave. Participants chose their favorite music for the training that included three month of two hours daily listening. The outcome was measured by means of standardized questionnaires and magnetoencephalography. We found a general reduction of tinnitus distress in all administered tinnitus questionnaires after the training. Additionally, tinnitus-related neural activity was reduced after the training. Nevertheless, notch width did not have an influence on the behavioral or neural effects of TMNMT. This could be due to a non-linear resolution of lateral inhibition in high frequencies. PMID:26406446

  6. Impact of Spectral Notch Width on Neurophysiological Plasticity and Clinical Effectiveness of the Tailor-Made Notched Music Training.

    PubMed

    Wunderlich, Robert; Lau, Pia; Stein, Alwina; Engell, Alva; Wollbrink, Andreas; Rudack, Claudia; Pantev, Christo

    2015-01-01

    Tinnitus, the ringing in the ears that is unrelated to any external source, causes a significant loss in quality of life, involving sleep disturbance and depression for 1 to 3% of the general population. While in the first place tinnitus may be triggered by damage to the inner ear cells, the neural generators of subjective tinnitus are located in central regions of the nervous system. A loss of lateral inhibition, tonotopical reorganization and a gain-increase in response to the sensory deprivation result in hypersensitivity and hyperactivity in certain regions of the auditory cortex. In the tailor-made notched music training (TMNMT) patients listen to music from which the frequency spectrum of the tinnitus has been removed. This evokes strong lateral inhibition from neurons tuned to adjacent frequencies onto the neurons involved in the tinnitus percept. A reduction of tinnitus loudness and tinnitus-related neural activity was achieved with TMNMT in previous studies. As the effect of lateral inhibition depends on the bandwidth of the notch, in the current study we altered the notch width to find the most effective notch width for TMNMT. We compared 1-octave notch width with ½-octave and ¼-octave. Participants chose their favorite music for the training that included three month of two hours daily listening. The outcome was measured by means of standardized questionnaires and magnetoencephalography. We found a general reduction of tinnitus distress in all administered tinnitus questionnaires after the training. Additionally, tinnitus-related neural activity was reduced after the training. Nevertheless, notch width did not have an influence on the behavioral or neural effects of TMNMT. This could be due to a non-linear resolution of lateral inhibition in high frequencies.

  7. Tailor-made tricalcium phosphate bone implant directly fabricated by a three-dimensional ink-jet printer.

    PubMed

    Igawa, Kazuyo; Mochizuki, Manabu; Sugimori, Osamu; Shimizu, Koutaro; Yamazawa, Kenji; Kawaguchi, Hiroshi; Nakamura, Kozo; Takato, Tsuyoshi; Nishimura, Ryouhei; Suzuki, Shigeki; Anzai, Masahiro; Chung, Ung-il; Sasaki, Nobuo

    2006-01-01

    Rapid prototyping (RP) is a molding technique that builds a three-dimensional (3D) model from computer-aided design (CAD) data. We fabricated new tailor-made bone implants (TIs) from alpha-tricalcium phosphate powder using an RP ink-jet printer based on computed tomography (CT) data, and evaluated their safety and efficacy. CT data of the skulls of seven beagle dogs were obtained and converted to CAD data, and bone defects were virtually made in the skull bilaterally. TIs were designed to fit the defects and were fabricated using the 3D ink-jet printer with six horizontal cylindrical holes running through the implants, designed for possible facilitation of vascular invasion and bone regeneration. As a control, hydroxyapatite implants (HIs) were cut manually from porous hydroxyapatite blocks. Then, craniectomy was performed to create real skull defects, and TIs and HIs were implanted. After implantation, CT was performed regularly, and the animals were euthanized at 24 weeks. No major side effects were observed. CT analysis showed narrowing of the cylindrical holes; bony bridging between the implants and the temporal bone was observed only for TIs. Histological analysis revealed substantial new bone formation inside the cylindrical holes in the TIs, while mainly connective tissues invaded the porous structures in HIs. Bone marrow was observed only in TIs. Osteoclasts were seen to resorb regenerated bone from inside the cylindrical holes and to invade and probably resorb the TIs. These data suggest that TIs are a safe and effective bone substitute, possessing osteoconductivity comparable with that of HIs.

  8. Enhancing inhibition-induced plasticity in tinnitus--spectral energy contrasts in tailor-made notched music matter.

    PubMed

    Stein, Alwina; Engell, Alva; Lau, Pia; Wunderlich, Robert; Junghoefer, Markus; Wollbrink, Andreas; Bruchmann, Maximilian; Rudack, Claudia; Pantev, Christo

    2015-01-01

    Chronic tinnitus seems to be caused by reduced inhibition among frequency selective neurons in the auditory cortex. One possibility to reduce tinnitus perception is to induce inhibition onto over-activated neurons representing the tinnitus frequency via tailor-made notched music (TMNM). Since lateral inhibition is modifiable by spectral energy contrasts, the question arises if the effects of inhibition-induced plasticity can be enhanced by introducing increased spectral energy contrasts (ISEC) in TMNM. Eighteen participants suffering from chronic tonal tinnitus, pseudo randomly assigned to either a classical TMNM or an ISEC-TMNM group, listened to notched music for three hours on three consecutive days. The music was filtered for both groups by introducing a notch filter centered at the individual tinnitus frequency. For the ISEC-TMNM group a frequency bandwidth of 3/8 octaves on each side of the notch was amplified, additionally, by about 20 dB. Before and after each music exposure, participants rated their subjectively perceived tinnitus loudness on a visual analog scale. During the magnetoencephalographic recordings, participants were stimulated with either a reference tone of 500 Hz or a test tone with a carrier frequency representing the individual tinnitus pitch. Perceived tinnitus loudness was significantly reduced after TMNM exposure, though TMNM type did not influence the loudness ratings. Tinnitus related neural activity in the N1m time window and in the so called tinnitus network comprising temporal, parietal and frontal regions was reduced after TMNM exposure. The ISEC-TMNM group revealed even enhanced inhibition-induced plasticity in a temporal and a frontal cortical area. Overall, inhibition of tinnitus related neural activity could be strengthened in people affected with tinnitus by increasing spectral energy contrast in TMNM, confirming the concepts of inhibition-induced plasticity via TMNM and spectral energy contrasts.

  9. Identification of a glycoprotein produced by enterotoxigenic Escherichia coli.

    PubMed

    Lindenthal, C; Elsinghorst, E A

    1999-08-01

    Enterotoxigenic Escherichia coli (ETEC) strain H10407 is capable of invading epithelial cell lines derived from the human ileocecum and colon in vitro. Two separate chromosomally encoded invasion loci (tia and tib) have been cloned from this strain. These loci direct nonadherent and noninvasive laboratory strains of E. coli to adhere to and invade cultured human intestinal epithelial cells. The tib locus directs the synthesis of TibA, a 104-kDa outer membrane protein that is directly correlated with the adherence and invasion phenotypes. TibA is synthesized as a 100-kDa precursor (preTibA) that must be modified for biological activity. Outer membranes of recombinant E. coli expressing TibA or preTibA were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted to nitrocellulose. The presence of glycoproteins was detected by oxidization of carbohydrates with periodate and labeling with hydrazide-conjugated digoxigenin. Only TibA could be detected as a glycoprotein. Complementation experiments with tib deletion mutants of ETEC strain H10407 demonstrate that the TibA glycoprotein is expressed in H10407, that the entire tib locus is required for TibA synthesis, and that TibA is the only glycoprotein produced by H10407. Protease treatment of intact H10407 cells removes the carbohydrates on TibA, suggesting that they are surface exposed. TibA shows homology with AIDA-I from diffuse-adhering E. coli and with pertactin precursor from Bordetella pertussis. Both pertactin and AIDA-I are members of the autotransporter family of outer membrane proteins and are afimbrial adhesins that play an important role in the virulence of these organisms. Analysis of the predicted TibA amino acid sequence indicates that TibA is also an autotransporter. Analysis of the tib locus DNA sequence revealed an open reading frame with similarity to RfaQ, a glycosyltransferase. The product of this tib locus open reading frame is proposed to be responsible for Tib

  10. Identification of a Glycoprotein Produced by Enterotoxigenic Escherichia coli

    PubMed Central

    Lindenthal, Christoph; Elsinghorst, Eric A.

    1999-01-01

    Enterotoxigenic Escherichia coli (ETEC) strain H10407 is capable of invading epithelial cell lines derived from the human ileocecum and colon in vitro. Two separate chromosomally encoded invasion loci (tia and tib) have been cloned from this strain. These loci direct nonadherent and noninvasive laboratory strains of E. coli to adhere to and invade cultured human intestinal epithelial cells. The tib locus directs the synthesis of TibA, a 104-kDa outer membrane protein that is directly correlated with the adherence and invasion phenotypes. TibA is synthesized as a 100-kDa precursor (preTibA) that must be modified for biological activity. Outer membranes of recombinant E. coli expressing TibA or preTibA were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted to nitrocellulose. The presence of glycoproteins was detected by oxidization of carbohydrates with periodate and labeling with hydrazide-conjugated digoxigenin. Only TibA could be detected as a glycoprotein. Complementation experiments with tib deletion mutants of ETEC strain H10407 demonstrate that the TibA glycoprotein is expressed in H10407, that the entire tib locus is required for TibA synthesis, and that TibA is the only glycoprotein produced by H10407. Protease treatment of intact H10407 cells removes the carbohydrates on TibA, suggesting that they are surface exposed. TibA shows homology with AIDA-I from diffuse-adhering E. coli and with pertactin precursor from Bordetella pertussis. Both pertactin and AIDA-I are members of the autotransporter family of outer membrane proteins and are afimbrial adhesins that play an important role in the virulence of these organisms. Analysis of the predicted TibA amino acid sequence indicates that TibA is also an autotransporter. Analysis of the tib locus DNA sequence revealed an open reading frame with similarity to RfaQ, a glycosyltransferase. The product of this tib locus open reading frame is proposed to be responsible for Tib

  11. Virulence Potential of Activatable Shiga Toxin 2d–Producing Escherichia coli Isolates from Fresh Produce

    PubMed Central

    Melton-Celsa, Angela R.; O'Brien, Alison D.; Feng, Peter C. H.

    2016-01-01

    Shiga toxin (Stx)–producing Escherichia coli (STEC) strains are food- and waterborne pathogens that are often transmitted via beef products or fresh produce. STEC strains cause both sporadic infections and outbreaks, which may result in hemorrhagic colitis and hemolytic uremic syndrome. STEC strains may elaborate Stx1, Stx2, and/or subtypes of those toxins. Epidemiological evidence indicates that STEC that produce subtypes Stx2a, Stx2c, and/or Stx2d are more often associated with serious illness. The Stx2d subtype becomes more toxic to Vero cells after incubation with intestinal mucus or elastase, a process named “activation.” Stx2d is not generally found in the E. coli serotypes most commonly connected to STEC outbreaks. However, STEC strains that are stx2d positive can be isolated from foods, an occurrence that gives rise to the question of whether those food isolates are potential human pathogens. In this study, we examined 14 STEC strains from fresh produce that were stx2d positive and found that they all produced the mucus-activatable Stx2d and that a subset of the strains tested were virulent in streptomycin-treated mice. PMID:26555533

  12. Virulence Potential of Activatable Shiga Toxin 2d-Producing Escherichia coli Isolates from Fresh Produce.

    PubMed

    Melton-Celsa, Angela R; O'Brien, Alison D; Feng, Peter C H

    2015-11-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are food- and waterborne pathogens that are often transmitted via beef products or fresh produce. STEC strains cause both sporadic infections and outbreaks, which may result in hemorrhagic colitis and hemolytic uremic syndrome. STEC strains may elaborate Stx1, Stx2, and/or subtypes of those toxins. Epidemiological evidence indicates that STEC that produce subtypes Stx2a, Stx2c, and/or Stx2d are more often associated with serious illness. The Stx2d subtype becomes more toxic to Vero cells after incubation with intestinal mucus or elastase, a process named "activation." Stx2d is not generally found in the E. coli serotypes most commonly connected to STEC outbreaks. However, STEC strains that are stx2d positive can be isolated from foods, an occurrence that gives rise to the question of whether those food isolates are potential human pathogens. In this study, we examined 14 STEC strains from fresh produce that were stx2d positive and found that they all produced the mucus-activatable Stx2d and that a subset of the strains tested were virulent in streptomycin-treated mice.

  13. Resistance of various shiga toxin-producing Escherichia coli to electrolyzed oxidizing water

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The resistance of thirty two strains of Escherichia coli O157:H7 and six major serotypes of non-O157 Shiga toxin- producing E. coli (STEC) plus E. coli O104 was tested against Electrolyzed oxidizing (EO) water using two different methods; modified AOAC 955.16 sequential inoculation method and minim...

  14. Characterization of CTX-M-14-producing Escherichia coli from food-producing animals.

    PubMed

    Liao, Xiao-Ping; Xia, Jing; Yang, Lei; Li, Liang; Sun, Jian; Liu, Ya-Hong; Jiang, Hong-Xia

    2015-01-01

    Bacterial resistance to the third-generation cephalosporin antibiotics has become a major concern for public health. This study was aimed to determine the characteristics and distribution of bla CTX-M-14, which encodes an extended-spectrum β-lactamase, in Escherichia coli isolated from Guangdong Province, China. A total of 979 E. coli isolates isolated from healthy or diseased food-producing animals including swine and avian were examined for bla CTX-M-14 and then the bla CTX-M-14 -positive isolates were detected by other resistance determinants [extended-spectrum β-lactamase genes, plasmid-mediated quinolone resistance, rmtB, and floR] and analyzed by phylogenetic grouping analysis, PCR-based plasmid replicon typing, multilocus sequence typing, and plasmid analysis. The genetic environments of bla CTX-M-14 were also determined by PCR. The results showed that fourteen CTX-M-14-producing E. coli were identified, belonging to groups A (7/14), B1 (4/14), and D (3/14). The most predominant resistance gene was bla TEM (n = 8), followed by floR (n = 7), oqxA (n = 3), aac(6')-1b-cr (n = 2), and rmtB (n = 1). Plasmids carrying bla CTX-M-14 were classified to IncK, IncHI2, IncHI1, IncN, IncFIB, IncF or IncI1, ranged from about 30 to 200 kb, and with insertion sequence of ISEcp1, IS26, or ORF513 located upstream and IS903 downstream of bla CTX-M-14. The result of multilocus sequence typing showed that 14 isolates had 11 STs, and the 11 STs belonged to five groups. Many of the identified sequence types are reported to be common in E. coli isolates associated with extraintestinal infections in humans, suggesting possible transmission of bla CTX-M-14 between animals and humans. The difference in the flanking sequences of bla CTX-M-14 between the 2009 isolates and the early ones suggests that the resistance gene context continues to evolve in E. coli of food producing animals. PMID:26528278

  15. Characterization of CTX-M-14-producing Escherichia coli from food-producing animals

    PubMed Central

    Liao, Xiao-Ping; Xia, Jing; Yang, Lei; Li, Liang; Sun, Jian; Liu, Ya-Hong; Jiang, Hong-Xia

    2015-01-01

    Bacterial resistance to the third-generation cephalosporin antibiotics has become a major concern for public health. This study was aimed to determine the characteristics and distribution of blaCTX-M-14, which encodes an extended-spectrum β-lactamase, in Escherichia coli isolated from Guangdong Province, China. A total of 979 E. coli isolates isolated from healthy or diseased food-producing animals including swine and avian were examined for blaCTX-M-14 and then the blaCTX-M-14 -positive isolates were detected by other resistance determinants [extended-spectrum β-lactamase genes, plasmid-mediated quinolone resistance, rmtB, and floR] and analyzed by phylogenetic grouping analysis, PCR-based plasmid replicon typing, multilocus sequence typing, and plasmid analysis. The genetic environments of blaCTX-M-14 were also determined by PCR. The results showed that fourteen CTX-M-14-producing E. coli were identified, belonging to groups A (7/14), B1 (4/14), and D (3/14). The most predominant resistance gene was blaTEM (n = 8), followed by floR (n = 7), oqxA (n = 3), aac(6′)-1b-cr (n = 2), and rmtB (n = 1). Plasmids carrying blaCTX-M-14 were classified to IncK, IncHI2, IncHI1, IncN, IncFIB, IncF or IncI1, ranged from about 30 to 200 kb, and with insertion sequence of ISEcp1, IS26, or ORF513 located upstream and IS903 downstream of blaCTX-M-14. The result of multilocus sequence typing showed that 14 isolates had 11 STs, and the 11 STs belonged to five groups. Many of the identified sequence types are reported to be common in E. coli isolates associated with extraintestinal infections in humans, suggesting possible transmission of blaCTX-M-14 between animals and humans. The difference in the flanking sequences of blaCTX-M-14 between the 2009 isolates and the early ones suggests that the resistance gene context continues to evolve in E. coli of food producing animals. PMID:26528278

  16. Comparison of non-O157 Shiga toxin-producing E. coli detection systems

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Category: methodology improvements Objective: To identify strengths and weaknesses of commercial Shiga toxin-producing E. coli detection systems and kits in a side by side fashion. Experimental Design: Three commercial Shiga toxin-producing E. coli detection tests (BAX, GDS, and GeneDisc) and two t...

  17. Expansion of Shiga Toxin-Producing Escherichia coli by Use of Bovine Antibiotic Growth Promoters.

    PubMed

    Kim, Jong-Chul; Chui, Linda; Wang, Yang; Shen, Jianzhong; Jeon, Byeonghwa

    2016-05-01

    Antibiotics are routinely used in food-producing animals to promote growth and prevent infectious diseases. We investigated the effects of bovine antibiotic growth promoters (bAGPs) on the propagation and spread of Shiga toxin (Stx)-encoding phages in Escherichia coli. Co-culture of E. coli O157:H7 and other E. coli isolated from cattle in the presence of sublethal concentrations of bAGPs significantly increased the emergence of non-O157, Stx-producing E. coli by triggering the SOS response system in E. coli O157:H7. The most substantial mediation of Stx phage transmission was induced by oxytetracyline and chlortetracycline, which are commonly used in agriculture. bAGPs may therefore contribute to the expansion of pathogenic Stx-producing E. coli.

  18. Expansion of Shiga Toxin–Producing Escherichia coli by Use of Bovine Antibiotic Growth Promoters

    PubMed Central

    Kim, Jong-Chul; Chui, Linda; Wang, Yang; Shen, Jianzhong

    2016-01-01

    Antibiotics are routinely used in food-producing animals to promote growth and prevent infectious diseases. We investigated the effects of bovine antibiotic growth promoters (bAGPs) on the propagation and spread of Shiga toxin (Stx)–encoding phages in Escherichia coli. Co-culture of E. coli O157:H7 and other E. coli isolated from cattle in the presence of sublethal concentrations of bAGPs significantly increased the emergence of non-O157, Stx-producing E. coli by triggering the SOS response system in E. coli O157:H7. The most substantial mediation of Stx phage transmission was induced by oxytetracyline and chlortetracycline, which are commonly used in agriculture. bAGPs may therefore contribute to the expansion of pathogenic Stx-producing E. coli. PMID:27088186

  19. Shiga toxin-producing E. coli and ruminant diets: A match made in heaven?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga-toxin producing E. coli (STEC) are also known as enterohemorrhagic E. coli (EHEC), which are pathogenic bacteria that can cause serious illnesses in humans who consume contaminated foods or water. This pathogen is commonly associated with cattle as it can survive within the intestinal tract a...

  20. Pathogenic Escherichia coli producing Extended-Spectrum β-Lactamases isolated from surface water and wastewater.

    PubMed

    Franz, Eelco; Veenman, Christiaan; van Hoek, Angela H A M; de Roda Husman, Ana; Blaak, Hetty

    2015-01-01

    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 < 0.001) and phylogenetic group B2 (X(2) = 23.5, P < 0.001). Finally, 84% of the pathogenic ESBL-producing E. coli isolates were resistant to three or more different classes of 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.

  1. Pathogenic Escherichia coli producing Extended-Spectrum β-Lactamases isolated from surface water and wastewater

    PubMed Central

    Franz, Eelco; Veenman, Christiaan; van Hoek, Angela H. A. M.; Husman, Ana de Roda; Blaak, Hetty

    2015-01-01

    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 (X2 = 14.7, P < 0.001) and phylogenetic group B2 (X2 = 23.5, P < 0.001). Finally, 84% of the pathogenic ESBL-producing E. coli isolates were resistant to three or more different classes of 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. PMID:26399418

  2. ESBL-producing uropathogenic Escherichia coli isolated from dogs and cats in Switzerland.

    PubMed

    Huber, Helen; Zweifel, Claudio; Wittenbrink, Max M; Stephan, Roger

    2013-03-23

    Extended-spectrum β-lactamase (ESBL)-producing Escherichia (E.) coli have emerged in human and veterinary medicine. The aim of this study was to investigate the presence of ESBL-producers among uropathogenic E. coli isolated from dogs and cats and to characterize detected ESBL-producing isolates by antibiotic susceptibility testing, identification of ESBL genes, multi-locus sequence typing (MLST), detection of putative virulence genes, and analysis of E. coli phylogroups. Among the 107 E. coli isolates derived from urinary samples (59 from dogs, 40 from cats), eight isolates from four different animals (two dogs, two cats) were found to be ESBL-producers. These isolates were of ST533/CTX-M-15/TEM/phylogroup B1 (four strains from one dog), ST410/CTX-M-15/TEM/phylogroup A (three strains, one from a dog and two from a cat), and ST648/CTX-M-15/phylogroup D (one strain from a cat). In terms of putative virulence factors, all isolates harbored lpfA, sat, and tsh, whereas iss was only detected in strains of ST533. Thus, ESBL-producers were detected among uropathogenic E. coli from Swiss companion animals and the eight CTX-M-15-producing isolates belonged to three sequence types (ST410, ST533, ST648) and three E. coli phylogroups (A, B1, D). For the first time, E. coli of ST533 carrying bla(CTX-M-15) were thereby detected in a dog.

  3. High Emergence of ESBL-Producing E. coli Cystitis: Time to Get Smarter in Cyprus

    PubMed Central

    Cantas, Leon; Suer, Kaya; Guler, Emrah; Imir, Turgut

    2016-01-01

    Background: Widespread prevalence of extended-spectrum βeta-lactamase producing Escherichia coli (ESBL-producing E. coli) limits the infection therapeutic options and is a growing global health problem. In this study our aim was to investigate the antimicrobial resistance profile of the E. coli in hospitalized and out-patients in Cyprus. Results: During the period 2010–2014, 389 strains of E. coli were isolated from urine samples of hospitalized and out-patients in Cyprus. ESBL-producing E. coli, was observed in 53% of hospitalized and 44% in out-patients, latest one being in 2014. All ESBL-producing E. coli remained susceptible to amikacin, carbapenems except ertapenem (in-patients = 6%, out-patients = 11%). Conclusion: High emerging ESBL-producing E. coli from urine samples in hospitalized and out-patients is an extremely worrisome sign of development of untreatable infections in the near future on the island. We therefore emphasize the immediate need for establishment of optimal therapy guidelines based on the country specific surveillance programs. The need for new treatment strategies, urgent prescription habit changes and ban of over-the-counter sale of antimicrobials at each segment of healthcare services is also discussed in this research. PMID:26793167

  4. Purely Chemical Approach for Preparation of d-α-Amino Acids via (S)-to-(R)-Interconversion of Unprotected Tailor-Made α-Amino Acids.

    PubMed

    Nian, Yong; Wang, Jiang; Zhou, Shengbin; Dai, Wenhao; Wang, Shuni; Moriwaki, Hiroki; Kawashima, Aki; Soloshonok, Vadim A; Liu, Hong

    2016-05-01

    Unnatural (R)-α-amino acids (α-AAs) are in growing demand in the biomedical research and pharmaceutical industries. In this work, we present development of a purely chemical approach for preparation of (R)-α-AAs via (S)-to-(R)-interconversion of natural and tailor-made (S)-α-AAs. The method can be used on free, unprotected α-AAs and features a remarkable structural generality including substrates bearing tertiary alkyl chains and reactive functional groups. These attractive characteristics, combined with simplicity of reaction conditions and virtually complete stereochemical outcome, constitute a true methodological advance in this area, rivaling previously reported chemical and biocatalytic approaches.

  5. Prevalence and diversity of enterotoxigenic Escherichia coli strains in fresh produce.

    PubMed

    Feng, Peter C H; Reddy, Shanker P

    2014-05-01

    Analysis of fresh produce showed that enterotoxigenic Escherichia coli (ETEC) strains are most often found in cilantro and parsley, with prevalence rates of approximately 0.3%. Some ETEC strains also carried Shiga toxigenic E. coli (STEC) genes but had no STEC adherence factors, which are essential to cause severe human illness. Most ETEC strains in produce carried stable toxin and/or labile toxin genes but belonged to unremarkable serotypes that have not been reported to have caused human illnesses.

  6. Gasifier feed: Tailor-made from Illinois coals. Interim final technical report, September 1, 1991--August 31, 1992

    SciTech Connect

    Ehrlinger, H.P. III; Lytle, J.; Frost, R.R.; Lizzio, A.; Kohlenberger, L.; Brewer, K.

    1992-12-31

    The main purpose of this project is to produce a feedstock from preparation plant fines from an Illinois coal that is ideal for a slurry fed, slagging, entrained-flow coal gasifier. The high sulfur content and high Btu value of Illinois coals are particularly advantageous in such a gasifier; preliminary calculations indicate that the increased cost of removing sulfur from the gas from a high sulfur coal is more than offset by the increased revenue from the sale of the elemental sulfur; additionally the high Btu Illinois coal concentrates more energy into the slurry of a given coal to water ratio. The Btu is higher not only because of the higher Btu value of the coal but also because Illinois coal requires less water to produce a pumpable slurry than western coal, i.e., as little as 30--35% water may be used for Illinois coal as compared to approximately 45% for most western coals. Destec Energy, a wholly-owned subsidiary of Dow Chemical Company, will provide guidelines and test compatibility of the slurries developed for gasification feedstock. Williams Technologies, Inc., will provide their expertise in long distance slurry pumping, and test selected products for viscosity, pumpability, and handleability. The Illinois State Geological Survey will study methods for producing clean coal/water slurries from preparation plant wastes including the concentration of pyritic sulfur into the coal slurry to increase the revenue from elemental sulfur produced during gasification operations, and decrease the pyritic sulfur content of the waste streams. ISGS will also test the gasification reactivity of the coals.

  7. Assessment of tailor-made prevention of atherosclerosis with folic acid supplementation: randomized, double-blind, placebo-controlled trials in each MTHFR C677T genotype.

    PubMed

    Miyaki, Koichi; Murata, Mitsuru; Kikuchi, Haruhito; Takei, Izumi; Nakayama, Takeo; Watanabe, Kiyoaki; Omae, Kazuyuki

    2005-01-01

    This study aimed at assessing the effect of folic acid supplementation quantitatively in each MTHFR C677T genotype and considered the efficiency of tailor-made prevention of atherosclerosis. Study design was genotype-stratified, randomized, double-blind, placebo-controlled trials. The setting was a Japanese company in the chemical industry. Subjects were 203 healthy men after exclusion of those who took folic acid or drugs known to effect folic acid metabolism. Intervention was folic acid 1 mg/day p.o. for 3 months. The primary endpoint was plasma total homocysteine level (tHcy). In all three genotypes, there were significant tHcy decreases. The greatest decrease was in the TT homozygote [6.61 (3.47-9.76) micromol/l] compared with other genotypes [CC: 2.59 (1.81-3.36), CT: 2.64 (2.16-3.13)], and there was a significant trend between the mutated allele number and the decrease. The tHcy were significantly lowered in all the genotypes, but the amount of the decrease differed significantly in each genotype, which was observed at both 1 and 3 months. Using these time-series data, the largest benefit obtained by the TT homozygote was appraised as 2.4 times compared with the CC homozygote. Taking into account the high allele frequency of this SNP, this quantitative assessment should be useful when considering tailor-made prevention of atherosclerosis with folic acid. PMID:15895286

  8. Prevalence of extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella pneumoniae in food-producing animals.

    PubMed

    Hiroi, Midori; Yamazaki, Fumie; Harada, Tetsuya; Takahashi, Naomi; Iida, Natsuko; Noda, Yoshihiro; Yagi, Miya; Nishio, Tomohiro; Kanda, Takashi; Kawamori, Fumihiko; Sugiyama, Kanji; Masuda, Takashi; Hara-Kudo, Yukiko; Ohashi, Norio

    2012-02-01

    To evaluate the diversity of extended-spectrum β-lactamases (ESBL) genes among food-producing animals, 48 isolates of ESBL-producing Escherichia coli isolates were obtained from rectal samples of broilers, layers, beef cattle and pigs, at the slaughterhouse level. ESBL-carrying E. coli were isolated from 60.0% of individual broiler rectal samples, 5.9% of layers, 12.5% of beef cattle and 3% of pigs. One ESBL-producing Klebsiella pneumoniae was isolated from a broiler. The ESBL-positive E. coli isolates from broilers harbored various ESBL genes: bla (SHV-12), bla(CTX-M-2), bla(CTX-M-14), bla(CTX-M-15) and bla(CTX-M-44). The plasmid DNAs were analyzed by restriction patterns. Homogeneous band patterns were yielded in those of K. pneumoniae and E. coli isolates harboring the bla(CTX-M-2) gene from different farms. No genetic relation between the 2 CTX-M-14 ESBL-producing strains was found by pulsed-field gel electrophoresis, although 2 plasmids in these strains, obtained from different broiler farms, were similar to each other. This study provides evidence that the proliferation of CTX-M-producing E. coli is due to the growth of indigenous CTX-M-producing strains and the possible emergence of strains that acquired CTX-M genes by horizontal transfer in different broiler farms. CTX-M-producing coliforms in broilers should be controlled due to the critical importance of cephalosporins and the zoonotic potential of ESBL-producing bacteria.

  9. Emergence of Carbapenemase-Producing Escherichia coli Isolated from Companion Animals in Algeria.

    PubMed

    Yousfi, Massilia; Touati, Abdelaziz; Mairi, Assia; Brasme, Lucien; Gharout-Sait, Alima; Guillard, Thomas; De Champs, Christophe

    2016-06-01

    The emergence and worldwide spread of carbapenemase-producing Enterobacteriaceae is of great concern to public health. The aim of this study was to investigate the occurrence of carbapenemase-producing Escherichia coli in companion animals in Algeria. Two hundred fecal samples were obtained from healthy and diseased dogs and cats in one veterinary office and private owners in Bejaia city, Algeria, during November 2014 to March 2015. Isolates were screened by polymerase chain reaction for the presence of carbapenemase, acquired plasmidic AmpC (pAmpC) and extended-spectrum beta-lactamase genes. Five carbapenemase-producing E. coli isolates were detected including four OXA-48-producing isolates and one isolate producing NDM-5. Coexpression of ESBL and pAmpC genes was observed in these isolates. Phylogenetic grouping revealed that these isolates belonged to A and D phylogroups. The results of this study show that carbapenemase-producing E. coli spread to the companion animals in Algeria.

  10. Gasifier feed: Tailor-made from Illinois coals. Final technical report, September 1, 1991--December 31, 1992

    SciTech Connect

    Ehrlinger, H.P. III; Lytle, J.M.; Frost, R.R.; Lizzio, A.A.; Kohlenberger, L.B.; Brewer, K.K. |||

    1992-12-31

    The main purpose of this project was to produce a feedstock from preparation plant fines from an Illinois (IL) coal that is ideal for a slurry fed, slagging, entrained-flow coal gasifier. The high-sulfur content and high-Btu value of IL coals are Particularly advantageous in such a gasifier; preliminary-calculations indicate that the increased cost of removing sulfur from the gas from a high-sulfur coal is more than offset b the increased revenue from the sale of the elemental sulfur; additionally the high-Btu IL coal concentrates more energy into the slurry of a given coal to water ratio. The Btu is--higher not only because of the hither Btu value of the coal but also because IL coal requires less water to produce a pumpable slurry than western coal, i.e., as little as 30--35% water may be used for IL coal as compared to approximately 45% for most western coals. During the contract extension, additional coal testing was completed confirming the fact that coal concentrates can be made from plant waste under a variety of flotation conditions 33 tests were conducted, yielding an average of 13326 Btu with 9.6% ash while recovering 86.0%-Of the energy value.

  11. Antimicrobial-resistant and ESBL-producing Escherichia coli in different ecological niches in Bangladesh

    PubMed Central

    Rashid, Mahmudur; Rakib, Mufti Mahmud; Hasan, Badrul

    2015-01-01

    Introduction The rapid and wide-scale environmental spread of multidrug-resistant bacteria in different ecosystems has become a serious issue in recent years. Objectives To investigate the epidemiology of antimicrobial resistance and extended spectrum beta-lactamase (ESBL) in Bangladeshi wild birds and aquatic environments, samples were taken from Open Bill Stork (Anastomus oscitans) (OBS) and the nearby water sources. Methods Water and fresh fecal samples were collected from several locations. All samples were processed and cultured for Escherichia coli and tested for antibiotic susceptibility against commonly used antibiotics. ESBL producers were characterized at genotypic level using polymerase chain reaction (PCR), sequencing, multilocus sequence typing, and rep-PCR. Results and discussion A total of 76 E. coli isolates from the 170 OBS and 8 E. coli isolates from three river sources were isolated. In total, 29% of E. coli isolated from OBS and all of the E. coli isolated from water sources were resistant to at least one of the tested antimicrobials. Resistant phenotypes were observed with all antimicrobials except tigecycline, gentamicin, imipenem, and chloramphenicol. Multidrug resistance was observed in 2.6% of OBS and 37.5% of the water isolates. Also, 1.2% of the ESBL-producing E. coli were isolated from OBS, whereas 50% of the E. coli isolated from water sources were ESBL producers possessing the CTX-M-15 gene. The most concerning aspect of our findings was the presence of human-associated E. coli sequence types in the water samples, for example, ST156-complex156, ST10-complex10 and ST46. Conclusion This study reports the presence of multidrug-resistant ESBL-producing E. coli in OBSs and nearby aquatic sources in Bangladesh. PMID:26193990

  12. High Prevalence of Mucosa-Associated E. coli Producing Cyclomodulin and Genotoxin in Colon Cancer

    PubMed Central

    Sauvanet, Pierre; Raisch, Jennifer; Delmas, Julien

    2013-01-01

    Some Escherichia coli strains produce toxins designated cyclomodulins (CMs) which interfere with the eukaryotic cell cycle of host cells, suggesting a possible link between these bacteria and cancers. There are relatively few data available concerning the colonization of colon tumors by cyclomodulin- and genotoxic-producing E. coli. We did a qualitative and phylogenetic analysis of mucosa-associated E. coli harboring cyclomodulin-encoding genes from 38 patients with colorectal cancer (CRC) and 31 with diverticulosis. The functionality of these genes was investigated on cell cultures and the genotoxic activity of strains devoid of known CM-encoding gene was investigated. Results showed a higher prevalence of B2 phylogroup E. coli harboring the colibatin-producing genes in biopsies of patients with CRC (55.3%) than in those of patients with diverticulosis (19.3%), (p<0.01). Likewise, a higher prevalence of B2 E. coli harboring the CNF1-encoding genes in biopsies of patients with CRC (39.5%) than in those of patients with diverticulosis (12.9%), (p = 0.01). Functional analysis revealed that the majority of these genes were functional. Analysis of the ability of E. coli to adhere to intestinal epithelial cells Int-407 indicated that highly adherent E. coli strains mostly belonged to A and D phylogroups, whatever the origin of the strains (CRC or diverticulosis), and that most E. coli strains belonging to B2 phylogroup displayed very low levels of adhesion. In addition, 27.6% (n = 21/76) E. coli strains devoid of known cyclomodulin-encoding genes induced DNA damage in vitro, as assessed by the comet assay. In contrast to cyclomodulin-producing E. coli, these strains mainly belonged to A or D E. coli phylogroups, and exhibited a non significant difference in the distribution of CRC and diverticulosis specimens (22% versus 32.5%, p = 0.91). In conclusion, cyclomodulin-producing E. coli belonging mostly to B2 phylogroup colonize the colonic mucosa of patients

  13. Antibacterial Activity of Some Plant Extracts Against Extended- Spectrum Beta-Lactamase Producing Escherichia coli Isolates

    PubMed Central

    Saeidi, Saeide; Amini Boroujeni, Negar; Ahmadi, Hassan; Hassanshahian, Mehdi

    2015-01-01

    Background: The extended-spectrum beta-lactamase (ESBL) -producing Escherichia coli isolates make many serious infections, especially urinary tract infections. Objectives: The purpose of this study was to determine the antibacterial activities of some natural plant extracts against ESBL-producing E. coli isolates, which harbor the TEM gene in urine samples of the patients who have urinary tract infections. Materials and Methods: Evaluation has to be exactly determined for both methods of disk diffusion test and polymerase chain reaction (PCR), separately. We evaluated 120 strains of E. coli isolates from the urine culture of the patients in Boo-Ali Hospital (Zahedan, south-eastern Iran) who were suffering from urinary tract infections. The ESBL-producing E. coli isolates were evaluated by disk diffusion test and PCR through TEM gene detection. The minimal inhibitory concentration (MIC) of commonly used antibiotics including ceftazidime, ceftriaxon, amikacin, gentamicin and ciprofloxacin along with the MIC of the alcoholic extract of different natural plants including Myrtus communis L (Myrtaceae), Amaranthus retraflexus (Amaranthaceae), Cyminum cuminum L (Apiaceae), Marrubium vulgare (Laminaceae) and Peganum. harmala (Zygrophyllaceae) against the ESBL-producing E. coli isolates, which harbor the TEM genes, were determined using the microdulition method. Results: Results of this study showed that in disk diffusion method, 80 samples of E. coli produced ESBLs. In PCR method, the TEM gene distribution in the isolated ESBL-producing organisms was 50 (41.6%). Amikacin was the most effective anti-bacterial agent and ciprofloxacin was the least effective against E. coli isolates. All the natural plant extracts mentioned above, especially P. harmala, were effective against the selected isolates of ESBL-producing E. coli. The most frequent ESBL rate producing E. coli isolates (32 out of 50) had MIC of 2.5 mg/mL in ethanol extract of P. harmala. Conclusions: The alcoholic

  14. Molecular epidemiology of Escherichia coli producing extended-spectrum beta-lactamases isolated in Rome, Italy.

    PubMed

    Carattoli, Alessandra; García-Fernández, Aurora; Varesi, Paola; Fortini, Daniela; Gerardi, Serena; Penni, Adriano; Mancini, Carlo; Giordano, Alessandra

    2008-01-01

    Escherichia coli strains producing extended-spectrum beta-lactamases (ESBLs) are a major problem in many different hospitals worldwide, causing outbreaks as well as sporadic infections. The prevalence of Escherichia coli ESBL producers was analyzed in a surveillance study performed on the population attending the Policlinico Umberto I, the largest university hospital in Rome, Italy. We also investigated genotypes, pathogenicity islands, and plasmids in the ESBL-positive E. coli isolates as further markers that are useful in describing the epidemiology of the infections. In this survey, 163 nonreplicate isolates of Escherichia coli were isolated from patients from 86 different wards, and 28 were confirmed as ESBL producers. A high prevalence (26/28) of CTX-M-15 producers was observed within the bacterial population circulating in this hospital, and the dissemination of this genetic trait was associated with the spread of related strains; however, these do not have the characteristics of a single epidemic clone spreading. The dissemination was also linked to horizontal transfer among the prevalent E. coli genotypes of multireplicon plasmids showing FIA, FIB, and FII replicons in various combinations, which are well adapted to the E. coli species. The analysis of related bacteria suggests a probable interpatient transmission occurring in several wards, causing small outbreaks. PMID:17959756

  15. Coenzyme B12 can be produced by engineered Escherichia coli under both anaerobic and aerobic conditions.

    PubMed

    Ko, Yeounjoo; Ashok, Somasundar; Ainala, Satish Kumar; Sankaranarayanan, Mugesh; Chun, Ah Yeong; Jung, Gyoo Yeol; Park, Sunghoon

    2014-12-01

    Coenzyme B12 (Vitamin B12 ) is one of the most complex biomolecules and an essential cofactor required for the catalytic activity of many enzymes. Pseudomonas denitrificans synthesizes coenzyme B12 in an oxygen-dependent manner using a pathway encoded by more than 25 genes that are located in six different operons. Escherichia coli, a robust and suitable host for metabolic engineering was used to produce coenzyme B12 . These genes were cloned into three compatible plasmids and expressed heterologously in E. coli BL21 (DE3). Real-time PCR, SDS-PAGE analysis and bioassay showed that the recombinant E. coli expressed the coenzyme B12 synthetic genes and successfully produced coenzyme B12 . However, according to the quantitative determination by inductively coupled plasma-mass spectrometry, the amount of coenzyme B12 produced by the recombinant E. coli (0.21 ± 0.02 μg/g cdw) was approximately 13-fold lower than that by P. denitrificans (2.75 ± 0.22 μg/g cdw). Optimization of the culture conditions to improve the production of coenzyme B12 by the recombinant E. coli was successful, and the highest titer (0.65 ± 0.03 μg/g cdw) of coenzyme B12 was obtained. Interestingly, although the synthesis of coenzyme B12 in P. denitrificans is strictly oxygen-dependent, the recombinant E. coli could produce coenzyme B12 under anaerobic conditions.

  16. Characterization of non-Shiga-toxin-producing Escherichia coli O157 strains isolated from dogs.

    PubMed

    Bentancor, A; Vilte, D A; Rumi, M V; Carbonari, C C; Chinen, I; Larzábal, M; Cataldi, A; Mercado, E C

    2010-01-01

    Shiga toxin-negative Escherichia coli O157 strains of various H types have been associated with diarrhea in children and are considered potentially pathogenic for humans. In this study, we describe non-Shiga toxin-producing E. coli O157 E. coli strains previously obtained from dogs in Argentina. Different E. coli phylogenetic lineages corresponding to flagellar types H16, H29 and H45 were identified. E. coli serotypes O157:H16 and O157:H45 contained intimin subtypes epsilon and alpha 1, respectively. Serotype O157:H45 carried the bfp gene encoding the bundle-forming pilus. Localized adherence-like patterns to HEp-2 cells were observed in O157:H16 strains, while O157:H45 adhered in a typical localized pattern. A total of eight different XbaI-pulse field electrophoresis patterns with more than 74 % similarity were identified among the nine E. coli O157:H16 strains. Our data emphasized the fact that dogs may harbor human pathogenic E. coli O157 which do not correspond to Shiga toxin-producing strains and whose potential human health hazard should not be underestimated. PMID:20461294

  17. The prevalence of verocytotoxin-producing Escherichia coli and antimicrobial resistance patterns of nonverocytotoxin-producing Escherichia coli and Salmonella in Ontario broiler chickens.

    PubMed Central

    Irwin, R J; McEwen, S A; Clarke, R C; Meek, A H

    1989-01-01

    The prevalence of verocytotoxin-producing Escherichia coli and Salmonella in Ontario broiler chickens was determined by culturing cloacal samples from 500 individual birds selected from 50 poultry farms. Resistance to antimicrobials was determined for each of the isolates. In addition, abattoir and farm-level management data were obtained to evaluate variables that may be considered risk factors for infection. The variables selected included: Percentage of birds condemned at slaughter, percentage of birds dead-on-arrival, bird weight, truck number, farm size, hatchery source, litter source and type, feed source, mortality levels, type of water drinker, water sanitization, down time, barn clean out and history of antibiotic treatment. None of the cloacal samples revealed the presence of verocytotoxin-producing E. coli, though 19/500 (3.8%) contained Salmonella organisms. Nine different Salmonella serotypes were isolated; the most common being S. hadar, S. heidelberg and S. mbandaka. Resistance to tetracycline and streptomycin was common among Salmonella (63%) and E. coli (25.2%) isolates. Resistance to two or more antimicrobials occurred in 420/500 (84%) of the E. coli isolates. No statistically significant associations between abattoir or farm-level management variables and the Salmonella-status of farms were demonstrated. PMID:2686829

  18. Effects of a tailor-made exercise program on exercise adherence and health outcomes in patients with knee osteoarthritis: a mixed-methods pilot study

    PubMed Central

    Lee, Fung-Kam Iris; Lee, Tze-Fan Diana; So, Winnie Kwok-Wei

    2016-01-01

    Introduction Previous studies showed that exercise intervention was effective in symptoms control of knee osteoarthritis (OA) but poor intervention adherence reduced the exercise effect. It has been suspected that the design of exercise intervention mainly from the health care professionals’ perspective could not address the patients’ barriers to exercise. Therefore, a tailor-made exercise program which incorporated the patient’s perspective in the design was developed and ready for evaluation. Objectives This pilot study estimated the effects of a tailor-made exercise program on exercise adherence and health outcomes, and explored the participants’ perception and experience of the program. Methods The intervention of this study was a 4-week community-based group exercise program, which required the participants to attend a 1-hour session each week. Thirty-four older people with knee OA were recruited to the program. Mixed-methods study design was used to estimate the effects of this program and explore the participants’ perception and experience of the program. Exercise adherence and performance in return-demonstration of the exercise were assessed at 12 weeks after the program. Disease-specific health status (Western Ontario and McMaster Universities Osteoarthritis Index), general health status (12-item Short Form of the Medical Outcome Study Questionnaire), knee range of motion, muscle strength, and endurance of the lower extremities (Timed-Stands Test) were measured at the beginning of the program and 12 weeks after. Six participants were interviewed individually on the 12th week. Results Thirty-three participants (75.0±7.3 years) completed the one-group pretest and post-test study. The participants’ exercise adherence was 91.4%±14.54%, and their correct performance in return-demonstration was 76.7%±21.75%. Most of the participants’ health outcomes significantly improved at posttests except the 12-item Short Form of the Medical Outcome Study

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

    PubMed

    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.

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

    PubMed

    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. PMID:27375249

  1. Isolation and characterization of verocytotoxin-producing Escherichia coli O157 from slaughter pigs and poultry.

    PubMed

    Heuvelink, A E; Zwartkruis-Nahuis, J T; van den Biggelaar, F L; van Leeuwen, W J; de Boer, E

    1999-11-01

    Rectal contents and tonsils from Dutch slaughter pigs collected immediately after slaughter were examined for the presence of verocytotoxin (VT)-producing Escherichia coli (VTEC) of serogroup O157 (O157 VTEC). In addition, fresh fecal material from poultry layer flocks and turkey flocks collected on poultry farms was examined for the presence of O157 VTEC. E. coli O157 strains were isolated from two (1.4%) of 145 pigs. The strains were isolated from samples of rectal contents, all samples of tonsils being negative. While all 501 fecal samples from chicken flocks were found negative, E. coli O157 strains were isolated from six (1.3%) of 459 pooled fecal samples from turkey flocks. One of the porcine isolates and one of the turkey isolates contained the VT2 gene, the E. coli attaching-and-effacing gene, as well as the enterohemorrhagic E. coli hemolysin gene. Production of VT was confirmed by cytotoxicity tests on Vero cells. Based on these characteristics, the two stains were regarded as potentially pathogenic for humans. The porcine and the turkey isolate were further characterized as being of phage types 4 and 14, respectively. While biochemically typical of E. coli O157, the remaining six isolates were nonverocytotoxigenic and negative for both the E. coli attaching-and-effacing gene and the enterohemorrhagic E. coli hemolysin gene. All eight E. coli O157 isolates did not carry genes that encode E. coli heat-labile and heat-stable enterotoxins. It was concluded that pigs and poultry can be a source of O157 VTEC strains characteristic of those causing illness in man. The extent to which pigs and poultry play a role in the epidemiology of human O157 VTEC infection needs further research. PMID:10573393

  2. Study protocol: münster tinnitus randomized controlled clinical trial-2013 based on tailor-made notched music training (TMNMT)

    PubMed Central

    2014-01-01

    Background Tinnitus is a result of hyper-activity/hyper-synchrony of auditory neurons coding the tinnitus frequency, which has developed to synchronous mass activity owing the lack of inhibition. We assume that removal of exactly these frequency components from an auditory stimulus will cause the brain to reorganize around tonotopic regions coding the tinnitus frequency. Based on this assumption a novel treatment for tonal tinnitus - tailor-made notched music training (TMNMT) (Proc Natl Acad Sci USA 107:1207–1210, 2010; Ann N Y Acad Sci 1252:253–258, 2012; Frontiers Syst Neurosci 6:50, 2012) has been introduced and will be tested in this clinical trial on a large number of tinnitus patients. Methods and design A randomized controlled trial (RCT) in parallel group design will be performed in a double-blinded manner. The choice of the intervention we are going to apply is based on two “proof of concept” studies in humans (Proc Natl Acad Sci USA 107:1207–1210, 2010; Ann N Y Acad Sci 1252:253–258, 2012; Frontiers Syst Neurosci 6:50, 2012; PloS One 6(9):e24685, 2011) and on a recent animal study (Front Syst Neurosci 7:21, 2013). The RCT includes 100 participants with chronic, tonal tinnitus who listened to tailor-made notched music (TMNM) for two hours a day for three months. The effect of TMNMT is assessed by the tinnitus handicap questionnaire and visual analogue scales (VAS) measuring perceived tinnitus loudness, distress and handicap. Discussion This is the first randomized controlled trial applying TMNMT on a larger number of patients with tonal tinnitus. Our data will verify more securely and reliably the effectiveness of this kind of completely non-invasive and low-cost treatment approach on tonal tinnitus. Trial registration Current Controlled Trials ISRCTN04840953 PMID:24581050

  3. A Tailor-Made City

    ERIC Educational Resources Information Center

    Kahama, Clement George

    1975-01-01

    Dodoma, future capital of Tanzania, is one of the first planned attempts at integrating man in his environment. The four principle elements of the Master Plan include: the residential communities, the national capital central spine, the system of open spaces and the transportation network. (BT)

  4. Teacher Training, Tailor-Made

    ERIC Educational Resources Information Center

    Newman, Katherine

    2009-01-01

    Family Partnerships for Achievement is not a course typical of most master's programs in education. The course was designed with one overriding goal: to prepare teachers to be effective in the Boston Public Schools (BPS). This goal drives every aspect of the Boston Teacher Residency (BTR), a district-based program for teacher training and…

  5. Oral tolerance failure upon neonatal gut colonization with Escherichia coli producing the genotoxin colibactin.

    PubMed

    Secher, Thomas; Payros, Delphine; Brehin, Camille; Boury, Michele; Watrin, Claude; Gillet, Marion; Bernard-Cadenat, Isabelle; Menard, Sandrine; Theodorou, Vassilia; Saoudi, Abdelhadi; Olier, Maiwenn; Oswald, Eric

    2015-06-01

    The intestinal barrier controls the balance between tolerance and immunity to luminal antigens. When this finely tuned equilibrium is deregulated, inflammatory disorders can occur. There is a concomitant increase, in urban populations of developed countries, of immune-mediated diseases along with a shift in Escherichia coli population from the declining phylogenetic group A to the newly dominant group B2, including commensal strains producing a genotoxin called colibactin that massively colonized the gut of neonates. Here, we showed that mother-to-offspring early gut colonization by colibactin-producing E. coli impairs intestinal permeability and enhances the transepithelial passage of luminal antigen, leading to an increased immune activation. Functionally, this was accompanied by a dramatic increase in local and systemic immune responses against a fed antigen, decreased regulatory T cell population, tolerogenic dendritic cells, and enhanced mucosal delayed-type hypersensitivity response. Conversely, the abolition of colibactin expression by mutagenesis abrogates the alteration of oral tolerance induced by neonatal colonization by E. coli. In conclusion, the vertical colonization by E. coli producing the genotoxin colibactin enhances intestinal translocation and subsequently alters oral tolerance. Thus, early colonization by E. coli from the newly dominant phylogenetic group B2, which produces colibactin, may represent a risk factor for the development of immune-mediated diseases. PMID:25824839

  6. Rapid, Multiplexed Characterization of Shiga Toxin-Producing Escherichia coli (STEC) Isolates Using Suspension Array Technology

    PubMed Central

    Carter, John M.; Lin, Andrew; Clotilde, Laurie; Lesho, Matthew

    2016-01-01

    Molecular methods have emerged as the most reliable techniques to detect and characterize pathogenic Escherichia coli. These molecular techniques include conventional single analyte and multiplex PCR, PCR followed by microarray detection, pulsed-field gel electrophoresis (PFGE), and whole genome sequencing. The choice of methods used depends upon the specific needs of the particular study. One versatile method involves detecting serogroup-specific markers by hybridization or binding to encoded microbeads in a suspension array. This molecular serotyping method has been developed and adopted for investigating E. coli outbreaks. The major advantages of this technique are the ability to simultaneously serotype E. coli and detect the presence of virulence and pathogenicity markers. Here, we describe the development of a family of multiplex molecular serotyping methods for Shiga toxin-producing E. coli, compare their performance to traditional serotyping methods, and discuss the cost-benefit balance of these methods in the context of various food safety objectives. PMID:27242670

  7. Rapid, Multiplexed Characterization of Shiga Toxin-Producing Escherichia coli (STEC) Isolates Using Suspension Array Technology.

    PubMed

    Carter, John M; Lin, Andrew; Clotilde, Laurie; Lesho, Matthew

    2016-01-01

    Molecular methods have emerged as the most reliable techniques to detect and characterize pathogenic Escherichia coli. These molecular techniques include conventional single analyte and multiplex PCR, PCR followed by microarray detection, pulsed-field gel electrophoresis (PFGE), and whole genome sequencing. The choice of methods used depends upon the specific needs of the particular study. One versatile method involves detecting serogroup-specific markers by hybridization or binding to encoded microbeads in a suspension array. This molecular serotyping method has been developed and adopted for investigating E. coli outbreaks. The major advantages of this technique are the ability to simultaneously serotype E. coli and detect the presence of virulence and pathogenicity markers. Here, we describe the development of a family of multiplex molecular serotyping methods for Shiga toxin-producing E. coli, compare their performance to traditional serotyping methods, and discuss the cost-benefit balance of these methods in the context of various food safety objectives. PMID:27242670

  8. Molecular epidemiology of VIM-1 producing Escherichia coli from Germany referred to the National Reference Laboratory.

    PubMed

    Kaase, Martin; Pfennigwerth, Niels; Lange, Felix; Anders, Agnes; Gatermann, Sören G

    2015-10-01

    The distribution of carbapenemase genes in Escherichia coli strains isolated between September 2009 and May 2013 in Germany was investigated. Out of 192 isolates with carbapenemase production OXA-48 was found in 44.8%, VIM-1 in 18.8%, NDM-1 in 11.5% and KPC-2 in 6.8%. Patients with VIM-1 producing E. coli (n=36) differed from patients with OXA-48 by an older age, less frequent mention of travel history and an increased proportion of clinical over screening specimens. These data might indicate that introduction from abroad is of minor importance for VIM-1 producing E. coli compared to other carbapenemases. Multilocus sequence typing revealed that E. coli with VIM-1 were mostly multiclonal, emphasizing the role of horizontal gene transfer in its spread. Susceptibility testing of VIM-1 producing E. coli demonstrated aztreonam susceptibility in 55.6%. Among non-β-lactams susceptibility rates of >90% were observed for amikacin, tigecycline, colistin, fosfomycin and nitrofurantoin.

  9. Fecal Colonization with Extended-Spectrum Beta-Lactamase and AmpC-Producing Escherichia coli

    PubMed Central

    El Mahdy, Taghrid S.; Shibl, Atef M.

    2016-01-01

    Background. Extended-spectrum β-lactamases (ESβLs) and AmpC β-lactamases cause β-lactam resistance in Escherichia coli. Fecal colonization by ESβL- and/or AmpC-positive E. coli is a source of nosocomial infections. Methods. In order to investigate inpatient fecal colonization by ESβLs and AmpC, antibiotic sensitivity tests were conducted and minimum inhibitory concentrations (MICs) were determined using the disk diffusion method and E-test, respectively. Characterization of ESβL and AmpC was performed using E-test strips, and a set of PCRs and DNA sequence analyses were used to characterize the ESβL and AmpC genes. Results. The whole collection of E. coli isolates (n = 50) was sensitive to imipenem, tigecycline, colistin, and fosfomycin, while 26% of the isolates showed reduced susceptibility to ceftazidime (MIC ≥ 4 μg/mL). ESβL was phenotypically identified in 26% (13/50) of cases, while AmpC activity was detected in two ESβL-producing E. coli isolates. All ESβL-producing E. coli were positive for the CTX-M gene, eleven isolates carried blaCTX-M-15, and two isolates carried blaCTX-M-14 gene. Two CTX-M-positive E. coli isolates carried blaCMY-2. Conclusions. The alimentary tract is a significant reservoir for ESβL- and/or AmpC-producing E. coli, which may lead to nosocomial infection. PMID:27340657

  10. Metabolic evolution of Escherichia coli strains that produce organic acids

    SciTech Connect

    Grabar, Tammy; Gong, Wei; Yocum, R Rogers

    2014-10-28

    This invention relates to the metabolic evolution of a microbial organism previously optimized for producing an organic acid in commercially significant quantities under fermentative conditions using a hexose sugar as sole source of carbon in a minimal mineral medium. As a result of this metabolic evolution, the microbial organism acquires the ability to use pentose sugars derived from cellulosic materials for its growth while retaining the original growth kinetics, the rate of organic acid production and the ability to use hexose sugars as a source of carbon. This invention also discloses the genetic change in the microorganism that confers the ability to use both the hexose and pentose sugars simultaneously in the production of commercially significant quantities of organic acids.

  11. Quantitative PCR measurements of Escherichia coli including shiga toxin-producing E. coli (STEC) in animal feces and environmental waters.

    PubMed

    Ahmed, W; Gyawali, P; Toze, S

    2015-03-01

    Quantitative PCR (qPCR) assays were used to determine the concentrations of E. coli including shiga toxin-producing E. coli (STEC) associated virulence genes (eaeA, stx1, stx2, and hlyA) in ten animal species (fecal sources) and environmental water samples in Southeast Queensland, Australia. The mean Log10 concentrations and standard deviations of E. coli 23S rRNA across fecal sources ranged from 1.3 ± 0.1 (horse) to 6.3 ± 0.4 (cattle wastewater) gene copies at a test concentration of 10 ng of DNA. The differences in mean concentrations of E. coli 23S rRNA gene copies among fecal source samples were significantly different from each other (P < 0.0001). Among the virulence genes, stx2 (25%, 95% CI, 17-33%) was most prevalent among fecal sources, followed by eaeA (19%, 95% CI, 12-27%), stx1 (11%, 95% CI, 5%-17%) and hlyA (8%, 95% CI, 3-13%). The Log10 concentrations of STEC virulence genes in cattle wastewater samples ranged from 3.8 to 5.0 gene copies at a test concentration of 10 ng of DNA. Of the 18 environmental water samples tested, three (17%) were positive for eaeA and two (11%) samples were also positive for the stx2 virulence genes. The data presented in this study will aid in the estimation of quantitative microbial risk assessment (QMRA) from fecal pollution of domestic and wild animals in drinking/recreational water catchments.

  12. Effect of the food matrix on pressure resistance of Shiga-toxin producing Escherichia coli.

    PubMed

    Li, Hui; Garcia-Hernandez, Rigoberto; Driedger, Darcy; McMullen, Lynn M; Gänzle, Michael

    2016-08-01

    The pressure resistance of Shiga-toxin producing Escherichia coli (STEC) depends on food matrix. This study compared the resistance of two five-strain E. coli cocktails, as well as the pressure resistant strain E. coli AW1.7, to hydrostatic pressure application in bruschetta, tzatziki, yoghurt and ground beef at 600 MPa, 20 °C for 3 min and during post-pressure survival at 4 °C. Pressure reduced STEC in plant and dairy products by more than 5 logs (cfu/ml) but not in ground beef. The pH affected the resistance of STEC to pressure as well as the post-pressure survival. E. coli with food constituents including calcium, magnesium, glutamate, caffeic acid and acetic acid were treated at 600 MPa, 20 °C. All compounds exhibited a protective effect on E. coli. The antimicrobial compounds ethanol and phenylethanol enhanced the inactivation by pressure. Calcium and magnesium also performed protective effects on E. coli during storage. Glutamate, glutamine or glutathione did not significantly influence the post-pressure survival over 12 days. Preliminary investigation on cell membrane was further performed through the use of fluorescence probe 1-N-phenylnaphthylamine. Pressure effectively permeabilised cell membrane, whereas calcium showed no effects on membrane permeabilisation.

  13. Development of a Multiplex PCR Assay for Detection of Shiga Toxin-Producing Escherichia coli, Enterohemorrhagic E. coli, and Enteropathogenic E. coli Strains

    PubMed Central

    Botkin, Douglas J.; Galli, Lucía; Sankarapani, Vinoth; Soler, Michael; Rivas, Marta; Torres, Alfredo G.

    2012-01-01

    Escherichia coli O157:H7 and other pathogenic E. coli strains are enteric pathogens associated with food safety threats and which remain a significant cause of morbidity and mortality worldwide. In the current study, we investigated whether enterohemorrhagic E. coli (EHEC), Shiga toxin-producing E. coli (STEC), and enteropathogenic E. coli (EPEC) strains can be rapidly and specifically differentiated with multiplex PCR (mPCR) utilizing selected biomarkers associated with each strain’s respective virulence genotype. Primers were designed to amplify multiple intimin (eae) and long polar fimbriae (lpfA) variants, the bundle-forming pilus gene bfpA, and the Shiga toxin-encoding genes stx1 and stx2. We demonstrated consistent amplification of genes specific to the prototype EHEC O157:H7 EDL933 (lpfA1-3, lpfA2-2, stx1, stx2, and eae-γ) and EPEC O127:H6 E2348/69 (eae-α, lpfA1-1, and bfpA) strains using the optimized mPCR protocol with purified genomic DNA (gDNA). A screen of gDNA from isolates in a diarrheagenic E. coli collection revealed that the mPCR assay was successful in predicting the correct pathotype of EPEC and EHEC clones grouped in the distinctive phylogenetic disease clusters EPEC1 and EHEC1, and was able to differentiate EHEC1 from EHEC2 clusters. The assay detection threshold was 2 × 104 CFU per PCR reaction for EHEC and EPEC. mPCR was also used to screen Argentinean clinical samples from hemolytic uremic syndrome and diarrheal patients, resulting in 91% sensitivity and 84% specificity when compared to established molecular diagnostic procedures. In conclusion, our mPCR methodology permitted differentiation of EPEC, STEC and EHEC strains from other pathogenic E. coli; therefore, the assay becomes an additional tool for rapid diagnosis of these organisms. PMID:22919600

  14. Real-time isothermal detection of Shiga toxin-producing Escherichia coli using recombinase polymerase amplification

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin (Stx) producing E. coli (STEC) are a major family of foodborne pathogens of immense public health, zoonotic and economic significance in the US and worldwide. To date, there are no published reports on use of recombinase polymerase amplification (RPA) for STEC detection. The primary goal...

  15. Characterization of shiga toxin subtypes and virulence genes in Porcine shiga toxin-producing Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Similar to ruminants, swine have been shown to be a reservoir for Shiga toxin-producing Escherichia coli (STEC), and pork products have been linked with outbreaks associated with STEC O157 and O111:H-. STEC strains, isolated in a previous study from fecal samples of late-finisher pigs, belonged to a...

  16. Gamma radiation inactivation of non-0157:H7 shiga-toxin producing Escherichia coli in foods

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Non-O157:H7 serovars of shiga-toxin producing Escherichia coli are emerging foodborne pathogens that have been associated with illness outbreaks and food product recalls on a global basis. Ionizing (gamma) radiation is a nonthermal food safety intervention technology that has been approved for use i...

  17. Shiga toxin-producing escherichia coli: detection, differentiation, and implications for food safety

    Technology Transfer Automated Retrieval System (TEKTRAN)

    All unprocessed food products typically harbor microorganisms. Some foods and the components that go into food production may contain pathogenic microorganisms such as Shiga toxin-producing Escherichia coli (STECs). When consumed, these STECs can cause serious illness or even death. In 2011, an out...

  18. Shiga toxin-producing Escherichia coli in swine: the public health perspective

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing Escherichia coli (STEC) strains are food-borne pathogens that are an important public health concern. STEC infection is associated with severe clinical diseases in humans, including hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS), which can lead to kidney failure ...

  19. Classification of shiga toxin-producing escherichia coli (STEC) serotypes with hyperspectral microscope imagery

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Non-O157:H7 Shiga toxin-producing Escherichia coli (STEC) strains such as O26, O45, O103, O111, O121 and O145 are recognized as serious outbreak to cause human illness due to their toxicity. Since a conventional microbiological method for cell counting is laborious and time-consuming process, optica...

  20. Development of an automated multiplexed immunomagnetic separation system for isolating Shiga toxin-producing Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In recent years, non-O157 Shiga toxin-producing Escherichia coli(STEC) have become an emerging problem. Efforts have been devoted to facilitating and speeding their detection, however, their isolation from high background microbiota foods remains problematic. To solve this problem, immunomagnetic se...

  1. A 7-plex microbead-based immunoassay for serotyping Shiga toxin-producing Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Serotyping of Shiga toxin-producing Escherichia coli (STEC) has been contingent upon the availability of antisera. Here we describe a 7-plex microbead-based immunoassay to simultaneously serotype seven STEC (i.e., belonging to serogroups O26, O45, O103, O111, O121, O145, and O157) by the Luminex xMA...

  2. Evaluation of beef trim sampling methods for detection of Shiga toxin-producing Escherichia coli (STEC)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Presence of Shiga toxin-producing Escherichia coli (STEC) is a major concern in ground beef. Several methods for sampling beef trim prior to grinding are currently used in the beef industry. The purpose of this study was to determine the efficacy of the sampling methods for detecting STEC in beef ...

  3. Drosophila yolk protein produced in E. coli is accumulated by mosquito ovaries.

    PubMed

    Bownes, M; Hurd, H; Büsgen, T; Servay, D; Alvis, S; Popovic, B; Bruce, S; Burns, I; Rothwell, K; Walkinshaw, Malcolm

    2002-10-01

    Despite similar functions, the yolk proteins of the higher dipteran flies and the vitellogenins found in other insects are unrelated at the sequence level and have evolved from different genes. Both are selectively endocytosed into the ovary via receptors belonging to the LDLR receptor subfamily. We cloned the Drosophila yp1 gene into an E. coli expression vector and showed that the yolk protein produced by E. coli is taken up into ovaries of both Drosophila melanogaster and the malaria mosquito Anopheles gambiae, which normally uses vitellogenin. PMID:12230547

  4. Tailor-made ion-imprinted polymer based on functionalized graphene oxide for the preconcentration and determination of trace copper in food samples.

    PubMed

    Liu, Yan; Qiu, Jian; Liu, Zhanchao; Ni, Liang; Jiang, Yinhua; Gong, Chongying; Meng, Xiangguo; Liu, Fangfang; Zhong, Guoxing

    2016-04-01

    A tailor-made Cu(II) ion-imprinted polymer based on large-surface-area graphene oxide sheets has been synthesized for the preconcentration and determination of trace copper from food samples by solid-phase extraction. Attributed to the ultrahigh surface area and hydrophilicity of graphene oxide, the Cu(II) ion-imprinted polymer prepared by the surface ion-imprinting technique exhibited a high binding capacity and a fast adsorption rate under the optimized experimental conditions. In the static adsorption experiments, the maximum adsorption capacity of Cu(II) ion-imprinted polymer is 109.38 mg/g at 25°C, which is much higher than that of the nonimprinted polymer (32.12 mg/g). Meanwhile, the adsorption is very rapid and equilibrium is reached after approximately 30 min. The adsorption mechanism is found to follow Langmuir adsorption model and the pseudo-second-order adsorption process. The Cu(II) ion-imprinted polymer was used for extracting and detecting Cu(II) in food samples combined with graphite flame atomic adsorption spectrometry with high recoveries in the range of 97.6-103.3%. The relative standard deviation and limit of detection of the method were evaluated as 1.2% and 0.37 μg/L, respectively. The results showed that the novel absorbent can be utilized as an effective material for the selective enrichment and determination of Cu(II) from food samples. PMID:26841822

  5. Developing and optimizing bacteriophage treatment to control enterohemorrhagic Escherichia coli on fresh produce.

    PubMed

    Snyder, Abigail B; Perry, Jennifer J; Yousef, Ahmed E

    2016-11-01

    Bacteriophages are potentially useful in controlling foodborne pathogens on minimally processed products since phage application is a non-destructive treatment. The purpose of this study was to evaluate the efficacy of a newly isolated environmental bacteriophage against enterohemorrhagic Escherichia coli on fresh produce, and optimize the treatment with consideration for potential application. Seven anti E. coli O157:H7 EDL933 bacteriophages were isolated from various sources; the most promising was isolated from municipal wastewater. This isolate (designated as E. coli phage OSY-SP) was propagated with the host, in a growth medium, to a titer of 10(8) PFU/ml. Before inoculation into fresh produce, E. coli phage OSY-SP was incubated with the host bacterium, spent medium was filter-sterilized, and the resulting crude lysate was used as a source of phage inocula for preliminary experiments. For optimized testing, phage in the crude lysate was purified by ultra-centrifugation and resuspension in phosphate-buffered saline. Efficacy of phage treatments was determined as a function of fresh produce type (cut green pepper or spinach leaves), treatment time (2 or 5min rinsing), and temperature of holding treated produce (4°C, 25°, or a combination of both temperatures). Cut green pepper was treated with UV light, to eliminate background microbiota, then spot-inoculated with E. coli O157:H7 EDL933 on cut edges, and the inoculum was allowed to dry. Because of its susceptibility to damage, baby spinach leaves were not subjected to a decontamination treatment. These leaves were inoculated with the green fluorescent protein-labeled E. coli O157:H7 B6-914 to facilitate inoculum enumeration in the presence of background microbiota. Phage suspension was applied to the inoculated fresh produce that was subsequently held for three days under variable storage conditions. The optimized phage treatment decreased the populations of pathogenic E. coli by 2.4-3.0logCFU/g on cut green

  6. An OXA-48-producing Escherichia coli isolated from a Danish patient with no hospitalization abroad.

    PubMed

    Gedebjerg, Anne; Hasman, Henrik; Sørensen, Christian Møller; Wang, Mikala

    2015-08-01

    Carbapenemase-producing organisms are disseminating globally and are now emerging as a worrying threat in Scandinavia. Before August 2013, OXA-48-producing organisms had not been detected in Danish patients. Here we report the isolation of an ST746 OXA-48-producing Escherichia coli with the plasmid pOXA-48a carrying the blaOXA-48 gene isolated from a Danish patient without history of hospitalization abroad. The patient reported tourist travel to Egypt and Turkey. The potential acquisition of carbapenemase-producing organisms by ingestion of contaminated food is discussed.

  7. First Report of Klebsiella pneumoniae-Carbapenemase-3-Producing Escherichia coli ST479 in Poland.

    PubMed

    Ojdana, Dominika; Sacha, Paweł; Olszańska, Dorota; Majewski, Piotr; Wieczorek, Piotr; Jaworowska, Jadwiga; Sieńko, Anna; Jurczak, Anna; Tryniszewska, Elżbieta

    2015-01-01

    An increase in the antibiotic resistance among members of the Enterobacteriaceae family has been observed worldwide. Multidrug-resistant Gram-negative rods are increasingly reported. The treatment of infections caused by Escherichia coli and other Enterobacteriaceae has become an important clinical problem associated with reduced therapeutic possibilities. Antimicrobial carbapenems are considered the last line of defense against multidrug-resistant Gram-negative bacteria. Unfortunately, an increase of carbapenem resistance due to the production of Klebsiella pneumoniae carbapenemase (KPC) enzymes has been observed. In this study we describe the ability of E. coli to produce carbapenemase enzymes based on the results of the combination disc assay with boronic acid performed according to guidelines established by the European Community on Antimicrobial Susceptibility Testing (EUCAST) and the biochemical Carba NP test. Moreover, we evaluated the presence of genes responsible for the production of carbapenemases (bla KPC, bla VIM, bla IMP, bla OXA-48) and genes encoding other β-lactamases (bla SHV, bla TEM, bla CTX-M) among E. coli isolate. The tested isolate of E. coli that possessed the bla KPC-3 and bla TEM-34 genes was identified. The tested strain exhibited susceptibility to colistin (0.38 μg/mL) and tigecycline (1 μg/mL). This is the first detection of bla KPC-3 in an E. coli ST479 in Poland. PMID:26339599

  8. Commensal E. coli Stx2 lysogens produce high levels of phages after spontaneous prophage induction.

    PubMed

    Iversen, Hildegunn; L' Abée-Lund, Trine M; Aspholm, Marina; Arnesen, Lotte P S; Lindbäck, Toril

    2015-01-01

    Enterohemorrhagic E. coli (EHEC) is a food-borne pathogen that causes disease ranging from uncomplicated diarrhea to life-threatening hemolytic uremic syndrome (HUS) and nervous system complications. Shiga toxin 2 (Stx2) is the major virulence factor of EHEC and is critical for development of HUS. The genes encoding Stx2 are carried by lambdoid bacteriophages and the toxin production is tightly linked to the production of phages during lytic cycle. It has previously been suggested that commensal E. coli could amplify the production of Stx2-phages and contribute to the severity of disease. In this study we examined the susceptibility of commensal E. coli strains to the Stx2-converting phage ϕ734, isolated from a highly virulent EHEC O103:H25 (NIPH-11060424). Among 38 commensal E. coli strains from healthy children below 5 years, 15 were lysogenized by the ϕ734 phage, whereas lytic infection was not observed. Three of the commensal E. coli ϕ734 lysogens were tested for stability, and appeared stable and retained the phage for at least 10 cultural passages. When induced to enter lytic cycle by H2O2 treatment, 8 out of 13 commensal lysogens produced more ϕ734 phages than NIPH-11060424. Strikingly, five of them even spontaneously (non-induced) produced higher levels of phage than the H2O2 induced NIPH-11060424. An especially high frequency of HUS (60%) was seen among children infected by NIPH-11060424 during the outbreak in 2006. Based on our findings, a high Stx2 production by commensal E. coli lysogens cannot be ruled out as a contributor to the high frequency of HUS during this outbreak. PMID:25692100

  9. Outbreak Caused by NDM-1- and RmtB-Producing Escherichia coli in Bulgaria

    PubMed Central

    Poirel, Laurent; Savov, Encho; Nazli, Arzu; Trifonova, Angelina; Todorova, Iva; Gergova, Ivanka

    2014-01-01

    Twelve consecutive carbapenem-resistant Escherichia coli isolates were recovered from patients (infection or colonization) hospitalized between March and September 2012 in different units at a hospital in Bulgaria. They all produced the carbapenemase NDM-1 and the extended-spectrum-β-lactamase CTX-M-15, together with the 16S rRNA methylase RmtB, conferring high-level resistance to all aminoglycosides. All those isolates were clonally related and belonged to the same sequence type, ST101. In addition to being the first to identify NDM-producing isolates in Bulgaria, this is the very first study reporting an outbreak of NDM-1-producing E. coli in the world. PMID:24514099

  10. Isolation of Shiga Toxin-Producing Escherichia coli from a South American Camelid (Lama guanicoe) with Diarrhea

    PubMed Central

    Mercado, E. C.; Rodríguez, S. M.; Elizondo, A. M.; Marcoppido, G.; Parreño, V.

    2004-01-01

    Shiga toxin-producing Escherichia coli belonging to serotype O26:H11 was isolated from a 2-month-old guanaco with severe watery diarrhea. E. coli colonies carried the stx1 and eae genes, showed localized adherence to HEp-2 cells, and produced enterohemolysin. A serological response to lipopolysaccharide O26 was observed at the onset of diarrhea. PMID:15472347

  11. Biofilm-Forming Abilities of Shiga Toxin-Producing Escherichia coli Isolates Associated with Human Infections

    PubMed Central

    Vogeleer, Philippe; Tremblay, Yannick D. N.; Jubelin, Grégory; Jacques, Mario

    2015-01-01

    Forming biofilms may be a survival strategy of Shiga toxin-producing Escherichia coli to enable it to persist in the environment and the food industry. Here, we evaluate and characterize the biofilm-forming ability of 39 isolates of Shiga toxin-producing Escherichia coli isolates recovered from human infection and belonging to seropathotypes A, B, or C. The presence and/or production of biofilm factors such as curli, cellulose, autotransporter, and fimbriae were investigated. The polymeric matrix of these biofilms was analyzed by confocal microscopy and by enzymatic digestion. Cell viability and matrix integrity were examined after sanitizer treatments. Isolates of the seropathotype A (O157:H7 and O157:NM), which have the highest relative incidence of human infection, had a greater ability to form biofilms than isolates of seropathotype B or C. Seropathotype A isolates were unique in their ability to produce cellulose and poly-N-acetylglucosamine. The integrity of the biofilms was dependent on proteins. Two autotransporter genes, ehaB and espP, and two fimbrial genes, z1538 and lpf2, were identified as potential genetic determinants for biofilm formation. Interestingly, the ability of several isolates from seropathotype A to form biofilms was associated with their ability to agglutinate yeast in a mannose-independent manner. We consider this an unidentified biofilm-associated factor produced by those isolates. Treatment with sanitizers reduced the viability of Shiga toxin-producing Escherichia coli but did not completely remove the biofilm matrix. Overall, our data indicate that biofilm formation could contribute to the persistence of Shiga toxin-producing Escherichia coli and specifically seropathotype A isolates in the environment. PMID:26712549

  12. Prevalence and characteristics of intimin-producing Escherichia coli strains isolated from healthy chickens in Korea.

    PubMed

    Oh, J-Y; Kang, M-S; An, B-K; Shin, E-G; Kim, M-J; Kim, Y-J; Kwon, Y-K

    2012-10-01

    Virulent Escherichia coli strains have commonly been associated with diarrheal illness in humans and animals. Typical enteropathogenic Escherichia coli (EPEC) with intimin gene (eaeA) and E. coli adherence factor plasmid, or atypical EPEC with only eaeA have been implicated in human cases. In the present study, we investigated the prevalence of virulence-associated genes including eaeA in the E. coli strains isolated from cloacal specimens of 184 chicken flocks in 7 provinces in Korea between 2009 and 2010. When 7 virulence genes (VT1, VT2, LT, and ST for enterotoxigenic E. coli; eaeA and bfpA for enteropathogenic E. coli; and aggR for enteroaggregative E. coli) were screened by multiplex PCR, a total of 30 E. coli strains carrying only the eaeA gene were detected from 184 flocks that were identified as atypical enteropathogenic Escherichia coli (aEPEC). The aEPEC strains were analyzed by eae subtyping, phylogenetic grouping PCR, and serotyping. Twelve (40%) of 30 aEPEC strains possessed an eae-β subtype, followed by θ (30%), ε (16.7%), and β1 (13.3%). Eight (26.7%) of 30 aEPEC strains were designated into the phylogenetic group A. Two (6.7%) and 3 (10%) aEPEC strains were classified into the phylogenetic group B2 and D, respectively. A total of 15 (50%) aEPEC strains were serotyped to groups O24, O25, O26, O71, O80, O103, and O157, and the remaining strains were nontypeable. In analyzing the genetic diversity among the 30 aEPEC isolates by the pulsed-field gel electrophoresis method with XbaI-digestion, the pulsed-field gel electrophoresis profiling produced 20 different patterns, but isolates within the same group did not show clear geographic or breed relationships. Our data indicate that healthy chickens may constitute an important natural reservoir of aEPEC strains, and suggest that transmission to humans could not be excluded. PMID:22991525

  13. Surface properties of the Vero cytotoxin-producing Escherichia coli O157:H7.

    PubMed Central

    Sherman, P; Soni, R; Petric, M; Karmali, M

    1987-01-01

    Strains of Escherichia coli serotype O157:H7 are Vero cytotoxin-producing enteric pathogens which have been associated with sporadic cases and outbreaks of hemorrhagic colitis and with the hemolytic uremic syndrome in humans. In addition to toxin production, adherence of many pathogenic bacteria to intestinal mucosal surfaces is a critical primary step in the pathogenesis of diarrheal diseases. Although E. coli serotype O157:H7 organisms adhere to intestinal epithelia of orally infected animals in a pattern morphologically identical to that previously described in adherent, effacing E. coli infections, the mechanisms of bacterial adherence are not known. To determine the cell surface adhesins which mediate attachment of E. coli O157:H7 to epithelial surfaces, we evaluated the surface properties of these organisms. Five strains isolated from children with the hemolytic uremic syndrome were grown both in broth cultures and on agar media. Adherence and invasion of E. coli O157:H7 in Intestine 407 and HEp-2 epithelial cell lines was quantitated using an enteroinvasive E. coli strain (serotype O164:NM) as a control. Cell surface properties of E. coli O157:H7 were evaluated by agglutination of a series of erythrocytes, transmission electron microscopy, DEAE-ion-exchange chromatography, and hydrophobic interaction chromatography. E. coli O157:H7 strains adhered to but did not invade either Intestine 407 or HEp-2 cells. Homologous O157:H7 rabbit antiserum blocked attachment of bacteria to tissue culture cells, in contrast to heterologous antiserum and preimmune rabbit serum, which did not inhibit attachment of E. coli O157:H7. None of the five O15:H7 isolates mediated mannose-resistant hemagglutination under any of the in vitro culture conditions. One isolate mediated mannose-sensitive hemagglutination after serial passage in broth cultures. Pili and fibrillae were not visualized by electron microscopy on nonhemagglutinating organisms, but pili were demonstrated on the one

  14. The discovery of cholera - like enterotoxins produced by Escherichia coli causing secretory diarrhoea in humans

    PubMed Central

    Sack, R. Bradley

    2011-01-01

    Non-vibrio cholera has been recognized as a clinical entity for as long as cholera was known to be caused by Vibrio cholerae. Until 1968, the aetiologic agent of this syndrome was not known. Following a series of studies in patients with non-vibrio cholera it was found that these patients had large concentrations of Escherichia coli in the small bowel and stools which produced cholera toxin-like enterotoxins, and had fluid and electrolyte transport abnormalities in the small bowel similar to patients with documented cholera. Furthermore, these patients developed antibodies to the cholera-like enterotoxin. Later studies showed that these strains, when fed to volunteers produced a cholera-like disease and that two enterotoxins were found to be produced by these organisms: a heat-labile enterotoxin (LT) which is nearly identical to cholera toxin, and a heat-stable enterotoxin (ST), a small molecular weight polypeptide. E. coli that produced one or both of these enterotoxins were designated enterotoxigenic E. coli (ETEC). ETEC are now known not only to cause a severe cholera-like illness, but to be the most common bacterial cause of acute diarrhoea in children in the developing world, and to be the most common cause of travellers’ diarrhoea in persons who visit the developing world. PMID:21415491

  15. Extended-Spectrum-β-Lactamase-Producing Escherichia coli as Intestinal Colonizers in the German Community

    PubMed Central

    Nickel, Silke; Pfeifer, Yvonne; Eller, Christoph; Krupa, Elzbieta; Lehner-Reindl, Verena; Höller, Christiane

    2014-01-01

    We determined the presence of extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli among 3,344 study participants from the German community. Intestinal colonization was detected in 211 persons (6.3%), without significant differences among the different age groups. The majority (95.2%) of isolates harbored CTX-M-type ESBL, with CTX-M-15 (46%) and CTX-M-1 (24.2%) as the most common types. The finding of ESBL producers and one isolate additionally producing carbapenemase OXA-244 indicates a risk of dissemination of resistant bacteria outside the hospitals. PMID:24295972

  16. Tailor-Made Stable Zr(IV)-Based Metal-Organic Frameworks for Laser Desorption/Ionization Mass Spectrometry Analysis of Small Molecules and Simultaneous Enrichment of Phosphopeptides.

    PubMed

    Chen, Lianfang; Ou, Junjie; Wang, Hongwei; Liu, Zhongshan; Ye, Mingliang; Zou, Hanfa

    2016-08-10

    Although thousands of metal-organic frameworks (MOFs) have been fabricated and widely applied in gas storage/separations, adsorption, catalysis, and so on, few kinds of MOFs have been used as adsorption materials while simultaneously serving as matrixes to analyze small molecules for laser desorption/ionization mass spectrometry (LDI-MS). Herein, a new concept is introduced to design and synthesize MOFs as both adsorption materials and matrixes according to the structure of ligands and common matrixes. The proof of concept design was demonstrated by selection of 2,5-pyridinedicarboxylic acid (PDC) and 2,5-dihydroxyterephthalic acid (DHT) as ligands for synthesis of MOFs. Two Zr(IV)-based MOFs of UiO-66-PDC and UiO-66-(OH)2 were synthesized and applied for the first time as new matrixes for analysis of small molecules by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Both of them showed low matrix interferences, high ionization efficiency, and good reproducibility when used as matrixes. A variety of small molecules, including saccharides, amino acids, nucleosides, peptides, alkaline drugs, and natural products, were analyzed. In addition, UiO-66-(OH)2 exhibited potential for application in the quantitative determination of glucose and pyridoxal 5'-phosphate. Furthermore, thanks to its intrinsically large surface area and highly ordered pores, UiO-66-(OH)2 also showed sensitive and specific enrichment of phosphopeptides prior to MS analysis. These results demonstrated that this strategy can be used to efficiently screen tailor-made MOFs as matrixes to analyze small molecules by MALDI-TOF-MS. PMID:27427857

  17. Combining Transcranial Direct Current Stimulation and Tailor-Made Notched Music Training to Decrease Tinnitus-Related Distress – A Pilot Study

    PubMed Central

    Teismann, Henning; Wollbrink, Andreas; Okamoto, Hidehiko; Schlaug, Gottfried; Rudack, Claudia; Pantev, Christo

    2014-01-01

    The central auditory system has a crucial role in tinnitus generation and maintenance. Curative treatments for tinnitus do not yet exist. However, recent attempts in the therapeutic application of both acoustic stimulation/training procedures and electric/magnetic brain stimulation techniques have yielded promising results. Here, for the first time we combined tailor-made notched music training (TMNMT) with transcranial direct current stimulation (tDCS) in an effort to modulate TMNMT efficacy in the treatment of 32 patients with tonal tinnitus and without severe hearing loss. TMNMT is characterized by regular listening to so-called notched music, which is generated by digitally removing the frequency band of one octave width centered at the individual tinnitus frequency. TMNMT was applied for 10 subsequent days (2.5 hours of daily treatment). During the initial 5 days of treatment and the initial 30 minutes of TMNMT sessions, tDCS (current strength: 2 mA; anodal (N = 10) vs. cathodal (N = 11) vs. sham (N = 11) groups) was applied simultaneously. The active electrode was placed on the head surface over left auditory cortex; the reference electrode was put over right supra-orbital cortex. To evaluate treatment outcome, tinnitus-related distress and perceived tinnitus loudness were assessed using standardized tinnitus questionnaires and a visual analogue scale. The results showed a significant treatment effect reflected in the Tinnitus Handicap Questionnaire that was largest after 5 days of treatment. This effect remained significant at the end of follow-up 31 days after treatment cessation. Crucially, tDCS did not significantly modulate treatment efficacy - it did not make a difference whether anodal, cathodal, or sham tDCS was applied. Possible explanations for the findings and functional modifications of the experimental design for future studies (e.g. the selection of control conditions) are discussed. PMID:24587113

  18. Review of Shiga-toxin-producing Escherichia coli (STEC) and their significance in dairy production.

    PubMed

    Farrokh, Choreh; Jordan, Kieran; Auvray, Frederic; Glass, Kathleen; Oppegaard, Hanne; Raynaud, Sabrina; Thevenot, Delphine; Condron, Robin; De Reu, Koen; Govaris, Alexander; Heggum, Klaus; Heyndrickx, Marc; Hummerjohann, Joerg; Lindsay, Denise; Miszczycha, Stephane; Moussiegt, Sylvie; Verstraete, Karen; Cerf, Olivier

    2013-03-15

    The involvement of the pathogenic Shiga-toxin-producing Escherichia coli (STEC; also called verocytotoxic-producing E. coli or VTEC) in sporadic cases and disease outbreaks is presently increasing. Infrequent cases are due to ingestion of milk and dairy products. As ruminants are healthy carriers of STEC and most dairy products may provide these bacteria with favourable conditions for their growth, milk and dairy products are a potential source of STEC. But not all STEC serotypes are pathogens; only relatively small numbers in the entire family of STEC are pathogenic. This review focuses on the recent advances in understanding of STEC and their significance in milk and dairy products. It is intended to gather the information that is needed to understand how these bacteria are described, detected and characterised, how they contaminate milk and grow in dairy products, and how the dairy industry can prevent them from affecting the consumer.

  19. Thiol-independent activity of a cholesterol-binding enterohemolysin produced by enteropathogenic Escherichia coli.

    PubMed

    Figueirêdo, P M S; Catani, C F; Yano, T

    2003-11-01

    Enterohemolysin produced by Escherichia coli associated with infant diarrhea showed characteristics similar to those of thiol-activated hemolysins produced by Gram-positive bacteria, including inactivation by cholesterol, lytic activity towards eukaryotic cells and thermoinstability. However, enterohemolysin activity was not inactivated by oxidation or by SH group-blocking agents (1 mM HgCl2, 1 mM iodoacetic acid) and the hemolysin (100 microg/ml) was not lethal to mice, in contrast to the lethality of the thiol-activated hemolysin family to animals. Earlier reports showed that intravenous injection of partially purified streptolysin O preparations (0.2 microg) was rapidly lethal to mice. These results suggest that E. coli enterohemolysin is not a thiol-activated hemolysin, despite its ability to bind cholesterol, probably due to the absence of free thiol-group(s) that characterize the active form of the thiol-activated hemolysin molecule.

  20. An outbreak of Vero cytotoxin producing Escherichia coli O157 infection associated with takeaway sandwiches.

    PubMed

    McDonnell, R J; Rampling, A; Crook, S; Cockcroft, P M; Wilshaw, G A; Cheasty, T; Stuart, J

    1997-12-12

    An outbreak of food poisoning due to Escherichia coli O157 phage type 2 Vero cytotoxin 2 affected 26 people in southern counties of England in May and June 1995. The organism was isolated from faecal specimens from 23 patients, 16 of whom lived in Dorset and seven in Hampshire. Isolates were indistinguishable by phage typing, Vero cytotoxin gene typing, restriction fragment length polymorphism, and pulsed field gel electrophoresis. Three associated cases, linked epidemiologically to the outbreak, were confirmed serologically by detection of antibodies to E. coli O157 lipopolysaccharide. Twenty-two of the 26 patients were adults: four were admitted to hospital with haemorrhagic colitis. Four cases were children: two were admitted to hospital with haemolytic uraemic syndrome (HUS). There were no deaths. Although E. coli O157 was not isolated from any food samples, illness was associated with having eaten cold meats in sandwiches bought from two sandwich producers, in Weymouth and in Portsmouth. Both shops were supplied by the same wholesaler, who kept no records and obtained cooked meats from several sources in packs that did not carry adequate identification marks. It was, therefore, impossible to trace back to the original producer or to investigate further to determine the origin of contamination with E. coli O157. To protect the public health it is essential that all wholesale packs of ready-to-eat food carry date codes and the producer's identification mark. Detailed record keeping should be part of hazard analysis critical control point (HACCP) systems and should be maintained throughout the chain of distribution from the producer to retail outlets.

  1. Shiga Toxin-Producing Escherichia coli in Peruvian Children with Bloody Diarrhea

    PubMed Central

    Llanos, Alejandro; Lee, Jorge; López, Francisco; Contreras, Carmen; Barletta, Francesca; Chea-Woo, Elsa; Ugarte, Claudia; Cleary, Thomas G.; Ochoa, Theresa J.

    2012-01-01

    Shiga toxin-producing Escherichia coli (STEC) are not routinely sought in clinical laboratories in developing counties. Among 131 bloody diarrhea samples in Peruvian children <5y of age, STEC was found in 9.2% and was associated with absence of fever, an observation that may increase suspicion of these pathogens. Because of the significant prevalence of STEC locally, proper diagnostics methods should be implemented in the region. PMID:22315000

  2. Multidrug Resistant CTX-M-Producing Escherichia coli: A Growing Threat among HIV Patients in India.

    PubMed

    Padmavathy, Kesavaram; Padma, Krishnan; Rajasekaran, Sikhamani

    2016-01-01

    Extended Spectrum β-Lactamases (ESBLs) confer resistance to third-generation cephalosporins and CTX-M types have emerged as the most prominent ESBLs worldwide. This study was designed to determine the prevalence of CTX-M positive ESBL-producing urinary E. coli isolates from HIV patients and to establish the association of multidrug resistance, phylogeny, and virulence profile with CTX-M production. A total of 57 ESBL producers identified among 76 E. coli strains isolated from HIV patients from South India were screened for bla CTX-M, AmpC production, multidrug resistance, and nine virulence associated genes (VAGs), fimH, pap, afa/dra, sfa/foc, iutA, fyuA, iroN, usp, and kpsMII. The majority (70.2%) of the ESBL producers harbored bla CTX-M and were AmpC coproducers. Among the CTX-M producers, 47.5% were found to be UPEC, 10% harbored as many as 7 VAGs, and 45% possessed kpsMII. Multidrug resistance (CIP(R)SXT(R)GEN(R)) was significantly more common among the CTX-M producers compared to the nonproducers (70% versus 41.2%). However, 71.4% of the multidrug resistant CTX-M producers exhibited susceptibility to nitrofurantoin thereby making it an effective alternative to cephalosporins/fluoroquinolones. The emergence of CTX-M-producing highly virulent, multidrug resistant uropathogenic E. coli is of significant public health concern in countries like India with a high burden of HIV/AIDS.

  3. Sorbitol non-fermenting shiga toxin-producing Escherichia coli in cattle on smallholdings.

    PubMed

    Islam, M Z; Christensen, J P; Biswas, P K

    2015-01-01

    We investigated faecal samples collected from the rectum of 518 cattle on 371 randomly selected smallholdings in Bangladesh for the presence of sorbitol non-fermenting (SN-F) shiga toxin-producing Escherichia coli (STEC). The SN-F isolates were tested for the presence of rfb O157, stx1, stx2, eae and hlyA genes by polymerase chain reaction (PCR). Seven SN-F isolates lacking these genes were profiled by pulsed-field gel electrophoresis (PFGE) to verify their clonality. SN-F E. coli was identified in 44 [8·5%, 95% confidence interval (CI) 6·4-11·2] samples; of these, 28 (5·4%, 95% CI 3·8-7·7) had shiga toxin-producing strains, although only two carried the rfb O157 gene. Thirteen isolates carried the hlyA gene while 18 harboured the eae gene. Based on PFGE, six pulsotypes were observed among the seven isolates that had no virulence genes. To the best of our knowledge this is the first report on shiga toxin-producing E. coli from direct rectal faecal samples of cattle on smallholdings.

  4. A PCR-ELISA for detecting Shiga toxin-producing Escherichia coli.

    PubMed

    Ge, Beilei; Zhao, Shaohua; Hall, Robert; Meng, Jianghong

    2002-03-01

    A sensitive and specific PCR-ELISA was developed to detect Escherichia coli O157:H7 and other Shiga toxin-producing E. coli (STEC) in food. The assay was based on the incorporation of digoxigenin-labeled dUTP and a biotin-labeled primer specific for Shiga toxin genes during PCR amplification. The labeled PCR products were bound to streptavidin-coated wells of a microtiter plate and detected by an ELISA. The specificity of the PCR was determined using 39 bacterial strains, including STEC, enteropathogenic E. coli, E. coli K12, and Salmonella. All of the STEC strains were positive, and non-STEC organisms were negative. The ELISA detecting system was able to increase the sensitivity of the PCR assay by up to 100-fold, compared with a conventional gel electrophoresis. The detection limit of the PCR-ELISA was 0.1-10 CFU dependent upon STEC serotypes, and genotypes of Shiga toxins. With the aid of a simple DNA extraction system, PrepMan, the PCR-ELISA was able to detect ca. 10(5) CFU of STEC per gram of ground beef without any culture enrichment. The entire procedure took about 6 h. Because of its microtiter plate format, PCR-ELISA is particularly suitable for large-scale screening and compatible with future automation.

  5. Prevalence and characteristics of Shiga toxin-producing Escherichia coli in Swiss raw milk cheeses collected at producer level.

    PubMed

    Stephan, R; Schumacher, S; Corti, S; Krause, G; Danuser, J; Beutin, L

    2008-07-01

    The aim of this study was to describe the prevalence, serotypes, and virulence genes of Shiga toxin-producing Escherichia coli (STEC) isolated from raw milk cheese samples collected at the producer level with the purpose of determining whether raw milk cheeses in Switzerland represent a potential source of STEC pathogenic for humans. Raw milk cheese samples (soft cheese, n = 52; semihard and hard cheese, n = 744; all produced from Swiss cows', goats', and sheep's milk) collected at the producer level throughout Switzerland within the national sampling plan during the period of March 2006 to December 2007 were analyzed. Of the 432 cheese samples obtained in the year 2006 and the 364 samples obtained in the year 2007, 16 (3.7%) and 23 (6.3%), respectively, were found to be stx positive. By colony dot-blot hybridization, non-O157 STEC strains were isolated from 16 samples. Of the 16 strains, 11 were typed into 7 E. coli O groups (O2, O15, O22, O91, O109, O113, O174), whereas 5 strains were nontypeable (ONT). Among the 16 STEC strains analyzed, stx(1) and stx(2) variants were detected in 1 and 15 strains, respectively. Out of the 15 strains with genes encoding for the Stx2 group, 4 strains were positive for stx(2), 6 strains for stx(2d2), 2 strains for stx(2-O118), 1 strain for stx(2-06), 1 strain for stx(2g), 1 strain for stx(2) and stx(2d2), and 1 strain for stx(2) and stx(2g). Furthermore, 3 STEC strains harbored E-hlyA as a further putative virulence factor. None of the strains tested positive for eae (intimin). Results obtained in this work reinforce the suggestion that semihard raw milk cheese may be a potential vehicle for transmission of pathogenic STEC to humans.

  6. Design of a recombinant Escherichia coli for producing L-phenylalanine from glycerol.

    PubMed

    Thongchuang, Mayura; Pongsawasdi, Piamsook; Chisti, Yusuf; Packdibamrung, Kanoktip

    2012-10-01

    A recombinant Escherichia coli was engineered to produce the commercially important amino acid L-phenylalanine (L-Phe) using glycerol as the carbon source. Compared to the conventionally used glucose and sucrose, glycerol is a less expensive carbon source. As phenylalanine dehydrogenase (PheDH) activity is involved in the last step of L-Phe synthesis in E. coli, a phenylalanine dehydrogenase gene (phedh) from the thermotolerant Bacillus lentus was cloned into pRSFDuet-1 (pPheDH) and expressed in E. coli BL21(DE3). The resulting clone had a limited ability to produce L-Phe from glycerol, possibly because of a poor glycerol uptake by the cell, or an inability to excrete L-Phe, or both. Therefore, yddG gene encoding an aromatic amino acid exporter and glpF gene encoding a glycerol transport facilitator were coexpressed with the phedh in a reengineered E. coli. In a glycerol medium, the maximum L-Phe production rates of the clones pPY (phedh and yddG genes) and pPYF (phedh, yddG and glpF genes) were 1.4- and 1.8-fold higher than the maximum production rate of the pPheDH clone. The better producing pPYF clone was further evaluated in a 5 l stirred-tank fermenter (37 °C, an aeration rate of 1 vvm, an agitation speed of 400 rpm). In the fermenter, the maximum concentration of L-Phe (366 mg/l) was achieved in a much shorter period compared to in the shake flasks. In the latter, the highest titer of L-Phe was only 76 % of the maximum value attained in the fermenter. PMID:22806734

  7. Extended spectrum betalactamase producing Enteroaggregative Escherichia coli from young children in Iran

    PubMed Central

    Khoshvaght, Hakimeh; Zeighami, Habib

    2014-01-01

    Aim The aim of this study was to investigate the frequency of betalactamase producing EAEC isolates among young children with diarrhea in Zanjan, Iran. Background Entero aggregative Escherichia coli (EAEC) is an emerging enteric pathogen associated with acute and persistent diarrhea and the evolution and spread of acquired extended spectrum betalactamases (ESBLs) among these strains has become a serious problem in the management of infectious diseases in developing countries. Patients and methods During the period from March 2011 to January 2012, 140 isolates of E. coli from diarrheal children aged 0–60 months and 90 isolates from age-matched controls without diarrhea were investigated for EAEC using PCR. Antimicrobial susceptibility testing was performed as CLSI guidelines and betalactamase genes, including bla TEM, bla CTX-M, bla IMP, bla VIM and bla NDM-1 investigated in EAEC isolates. Results In this study, EAEC was detected with slightly higher frequency in children with (8%) than in children without (4.6%) diarrhea (P > 0.05). Diarrheagenic E. coli exhibited high level resistance to aztreonam (80.7%), amoxicillin (74.4%) and tetracycline (69.3%). Also, 86.4% of E. coli isolates were resistant to at least three different classes of antimicrobial agents and considered as multidrug resistance. Molecular characterization of betalactamase genes showed that bla TEM was the most frequently isolated betalactamase. It was detected in 78.9% of ESBL producing EAEC isolates. Also, the frequency of bla CTX-M was 63.1% (12/19) and 8 (42.1%) isolates carried the bla TEM and bla CTX-M, simultaneously. None MBL producing EAEC was detected in our study. Conclusion Our results indicate that ESBLs especially bla TEM and bla CTX-M are widespread among EAEC isolates and appropriate surveillance and control measures are essential to prevent further dissemination of betalactamases in our country. PMID:24834305

  8. Diminazene aceturate: an antibacterial agent for Shiga-toxin-producing Escherichia coli O157:H7

    PubMed Central

    Wu, Si-Ying; Park, Gil-Yong; Kim, So-Hee; Hulme, John; An, Seong Soo A

    2016-01-01

    The aim of this study was to investigate the bacteriostatic and bactericidal effects of diminazene aceturate (DA) against five strains of pathogenic bacteria and two strains of nonpathogenic bacteria. The results showed that 5 μg/mL of DA suppressed the growth of pathogenic Escherichia coli by as much as 77% compared with the controls. Enterohemorrhagic E. coli EDL933 (an E. coli O157:H7 strain) was the most sensitive to DA with a minimum inhibitory concentration of 20 μg/mL. Additional investigations showed that DA induced the highest level of intracellular reactive oxygen species in EDL933. A positive correlation between the reactive oxygen species levels and DA concentration was demonstrated. DA (5 μg/mL) was also a potent uncoupler, inducing a stationary phase collapse (70%–75%) in both strains of E. coli O157:H7. Further investigation showed that the collapse was due to the NaCl:DA ratio in the broth and was potassium ion dependent. A protease screening assay was conducted to elucidate the underlying mechanism. It was found that at neutral pH, the hydrolysis of H-Asp-pNA increased by a factor of 2–3 in the presence of DA, implying that DA causes dysregulation of the proton motive force and a decrease in cellular pH. Finally, a commercial verotoxin test showed that DA did not significantly increase toxin production in EDL933 and was a suitable antibacterial agent for Shiga-toxin-producing E. coli. PMID:27789937

  9. Molecular Diversity and Plasmid Analysis of KPC-Producing Escherichia coli.

    PubMed

    Chavda, Kalyan D; Chen, Liang; Jacobs, Michael R; Bonomo, Robert A; Kreiswirth, Barry N

    2016-07-01

    The emergence and spread of Klebsiella pneumoniae carbapenemase (KPC) among Enterobacteriaceae presents a major public health threat to the world. Although not as common as in K. pneumoniae, KPC is also found in Escherichia coli strains. Here, we genetically characterized 9 carbapenem-resistant E. coli strains isolated from six hospitals in the United States and completely sequenced their blaKPC-harboring plasmids. The nine strains were isolated from different geographical locations and belonged to 8 different E. coli sequence types. Seven blaKPC-harboring plasmids belonged to four different known incompatibility groups (IncN, -FIA, -FIIK2, and -FIIK1) and ranged in size from ∼16 kb to ∼241 kb. In this analysis, we also identified two plasmids that have novel replicons: (i) pBK28610, which is similar to p34978-3 with an insertion of Tn4401b, and (ii) pBK31611, which does not have an apparent homologue in the GenBank database. Moreover, we report the emergence of a pKP048-like plasmid, pBK34397, in E. coli in the United States. Meanwhile, we also found examples of interspecies spread of blaKPC plasmids, as pBK34592 is identical to pBK30683, isolated from K. pneumoniae In addition, we discovered examples of acquisition (pBK32602 acquired an ∼46-kb fragment including a novel replication gene, along with Tn4401b and other resistance genes) and/or loss (pKpQIL-Ec has a 14.5-kb deletion compared to pKpQIL-10 and pBK33689) of DNA, demonstrating the plasticity of these plasmids and their rapid evolution in the clinic. Overall, our study shows that the spread of blaKPC-producing E. coli is largely due to horizontal transfer of blaKPC-harboring plasmids and related mobile elements into diverse genetic backgrounds. PMID:27114279

  10. PL3 Amidase, a Tailor-made Lysin Constructed by Domain Shuffling with Potent Killing Activity against Pneumococci and Related Species

    PubMed Central

    Blázquez, Blas; Fresco-Taboada, Alba; Iglesias-Bexiga, Manuel; Menéndez, Margarita; García, Pedro

    2016-01-01

    that the structure/function-based domain shuffling approach is a successful method to construct tailor-made endolysins with higher bactericidal activities than their parental enzymes. PMID:27516758

  11. PL3 Amidase, a Tailor-made Lysin Constructed by Domain Shuffling with Potent Killing Activity against Pneumococci and Related Species.

    PubMed

    Blázquez, Blas; Fresco-Taboada, Alba; Iglesias-Bexiga, Manuel; Menéndez, Margarita; García, Pedro

    2016-01-01

    that the structure/function-based domain shuffling approach is a successful method to construct tailor-made endolysins with higher bactericidal activities than their parental enzymes. PMID:27516758

  12. The polymorphic aggregative phenotype of Shiga toxin-producing Escherichia coli O111 depends on rpoS and curli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Escherichia coli O111 is an emerging non-O157:H7 Shiga toxin-producing E. coli (STEC). We previously reported that outbreak and environmental, but not sporadic case, strains of STEC O111 share a distinct aggregation phenotype (M. E. Diodati, A. H. Bates, M. B. Cooley, S. Walker, R. E. Mandrell, and ...

  13. Shiga toxin-producing Escherichia coli and rectoanal junction persistence in ruminants: a study of bacterial-epithelial interactions.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Escherichia coli O157:H7 (O157) was the first Shiga toxin-producing E. coli serotype to be associated with bloody diarrhea or hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS) in humans. It has since been implicated in several outbreaks in the U.S. and globally. Non-O157 STEC have not bee...

  14. Detection of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Market-Ready Chickens in Zambia.

    PubMed

    Chishimba, K; Hang'ombe, B M; Muzandu, K; Mshana, S E; Matee, M I; Nakajima, C; Suzuki, Y

    2016-01-01

    The frequent administering of antibiotics in the treatment of poultry diseases may contribute to emergence of antimicrobial-resistant strains. The objective of this study was to detect the presence of extended-spectrum β-lactamase- (ESBL-) producing Escherichia coli in poultry in Zambia. A total of 384 poultry samples were collected and analyzed for ESBL-producing Escherichia coli. The cultured E. coli isolates were subjected to antimicrobial susceptibility tests and the polymerase chain reaction for detection of bla CTX-M, bla SHV, and bla TEM genes. Overall 20.1%, 77/384, (95% CI; 43.2-65.5%) of total samples analyzed contained ESBL-producing Escherichia coli. The antimicrobial sensitivity test revealed that 85.7% (66/77; CI: 75.7-92) of ESBL-producing E. coli isolates conferred resistance to beta-lactam and other antimicrobial agents. These results indicate that poultry is a potential reservoir for ESBL-producing Escherichia coli. The presence of ESBL-producing Escherichia coli in poultry destined for human consumption requires strengthening of the antibiotic administering policy. This is important as antibiotic administration in food animals is gaining momentum for improved animal productivity in developing countries such as Zambia. PMID:27190518

  15. Detection of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Market-Ready Chickens in Zambia

    PubMed Central

    Chishimba, K.; Hang'ombe, B. M.; Muzandu, K.; Mshana, S. E.; Matee, M. I.; Nakajima, C.; Suzuki, Y.

    2016-01-01

    The frequent administering of antibiotics in the treatment of poultry diseases may contribute to emergence of antimicrobial-resistant strains. The objective of this study was to detect the presence of extended-spectrum β-lactamase- (ESBL-) producing Escherichia coli in poultry in Zambia. A total of 384 poultry samples were collected and analyzed for ESBL-producing Escherichia coli. The cultured E. coli isolates were subjected to antimicrobial susceptibility tests and the polymerase chain reaction for detection of blaCTX-M, blaSHV, and blaTEM genes. Overall 20.1%, 77/384, (95% CI; 43.2–65.5%) of total samples analyzed contained ESBL-producing Escherichia coli. The antimicrobial sensitivity test revealed that 85.7% (66/77; CI: 75.7–92) of ESBL-producing E. coli isolates conferred resistance to beta-lactam and other antimicrobial agents. These results indicate that poultry is a potential reservoir for ESBL-producing Escherichia coli. The presence of ESBL-producing Escherichia coli in poultry destined for human consumption requires strengthening of the antibiotic administering policy. This is important as antibiotic administration in food animals is gaining momentum for improved animal productivity in developing countries such as Zambia. PMID:27190518

  16. Prevalence of β-Lactamase Producing Escherichia coli from Retail Meat in Turkey.

    PubMed

    Pehlivanlar Önen, Sevda; Aslantaş, Özkan; Şebnem Yılmaz, Ebru; Kürekci, Cemil

    2015-09-01

    Extended spectrum β-lactamase (ESBL) and plasmid-mediated AmpC β-lactamase (pAmpC) producing Escherichia coli have been shown to be present in humans and animals representing a significant problem worldwide. This study aimed to search the presence of ESBL and/or AmpC-producing E. coli in retail meats (chicken and beef) in Turkey. A total of 88 β-lactamase-producing E. coli were isolated from chicken (n = 81/100) and beef meat (n = 7/100) samples and their susceptibility to several antimicrobials were tested using disc diffusion method. E. coli isolates were further characterized for their phylogenetic groups. β-Lactamase encoding (blaTEM , blaSHV , blaOXA , blaCTX-M , and blaAmpC ) and quinolone resistance genes (qnrA, qnrB, qnrS, qepA, and acc(6')-Ib-cr) were also secreened by polymerase chain reaction (PCR). However, in regard to β-lactamase genes, 84 of 88 isolates were positive for blaCTX-M-1 (n = 39), blaCTX-M-3 (n = 5), blaCTX-M-15 (n = 4), blaTEM-1b (n = 2), blaSHV-12 (n = 1), blaCTX-M-1 /blaTEM-1b (n = 10), blaCTX-M-1 /blaTEM-1b /blaSHV-5 (n = 1), blaCTX-M-1 /blaCMY-2 (n = 1) and blaTEM-1b /blaCMY-2 (n = 6), blaCTX-M-15 /blaSHV-12 (n = 1), blaCTX-M-15 /blaTEM-1b (n = 1), blaTEM-1b /blaSHV-12 (n = 1), and blaCMY-2 (n = 12) genes. Resistance to cefuroxime (75.6% and 85.7%), nalidixic acid (89% and 85.7%), tetracycline (91.4% and 100%), streptomycin (40.2% and 100%), and trimethoprim-sulfamethoxazole (36.6% and 85.7%) was observed among strains isolated from chicken and beef, respectively. However, all isolates were found to be susceptible to amikacin, imipenem, and cefepime. Resistance to ampicillin and cefoxitin was significantly linked to blaCMY-2 gene, while there was a significant correlation between CTX-M type ESBL and antimicrobial resistance to cefuroxime and streptomycin (P < 0.05). The results of this study suggest that raw chicken retail meats are highly contaminated with ESBL-producing E. coli implementing a great risk to human health in

  17. Clonal diversity of Shiga toxin-producing Escherichia coli O103:H2/H(-) in Germany.

    PubMed

    Prager, Rita; Liesegang, Almut; Voigt, W; Rabsch, W; Fruth, Angelika; Tschäpe, H

    2002-07-01

    Shiga toxin producing Escherichia coli O103:H2/H(-) belong to the third most frequently isolated EHEC serotypes in Germany following isolates of O157:H7/H(-) and O26:H11/H(-). A total of 145 respective E. coli 103 isolates from single cases of diarrhoea and haemolytic uremic syndrome (HUS) in 1997-2000 were characterised by a range of molecular subtyping methods (PFGE, P-gene profiling, ribotyping, electrotyping) and phage typing in order to analyse their genetic relatedness and the practicability for new epidemiological tracing back. All isolates cluster into a distinct EHEC subgroup and reveal a high clonal diversity together with a considerable stability. Since strains of this serotype rank up to the third most frequently isolated EHEC in Germany a large population of this serotype, and therefore, a great supply of such strains may exist in this country. PMID:12798005

  18. Non-O157 verotoxin-producing Escherichia coli: a problem, paradox, and paradigm.

    PubMed

    Bettelheim, Karl A

    2003-04-01

    The problems associated with identification and characterization of non-O157 verotoxin-producing Escherichia coli (VTEC) are discussed. The paradox of VTEC is that most reports of human illnesses are associated with serotypes such as O157:H7, O111:H- (nonmotile), O26:H11, and O113:H21, which are rarely found in domestic animals. However, those VTEC serotypes commonly found in domestic animals, especially ruminants, rarely cause human illnesses. When they cause human illnesses, the symptoms are similar to those caused by the serotypes E. coli O157:H7, O111:H-, O26:H11, and O113:H21. The impact of VTEC on human and animal health is also addressed. The VTEC and their toxicity are considered as a paradigm for emerging pathogens. The question on how such pathogens could arise from a basic commensal population is also addressed.

  19. Shiga Toxin/Verocytotoxin-Producing Escherichia coli Infections: Practical Clinical Perspectives.

    PubMed

    Davis, T Keefe; Van De Kar, Nicole C A J; Tarr, Phillip I

    2014-08-01

    Escherichia coli strains that produce Shiga toxins/verotoxins are rare, but important, causes of human disease. They are responsible for a spectrum of illnesses that range from the asymptomatic to the life-threatening hemolytic-uremic syndrome; diseases caused by E. coli belonging to serotype O157:H7 are exceptionally severe. Each illness has a fairly predictable trajectory, and good clinical practice at one phase can be inappropriate at other phases. Early recognition, rapid and definitive microbiology, and strategic selection of tests increase the likelihood of good outcomes. The best management of these infections consists of avoiding antibiotics, antimotility agents, and narcotics and implementing aggressive intravenous volume expansion, especially in the early phases of illness.

  20. Promising Nucleic Acid Lateral Flow Assay Plus PCR for Shiga Toxin-Producing Escherichia coli.

    PubMed

    Terao, Yoshitaka; Takeshita, Kana; Nishiyama, Yasutaka; Morishita, Naoki; Matsumoto, Takashi; Morimatsu, Fumiki

    2015-08-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) is a frequent cause of foodborne infections, and methods for rapid and reliable detection of STEC are needed. A nucleic acid lateral flow assay (NALFA) plus PCR was evaluated for detecting STEC after enrichment. When cell suspensions of 45 STEC strains, 14 non-STEC strains, and 13 non-E. coli strains were tested with the NALFA plus PCR, all of the STEC strains yielded positive results, and all of the non-STEC and non-E. coli strains yielded negative results. The lower detection limit for the STEC strains ranged from 0.1 to 1 pg of genomic DNA (about 20 to 200 CFU) per test, and the NALFA plus PCR was able to detect Stx1- and Stx2-producing E. coli strains with similar sensitivities. The ability of the NALFA plus PCR to detect STEC in enrichment cultures of radish sprouts, tomato, raw ground beef, and beef liver inoculated with 10-fold serially diluted STEC cultures was comparable to that of a real-time PCR assay (at a level of 100 to 100,000 CFU/ml in enrichment culture). The bacterial inoculation test in raw ground beef revealed that the lower detection limit of the NALFA plus PCR was also comparable to that obtained with a real-time PCR assay that followed the U.S. Department of Agriculture guidelines. Although further evaluation is required, these results suggest that the NALFA plus PCR is a specific and sensitive method for detecting STEC in a food manufacturing plant. PMID:26219371

  1. Fusion tags for protein solubility, purification and immunogenicity in Escherichia coli: the novel Fh8 system

    PubMed Central

    Costa, Sofia; Almeida, André; Castro, António; Domingues, Lucília

    2014-01-01

    Proteins are now widely produced in diverse microbial cell factories. The Escherichia coli is still the dominant host for recombinant protein production but, as a bacterial cell, it also has its issues: the aggregation of foreign proteins into insoluble inclusion bodies is perhaps the main limiting factor of the E. coli expression system. Conversely, E. coli benefits of cost, ease of use and scale make it essential to design new approaches directed for improved recombinant protein production in this host cell. With the aid of genetic and protein engineering novel tailored-made strategies can be designed to suit user or process requirements. Gene fusion technology has been widely used for the improvement of soluble protein production and/or purification in E. coli, and for increasing peptide’s immunogenicity as well. New fusion partners are constantly emerging and complementing the traditional solutions, as for instance, the Fh8 fusion tag that has been recently studied and ranked among the best solubility enhancer partners. In this review, we provide an overview of current strategies to improve recombinant protein production in E. coli, including the key factors for successful protein production, highlighting soluble protein production, and a comprehensive summary of the latest available and traditionally used gene fusion technologies. A special emphasis is given to the recently discovered Fh8 fusion system that can be used for soluble protein production, purification, and immunogenicity in E. coli. The number of existing fusion tags will probably increase in the next few years, and efforts should be taken to better understand how fusion tags act in E. coli. This knowledge will undoubtedly drive the development of new tailored-made tools for protein production in this bacterial system. PMID:24600443

  2. Molecular characterization and phylogeny of Shiga toxin-producing E. coli (STEC) from imported beef meat in Malaysia.

    PubMed

    Abuelhassan, Nawal Nouridaim; Mutalib, Sahilah Abdul; Gimba, Fufa Ido; Yusoff, Wan Mohtar

    2016-09-01

    This study aimed at determining the presence and characterization of Escherichia coli and Shiga toxin-producing E. coli (STEC) from imported frozen beef meats. Seventy-four (74) frozen imported beef meat samples from two countries, India (42 samples) and Australia (32 samples), were collected and tested for E. coli. These samples were purchased from the frozen meat sections of five different supermarkets in different locations in Selangor, Malaysia, from April 2012 to October 2014. A total of 222 E. coli strains were isolated from the meat samples; 126 strains were isolated from country A (India), and 96 E. coli strains were from country of origin B (Australia), respectively. A total of 70 E. coli strains were identified and characterized. All E. coli strains were isolated into Fluorocult medium and identified using API 20E kit. All selected E. coli strains were characterized for Shiga toxin genes (stx1 and stx2). All biochemically identified E. coli in this study were further subjected to molecular detection through polymerase chain reaction (PCR) amplification and characterization using 16S ribosomal RNA (rRNA) gene of Shiga toxin-producing E. coli. Of the 70 E. coli strains, 11 strains were positive for both Shiga toxin genes (stx1 and stx2) and 11 (11/70) strains were positive for stx1 gene, while 25 (25/70) strains were positive for stx2 gene. The analysis of 16S rRNA gene of all the E. coli isolates in this study was successfully sequenced and analyzed, and based on sequence data obtained, a phylogenetic tree of the 16S rRNA gene was performed using Clustal W programme in MEGA 6.06 software. Phylogenetic tree showed that the E. coli isolates in our study cluster with the strain of E. coli isolated in other countries, which further confirm that the isolates of E. coli in this study are similar to those obtained in other studies. As a result, all the strains obtained in this study proved to be a strain of pathogenic E. coli, which may cause a serious outbreak

  3. Molecular characterization and phylogeny of Shiga toxin-producing E. coli (STEC) from imported beef meat in Malaysia.

    PubMed

    Abuelhassan, Nawal Nouridaim; Mutalib, Sahilah Abdul; Gimba, Fufa Ido; Yusoff, Wan Mohtar

    2016-09-01

    This study aimed at determining the presence and characterization of Escherichia coli and Shiga toxin-producing E. coli (STEC) from imported frozen beef meats. Seventy-four (74) frozen imported beef meat samples from two countries, India (42 samples) and Australia (32 samples), were collected and tested for E. coli. These samples were purchased from the frozen meat sections of five different supermarkets in different locations in Selangor, Malaysia, from April 2012 to October 2014. A total of 222 E. coli strains were isolated from the meat samples; 126 strains were isolated from country A (India), and 96 E. coli strains were from country of origin B (Australia), respectively. A total of 70 E. coli strains were identified and characterized. All E. coli strains were isolated into Fluorocult medium and identified using API 20E kit. All selected E. coli strains were characterized for Shiga toxin genes (stx1 and stx2). All biochemically identified E. coli in this study were further subjected to molecular detection through polymerase chain reaction (PCR) amplification and characterization using 16S ribosomal RNA (rRNA) gene of Shiga toxin-producing E. coli. Of the 70 E. coli strains, 11 strains were positive for both Shiga toxin genes (stx1 and stx2) and 11 (11/70) strains were positive for stx1 gene, while 25 (25/70) strains were positive for stx2 gene. The analysis of 16S rRNA gene of all the E. coli isolates in this study was successfully sequenced and analyzed, and based on sequence data obtained, a phylogenetic tree of the 16S rRNA gene was performed using Clustal W programme in MEGA 6.06 software. Phylogenetic tree showed that the E. coli isolates in our study cluster with the strain of E. coli isolated in other countries, which further confirm that the isolates of E. coli in this study are similar to those obtained in other studies. As a result, all the strains obtained in this study proved to be a strain of pathogenic E. coli, which may cause a serious outbreak

  4. Antibacterial activities of essential oils from Iranian medicinal plants on extended-spectrum β-lactamase-producing Escherichia coli.

    PubMed

    Sharifi-Rad, J; Mnayer, D; Roointan, A; Shahri, F; Ayatollahi, S A M; Sharifi-Rad, M; Molaee, N; Sharifi-Rad, M

    2016-01-01

    The extended-spectrum beta-lactamase (ESBL) -producing Escherichia coli strains can lead to various infections particularly urinary tract infections. The main objective of this investigation was to evaluate the antibacterial activities of essential oils (EOs) from different Iranian medicinal plants against TEM gene positive ESBL-producing E. coli strains isolated from urine samples of patients with urinary tract infections. EOs were extracted using hydrodistillation method. E. coli strains were isolated by different specific Medias. ESBL-producing E. coli strains were isolated from urine samples of patients with urinary tract infections in Shiraz hospital, Iran. Then, ESBL- producing strains were identified using double disk synergy test, phenotypic disc confirmatory test and polymerase chain reaction (PCR) for TEM gene detection. The antibacterial activity of the EOs from different plants (Achillea wilhelmsii C. Koch, Echinophora platyloba DC., Lallemantia royleana, Nepeta persica Boiss., Pulicaria vulgaris Gaertn., Salvia nemorosa, and Satureja intermedia C.A.Mey) and antibiotics against ESBL-producing strains was studied using the microdilution method for the evaluation of the minimum inhibitory concentration (MIC). The 103 out of 295 E. coli strains with 97 (90.65%) TEM gene distributions were identified as ESBL-producing strains. All of the EOs derived from different plants displayed high inhibitory effects against ESBL-producing E. coli strains. The results of our investigations may propose a good treatment option against resistant infectious bacteria. PMID:27650980

  5. Metabolic design of a platform Escherichia coli strain producing various chorismate derivatives.

    PubMed

    Noda, Shuhei; Shirai, Tomokazu; Oyama, Sachiko; Kondo, Akihiko

    2016-01-01

    A synthetic metabolic pathway suitable for the production of chorismate derivatives was designed in Escherichia coli. An L-phenylalanine-overproducing E. coli strain was engineered to enhance the availability of phosphoenolpyruvate (PEP), which is a key precursor in the biosynthesis of aromatic compounds in microbes. Two major reactions converting PEP to pyruvate were inactivated. Using this modified E.coli as a base strain, we tested our system by carrying out the production of salicylate, a high-demand aromatic chemical. The titer of salicylate reached 11.5 g/L in batch culture after 48 h cultivation in a 2-liter jar fermentor, and the yield from glucose as the sole carbon source exceeded 40% (mol/mol). In this test case, we found that pyruvate was synthesized primarily via salicylate formation and the reaction converting oxaloacetate to pyruvate. In order to demonstrate the generality of our designed strain, we employed this platform for the production of each of 7 different chorismate derivatives. Each of these industrially important chemicals was successfully produced to levels of 1-3g/L in test tube-scale culture.

  6. Occurrence of Shiga toxin-producing Escherichia coli in a Spanish raw ewe's milk cheese.

    PubMed

    Caro, Irma; García-Armesto, María R

    2007-05-30

    The aim of the present study was to investigate the occurrence of Escherichia coli O157:H7 and other Shiga toxin-producing E. coli (STEC) in 'Castellano' cheese, a non-cooked and hard or semi-hard Spanish cheese made from ewe's milk. A total of 83 raw milk cheese samples with different ripening times (2.5, 6 and 12 months) were taken at 30 cheese factories. Samples were examined for the presence of STEC using in the first stage the Association of Official Analytical Chemists (AOAC) official method number 997.11, and then, in the second stage, isolates were tested for virulence genes using genotypic (PCR) methods. Three STEC strains were detected in two samples (2.4%) of 'Castellano' cheese, one with 2.5 and the other one with 12 month-ripening period. From those STEC isolates, two were identified as E. coli O14 and the third presented an O-specific polysaccharide not-groupable serologically (ONG). PCR showed that all isolates were characterized by harbouring the Shiga toxin (stx) stx1 gene and by the absence of the genes for stx2, eaeA, and ehxA virulence factors. This study revealed the potential of STEC to survive in long-ripened-hard cheeses.

  7. Epidemiological factors associated with ESBL- and non ESBL-producing E. coli causing urinary tract infection in general practice.

    PubMed

    Hertz, Frederik Boëtius; Schønning, Kristian; Rasmussen, Steen Christian; Littauer, Pia; Knudsen, Jenny Dahl; Løbner-Olesen, Anders; Frimodt-Møller, Niels

    2016-01-01

    The purpose of the study was to evaluate how use of antibiotics precedes the presence of ESBL-producing E.coli in general practice. The authors performed a triple-case-control study where three case groups were individually compared to a single control group of uninfected individuals. Urine samples were prospectively collected and retrospective statistical analyses were done. This study included 98 cases with urinary tract infection (UTI) caused by ESBL-producing E. coli, 174 with antibiotic-resistant (non-ESBL) E. coli, 177 with susceptible E. coli and 200 with culture negative urine samples. Case groups had significantly higher use of antibiotics than the control group within 30 days before infection (p < 0.0001). The ESBL group had significantly more hospital admissions than the other case groups (p < 0.05). Hospital admission was an independent risk factor for community onset UTI by ESBL-producing E. coli. Exposure to antibiotics was a risk factor for UTI with E. coli, while prior antibiotic usage was not an indisputable predictor for infection with ESBL-producing E.coli in general practice. PMID:26523346

  8. Epidemiological factors associated with ESBL- and non ESBL-producing E. coli causing urinary tract infection in general practice.

    PubMed

    Hertz, Frederik Boëtius; Schønning, Kristian; Rasmussen, Steen Christian; Littauer, Pia; Knudsen, Jenny Dahl; Løbner-Olesen, Anders; Frimodt-Møller, Niels

    2016-01-01

    The purpose of the study was to evaluate how use of antibiotics precedes the presence of ESBL-producing E.coli in general practice. The authors performed a triple-case-control study where three case groups were individually compared to a single control group of uninfected individuals. Urine samples were prospectively collected and retrospective statistical analyses were done. This study included 98 cases with urinary tract infection (UTI) caused by ESBL-producing E. coli, 174 with antibiotic-resistant (non-ESBL) E. coli, 177 with susceptible E. coli and 200 with culture negative urine samples. Case groups had significantly higher use of antibiotics than the control group within 30 days before infection (p < 0.0001). The ESBL group had significantly more hospital admissions than the other case groups (p < 0.05). Hospital admission was an independent risk factor for community onset UTI by ESBL-producing E. coli. Exposure to antibiotics was a risk factor for UTI with E. coli, while prior antibiotic usage was not an indisputable predictor for infection with ESBL-producing E.coli in general practice.

  9. Behavior of shiga toxin-producing Escherichia coli, enteroinvasive E. coli, enteropathogenic E. coli and enterotoxigenic E. coli strains on whole and sliced jalapeño and serrano peppers.

    PubMed

    Gómez-Aldapa, Carlos A; Rangel-Vargas, Esmeralda; Gordillo-Martínez, Alberto J; Castro-Rosas, Javier

    2014-06-01

    The behavior of enterotoxigenic Escherichia coli (ETEC), enteropathogenic E. coli (EPEC), enteroinvasive E. coli (EIEC) and non-O157 shiga toxin-producing E. coli (non-O157-STEC) on whole and slices of jalapeño and serrano peppers as well as in blended sauce at 25 ± 2 °C and 3 ± 2 °C was investigated. Chili peppers were collected from markets of Pachuca city, Hidalgo, Mexico. On whole serrano and jalapeño stored at 25 ± 2 °C or 3 ± 2 °C, no growth was observed for EPEC, ETEC, EIEC and non-O157-STEC rifampicin resistant strains. After twelve days at 25 ± 2 °C, on serrano peppers all diarrheagenic E. coli pathotypes (DEP) strains had decreased by a total of approximately 3.7 log, whereas on jalapeño peppers the strains had decreased by approximately 2.8 log, and at 3 ± 2 °C they decreased to approximately 2.5 and 2.2 log respectively, on serrano and jalapeño. All E. coli pathotypes grew onto sliced chili peppers and in blended sauce: after 24 h at 25 ± 2 °C, all pathotypes had grown to approximately 3 and 4 log CFU on pepper slices and sauce, respectively. At 3 ± 2 °C the bacterial growth was inhibited. PMID:24549200

  10. Behavior of shiga toxin-producing Escherichia coli, enteroinvasive E. coli, enteropathogenic E. coli and enterotoxigenic E. coli strains on whole and sliced jalapeño and serrano peppers.

    PubMed

    Gómez-Aldapa, Carlos A; Rangel-Vargas, Esmeralda; Gordillo-Martínez, Alberto J; Castro-Rosas, Javier

    2014-06-01

    The behavior of enterotoxigenic Escherichia coli (ETEC), enteropathogenic E. coli (EPEC), enteroinvasive E. coli (EIEC) and non-O157 shiga toxin-producing E. coli (non-O157-STEC) on whole and slices of jalapeño and serrano peppers as well as in blended sauce at 25 ± 2 °C and 3 ± 2 °C was investigated. Chili peppers were collected from markets of Pachuca city, Hidalgo, Mexico. On whole serrano and jalapeño stored at 25 ± 2 °C or 3 ± 2 °C, no growth was observed for EPEC, ETEC, EIEC and non-O157-STEC rifampicin resistant strains. After twelve days at 25 ± 2 °C, on serrano peppers all diarrheagenic E. coli pathotypes (DEP) strains had decreased by a total of approximately 3.7 log, whereas on jalapeño peppers the strains had decreased by approximately 2.8 log, and at 3 ± 2 °C they decreased to approximately 2.5 and 2.2 log respectively, on serrano and jalapeño. All E. coli pathotypes grew onto sliced chili peppers and in blended sauce: after 24 h at 25 ± 2 °C, all pathotypes had grown to approximately 3 and 4 log CFU on pepper slices and sauce, respectively. At 3 ± 2 °C the bacterial growth was inhibited.

  11. Carriage of Escherichia coli Producing CTX-M-Type Extended-Spectrum β-Lactamase in Healthy Vietnamese Individuals

    PubMed Central

    Ueda, Shuhei; Bui, Thi Kim Ngan; Hamamoto, Kouta; Toyosato, Takehiko; Le, Danh Tuyen; Yamamoto, Yoshimasa

    2015-01-01

    Healthy carriage of CTX-M-type extended-spectrum β-lactamase (ESBL)-producing Escherichia coli was examined by thrice collecting fecal samples from the same 199 healthy Vietnamese subjects every 6 months. Using pulsed-field gel electrophoresis (PFGE), identical PFGE patterns throughout the three samplings were not observed, although prevalence of E. coli in the subjects was around 50% in the three samplings. Our results suggested a short carriage period of the CTX-M-type ESBL-producing E. coli in healthy Vietnamese subjects. PMID:26195526

  12. Carriage of Escherichia coli Producing CTX-M-Type Extended-Spectrum β-Lactamase in Healthy Vietnamese Individuals.

    PubMed

    Bui, Thi Mai Huong; Hirai, Itaru; Ueda, Shuhei; Bui, Thi Kim Ngan; Hamamoto, Kouta; Toyosato, Takehiko; Le, Danh Tuyen; Yamamoto, Yoshimasa

    2015-10-01

    Healthy carriage of CTX-M-type extended-spectrum β-lactamase (ESBL)-producing Escherichia coli was examined by thrice collecting fecal samples from the same 199 healthy Vietnamese subjects every 6 months. Using pulsed-field gel electrophoresis (PFGE), identical PFGE patterns throughout the three samplings were not observed, although prevalence of E. coli in the subjects was around 50% in the three samplings. Our results suggested a short carriage period of the CTX-M-type ESBL-producing E. coli in healthy Vietnamese subjects. PMID:26195526

  13. Diagnosis of Shiga toxin producing Escherichia coli infection, contribution of genetic amplification technique.

    PubMed

    El Sayed Zaki, Maysaa; El-Adrosy, Hala

    2007-02-01

    There has been no culture method of choice for detecting non-O157 Shiga toxin-producing Escherichia coli strains (STEC) because of their biochemical diversity The aim of this study was the assessment of verotoxin gene detection (VT1/VT2) within STEC PCR compared with the Vero cells cytotoxicity among O157 and non-O157 STEC serotypes. Stool cultures were performed on Tryptic Soy Broth and sorbitol MacConkey agar with cefixitime and tellurite supplements which were identified as Escherichia coli (E. coli) by BBL crystal. Further identifications were performed including verotoxin production assessment by Vero cells cytotoxicity assay, PCR for specific VT1/VT2 genotyping, and isolates were plated on blood agar and tested for enterohemolysis. Vero cells cytotoxicity assay revealed that 58 of E. coli isolates (71.6%) were STEC. In PCR, 33 (56.9%) of the 58 strains were positive for the VT2 gene, 24 (41.4%) were positive for the VT1 gene and one isolate was positive for both genes. In comparison to Vero cells cytotoxicity, the sensitivity, specificity of PCR were 100%. In comparative study between verotoxin assessment by Vero cells cytotoxicity and enterohemolytic activity, concordance positive results between both were 53 (91.4%). The most common serogroups of STEC were O157 (33%) and O26 (20%). From this study we can conclude that enterohemolysin production can be used as surrogate marker for STEC. The most rapid and promising approach for detection of STEC is by molecular method.

  14. Six Novel O Genotypes from Shiga Toxin-Producing Escherichia coli.

    PubMed

    Iguchi, Atsushi; Iyoda, Sunao; Seto, Kazuko; Nishii, Hironobu; Ohnishi, Makoto; Mekata, Hirohisa; Ogura, Yoshitoshi; Hayashi, Tetsuya

    2016-01-01

    Serotyping is one of the typing techniques used to classify strains within the same species. O-serogroup diversification shows a strong association with the genetic diversity of O-antigen biosynthesis genes. In a previous study, based on the O-antigen biosynthesis gene cluster (O-AGC) sequences of 184 known Escherichia coli O serogroups (from O1 to O187), we developed a comprehensive and practical molecular O serogrouping (O genotyping) platform using a polymerase chain reaction (PCR) method, named E. coli O-genotyping PCR. Although, the validation assay using the PCR system showed that most of the tested strains were successfully classified into one of the O genotypes, it was impossible to classify 6.1% (35/575) of the strains, suggesting the presence of novel O genotypes. In this study, we conducted sequence analysis of O-AGCs from O-genotype untypeable Shiga toxin-producing E. coli (STEC) strains and identified six novel O genotypes; OgN1, OgN8, OgN9, OgN10, OgN12 and OgN31, with unique wzx and/or wzy O-antigen processing gene sequences. Additionally, to identify these novel O-genotypes, we designed specific PCR primers. A screen of O genotypes using O-genotype untypeable strains showed 13 STEC strains were classified into five novel O genotypes. The O genotyping at the molecular level of the O-AGC would aid in the characterization of E. coli isolates and will assist future studies in STEC epidemiology and phylogeny.

  15. Six Novel O Genotypes from Shiga Toxin-Producing Escherichia coli

    PubMed Central

    Iguchi, Atsushi; Iyoda, Sunao; Seto, Kazuko; Nishii, Hironobu; Ohnishi, Makoto; Mekata, Hirohisa; Ogura, Yoshitoshi; Hayashi, Tetsuya

    2016-01-01

    Serotyping is one of the typing techniques used to classify strains within the same species. O-serogroup diversification shows a strong association with the genetic diversity of O-antigen biosynthesis genes. In a previous study, based on the O-antigen biosynthesis gene cluster (O-AGC) sequences of 184 known Escherichia coli O serogroups (from O1 to O187), we developed a comprehensive and practical molecular O serogrouping (O genotyping) platform using a polymerase chain reaction (PCR) method, named E. coli O-genotyping PCR. Although, the validation assay using the PCR system showed that most of the tested strains were successfully classified into one of the O genotypes, it was impossible to classify 6.1% (35/575) of the strains, suggesting the presence of novel O genotypes. In this study, we conducted sequence analysis of O-AGCs from O-genotype untypeable Shiga toxin-producing E. coli (STEC) strains and identified six novel O genotypes; OgN1, OgN8, OgN9, OgN10, OgN12 and OgN31, with unique wzx and/or wzy O-antigen processing gene sequences. Additionally, to identify these novel O-genotypes, we designed specific PCR primers. A screen of O genotypes using O-genotype untypeable strains showed 13 STEC strains were classified into five novel O genotypes. The O genotyping at the molecular level of the O-AGC would aid in the characterization of E. coli isolates and will assist future studies in STEC epidemiology and phylogeny. PMID:27242776

  16. Inactivation of Escherichia coli ATCC 11775 in fresh produce using atmospheric pressure cold plasma

    NASA Astrophysics Data System (ADS)

    Bermudez-Aguirre, Daniela; Wemlinger, Erik; Barbosa-Canovas, Gustavo; Pedrow, Patrick; Garcia-Perez, Manuel

    2011-10-01

    Food-borne outbreaks are associated with the presence of pathogenic bacteria in food products such as fresh produce. One of the target microorganisms is Escherichia coli which exhibits resistance to being inactivated with conventional disinfection methods for vegetables. Atmospheric pressure cold plasma (APCP) was tested to disinfect three vegetables with challenge surfaces, lettuce, carrots and tomatoes. The produce was inoculated with the bacteria to reach an initial microbial concentration of 107 cfu/g. Vegetables were initially exposed to the APCP discharges from a needle array at 5.7 kV RMS in argon, processing times of 0.5, 3 and 5 min. Initial results indicate that microbial decontamination is effective on the lettuce (1.2 log reduction) when compared with other vegetables. To claim disinfection, a 3 log reduction or more is needed, which makes APCP treatment very promising technology for decontamination of produce. We propose that with method refinements full disinfection can be achieved using APCP.

  17. Caffeic acid production enhancement by engineering a phenylalanine over-producing Escherichia coli strain.

    PubMed

    Huang, Qin; Lin, Yuheng; Yan, Yajun

    2013-12-01

    Caffeic acid is a plant-specific phenylpropanoic acid with multiple health-improving effects reported, and its therapeutic derivatives have also been studied throughout the last decade. To meet its market need and achieve high-level production, microbial production of caffeic acid approaches have been developed in metabolically engineered Escherichia coli. In our previous work, we have established the first artificial pathway that realized de novo production of caffeic acid using E. coli endogenous 4-hydroxyphenylacetate 3-hydroxylase (4HP3H). In this work, we exploited the catalytic potential of 4HPA3H in the whole-cell bioconversion study and produced 3.82 g/L (461.12 mg/L/OD) caffeic acid from p-coumaric acid, a direct precursor. We further engineered a phenylalanine over-producer into a tyrosine over-producer and then introduced the artificial pathway. After adjusting the expression strategy and optimizing the inoculants timing, de novo production of caffeic acid reached 766.68 mg/L. Both results from the direct precursor and simple carbon sources represent the highest titers of caffeic acid from microbial production so far.

  18. The microcosm mediates the persistence of shiga toxin-producing Escherichia coli in freshwater ecosystems.

    PubMed

    Mauro, Steven A; Opalko, Hannah; Lindsay, Kyle; Colon, Michael P; Koudelka, Gerald B

    2013-08-01

    Water is a major route for infection of humans by exotoxin-producing bacteria, including Shiga toxin-producing Escherichia coli (STEC). While STEC has the potential to be present in nearly every type of water source, its distribution is sporadic, and an understanding of factors that govern its emergence and persistence within water is lacking. In this study, we examined the influence of microbe content on STEC persistence in freshwater. We found that depletion of microbes in the water leads to a considerable increase in the persistence of STEC, an effect that can be mitigated by adding grazing protists to the water. STEC strains appear to be more resistant to the impact of grazing protists than E. coli strains that lack the Shiga toxin (stx) gene. Our results demonstrate that the microcosm can dramatically influence the persistence of STEC in aquatic ecosystems and that the overall impact by microbes on STEC strains is fundamentally different from that of non-STEC strains of bacteria. Overall, these results provide insight into why STEC and possibly other exotoxin-producing bacterial pathogens display such variability in abundance, distribution, and persistence in aquatic ecosystems.

  19. Distribution, Numbers, and Diversity of ESBL-Producing E. coli in the Poultry Farm Environment

    PubMed Central

    Blaak, Hetty; van Hoek, Angela H. A. M.; Hamidjaja, Raditijo A.; van der Plaats, Rozemarijn Q. J.; Kerkhof-de Heer, Lianne; de Roda Husman, Ana Maria; Schets, Franciska M.

    2015-01-01

    This study aimed to discern the contribution of poultry farms to the contamination of the environment with ESBL-producing Escherichia coli and therewith, potentially to the spread of these bacteria to humans and other animals. ESBL-producing E. coli were detected at all investigated laying hen farms (n = 5) and broiler farms (n = 3) in 65% (46/71) and 81% (57/70) of poultry faeces samples, respectively. They were detected in rinse water and run-off water (21/26; 81%), other farm animals (11/14; 79%), dust (21/35; 60%), surface water adjacent to farms (20/35; 57%), soil (48/87; 55%), on flies (11/73; 15%), and in barn air (2/33; 6%). The highest prevalence and concentrations in the outdoor environment were observed in soil of free-range areas at laying hen farms (100% of samples positive, geometric mean concentration 2.4×104 cfu/kg), and surface waters adjacent to broiler farms during, or shortly after, cleaning between production rounds (91% of samples positive, geometric mean concentration 1.9×102 cfu/l). The diversity of ESBL-producing E. coli variants with respect to sequence type, phylogenetic group, ESBL-genotype and antibiotic resistance profile was high, especially on broiler farms where on average 16 different variants were detected, and the average Simpson’s Indices of diversity (SID; 1–D) were 0.93 and 0.94 among flock and environmental isolates respectively. At laying hen farms on average nine variants were detected, with SIDs of 0.63 (flock isolates) and 0.77 (environmental isolates). Sixty percent of environmental isolates were identical to flock isolates at the same farm. The highest proportions of ‘flock variants’ were observed in dust (94%), run-off gullies (82%), and barn air (67%), followed by surface water (57%), soil (56%), flies (50%) and other farm animals (35%).The introduction of ESBL-producing E. coli from poultry farms to the environment may pose a health risk if these bacteria reach places where people may become exposed. PMID

  20. Roasting coffee beans produces compounds that induce prophage lambda in E. coli and are mutagenic in E. coli and S. typhimurium.

    PubMed

    Kosugi, A; Nagao, M; Suwa, Y; Wakabayashi, K; Sugimura, T

    1983-03-01

    Freshly brewed blended coffee, instant coffee and instant caffeine-free coffee induced prophage lambda in lysogenic E. coli K12, strain GY5027. Because coffee prepared from green beans by the same extraction method as used for freshly brewed blended coffee had no prophage-inducing activity, this activity may be attributed to compounds produced in the roasting process. Roasting also produced compounds that were mutagenic in S. typhimurium TA100 and E. coli WP2 uvrA/pKM101. PMID:6220221

  1. Methods for the detection and isolation of Shiga toxin-producing Escherichia coli.

    PubMed

    De Boer E; Heuvelink, A E

    2000-01-01

    Shiga toxin-producing Escherichia coli (STEC) are an important cause of haemorrhagic colitis and the diarrhoea-associated form of the haemolytic uraemic syndrome. Of the numerous serotypes of E. coli that have been shown to produce Shiga toxin (Stx), E. coli O157:H7 and E. coli O157:NM (non-motile) are most frequently implicated in human disease. Early recognition of STEC infections is critical for effective treatment of patients. Furthermore, rapid microbiological diagnosis of individual patients enables the prompt notification of outbreaks and implementation of control measures to prevent more cases. Most human infections caused by STEC have been acquired by the consumption of contaminated foods, especially those of bovine origin such as undercooked ground beef and unpasteurized cows' milk, and by person-to-person contacts. To identify the reservoirs of STEC and the routes of transmission to man, sensitive methods are needed as these pathogens may only be present in food, environmental and faecal samples in small numbers. In addition, sensitive and rapid detection methods are necessary for the food industry to ensure a safe supply of foods. Sensitive methods are also needed for surveillance programmes in risk assessment studies, and for studies on survival and growth of STEC strains. Cultural methods for the enrichment, isolation and confirmation of O157 STEC are still evolving. Several selective enrichment media have been described, of which modified tryptone soy broth with novobiocin and modified E. coli broth with novobiocin, seem to be the most appropriate. These media are minimally-selective broths that give a somewhat limited differential specificity favouring isolation of O157 STEC, as opposed to other Gram-negative bacteria, in the sample. An incubation temperature of 41-42 degrees C further enhances selectivity. The occurrence of heat-, freeze-, acid- or salt-stressed STEC in foods means that it is important to be able to detect cells that are in a

  2. Construction and Characterization of an Escherichia coli Mutant Producing Kdo2-Lipid A

    PubMed Central

    Wang, Jianli; Ma, Wenjian; Wang, Zhou; Li, Ye; Wang, Xiaoyuan

    2014-01-01

    3-deoxy-d-manno-oct-2-ulosonic acid (Kdo)2-lipid A is the conserved structure domain of lipopolysaccharide found in most Gram-negative bacteria, and it is believed to stimulate the innate immune system through the TLR4/MD2 complex. Therefore, Kdo2-lipid A is an important stimulator for studying the mechanism of the innate immune system and for developing bacterial vaccine adjuvants. Kdo2-lipid A has not been chemically synthesized to date and could only be isolated from an Escherichia coli mutant strain, WBB06. WBB06 cells grow slowly and have to grow in the presence of tetracycline. In this study, a novel E. coli mutant strain, WJW00, that could synthesize Kdo2-lipid A was constructed by deleting the rfaD gene from the genome of E. coli W3110. The rfaD gene encodes ADP-l-glycero-d-manno-heptose-6-epimerase RfaD. Based on the analysis by SDS-PAGE, thin layer chromatography (TLC) and electrospray ionization mass spectrometry (ESI/MS), WJW00 could produce similar levels of Kdo2-lipid A to WBB06. WJW00 cells grow much better than WBB06 cells and do not need to add any antibiotics during growth. Compared with the wild-type strain, W3110, WJW00 showed increased hydrophobicity, higher cell permeability, greater autoaggregation and decreased biofilm-forming ability. Therefore, WJW00 could be a more suitable strain than WBB06 for producing Kdo2-lipid A and a good base strain for developing lipid A adjuvants. PMID:24633251

  3. Introduction to the food safety concerns of verotoxin-producing Escherichia coli.

    PubMed

    Hussein, Hussein S; Omaye, Stanley T

    2003-04-01

    Verotoxin-producing Escherichia coli (VTEC) have emerged in the past two decades as food-borne pathogens that can cause major outbreaks of human illnesses worldwide. The number of outbreaks has increased in recent years due to changes in food production and processing systems, eating habits, microbial adaptation, and methods of VTEC transmission. The human illnesses range from mild diarrhea to hemolytic uremic syndrome (HUS) that can lead to death. The VTEC outbreaks have been attributed to O157:H7 and non-O157:H7 serotypes of E. coli. These E. coli serotypes include motile (e.g., O26:H11 and O104:H21) and nonmotile (e.g., O111:H-, O145:H-, and O157:H-) strains. In the United States, E. coli O157:H7 has been the major cause of VTEC outbreaks. Worldwide, however, non-O157:H7 VTEC (e.g., members of the O26, O103, O111, O118, O145, and O166 serogroups) have caused approximately 30% of the HUS cases in the past decade. Because large numbers of the VTEC outbreaks have been attributed to consumption of ruminant products (e.g., ground beef), cattle and sheep are considered reservoirs of these food-borne pathogens. Because of the food safety concern of VTEC, a global perspective on this problem is addressed (Exp Biol Med Vol. 228, No. 4). The first objective was to evaluate the known non-O157:H7 VTEC strains and the limitations associated with their detection and characterization. The second objective was to identify the VTEC serotypes associated with outbreaks of human illnesses and to provide critical evaluation of their virulence. The third objective was to determine the rumen effect on survival of E. coli O157:H7 as a VTEC model. The fourth objective was to explore the role of intimins in promoting attaching and effacing lesions in humans. Finally, the ability of VTEC to cause persistent infections in cattle was evaluated.

  4. Validation of pepperoni process for control of Shiga toxin-producing Escherichia coli.

    PubMed

    Glass, Kathleen A; Kaspar, Charles W; Sindelar, Jeffrey J; Milkowski, Andrew L; Lotz, Brian M; Kang, Jihun; Faith, Nancy G; Enache, Elena; Kataoka, A I; Henry, Craig

    2012-05-01

    The objective of this study was to compare the survival of non-O157 Shiga toxin-producing Escherichia coli (STEC) with E. coli O157:H7 during pepperoni production. Pepperoni batter was inoculated with 7 log CFU/g of a seven-strain STEC mixture, including strains of serotypes O26, O45, O103, O111, O121, O145, and O157. Sausages were fermented to pH ≤4.8, heated at 53.3°C for 1 h, and dried for up to 20 days. STEC strains were enumerated at designated intervals on sorbitol MacConkey (SMAC) and Rainbow (RA) agars; enrichments were completed in modified EC (mEC) broth and nonselective tryptic soy broth (TSB). When plated on SMAC, total E. coli populations decreased 2.6 to 3.5 log after the 1-h heating step at 53.3°C, and a 4.9- to 5-log reduction was observed after 7 days of drying. RA was more sensitive in recovering survivors; log reductions on it were 1.9 to 2.6, 3.8 to 4.2, and 4.6 to 5.3 at the end of cook, and at day 7 and day 14 of drying, respectively. When numbers were less than the limit of detection by direct plating on days 14 and 20 of drying (representing a 5-log kill), no more than one of three samples in each experiment was positive by enrichment with mEC broth; however, STEC strains were recovered in TSB enrichment. Freezing the 7-day dried sausage for 2 to 3 weeks generated an additional 1- to 1.5-log kill. Confirmation by PCR revealed that O103 and O157 had the greatest survival during pepperoni productions, but all serotypes except O111 and O121 were occasionally recovered during drying. This study suggests that non-O157 STEC s trains have comparable or less ability than E. coli O157 to survive the processing steps involved in the manufacture of pepperoni. Processes suitable for control of E. coli O157 will similarly inactivate the other STEC strains tested in this study.

  5. Attaching and effacing Escherichia coli and Shiga toxin-producing E. coli in children with acute diarrhoea and controls in Teresina/PI, Brazil.

    PubMed

    Nunes, Maria do Rosário Conceição Moura; Magalhães, Paula Prazeres; Macêdo, Antônio da Silva; Franco, Roger Teixeira; Penna, Francisco José; Mendes, Edilberto Nogueira

    2012-01-01

    This 3.5-year prospective study was conducted to ascertain the level of attaching and effacing Escherichia coli (AEEC) associated diarrhoea in children from Teresina, a northeastern state of Brazil. Passed faecal specimens from 400 patients (250 with and 150 without diarrhoea) up to 60 months of age attending from 2004 to 2007 at two public hospitals were investigated. Conventional microbiology methods and PCR were employed. Escherichia coli was isolated from 390 children, 240 of them with diarrhoea. A total of 117 AEEC strains were cultivated from specimens from 63 children, 37 with and 26 without diarrhoea. No association between AEEC and diarrhoea was observed. Atypical enteropathogenic E. coli (a-EPEC) (79.4%) was more commonly found than typical EPEC (t-EPEC). Association between EPEC and EPEC subtypes and diarrhoea was not detected. Mixed infection by t-EPEC and a-EPEC and infection by Shiga toxin-producing E. coli (STEC) were rare. Enteropathogenic E. coli was more common in males and in children aged less than 12 months. Correlation between serotyping and PCR results was 0.19. High resistance rates of AEEC to ampicillin, cephalotin, and trimethoprim-sulfamethoxazole were found. In conclusion, EPEC is very common in children with diarrhoea and controls in the population we studied, with a-EPEC predominating. This diarrhoeagenic E. coli (DEC) pathotype is more common in infant males and is resistant to drugs frequently used in clinical practice.

  6. Shiga Toxin–Producing Escherichia coli O157, England and Wales, 1983–2012

    PubMed Central

    Byrne, Lisa; Smith, Geraldine A.; Elson, Richard; Harris, John P.; Salmon, Roland; Smith, Robert; O’Brien, Sarah J.; Adak, Goutam K.; Jenkins, Claire

    2016-01-01

    We evaluated clinical Shiga toxin–producing Escherichia coli O157 infections in England and Wales during 1983–2012 to describe changes in microbiological and surveillance methods. A strain replacement event was captured; phage type (PT) 2 decreased to account for just 3% of cases by 2012, whereas PT8 and PT21/28 strains concurrently emerged, constituting almost two thirds of cases by 2012. Despite interventions to control and reduce transmission, incidence remained constant. However, sources of infection changed over time; outbreaks caused by contaminated meat and milk declined, suggesting that interventions aimed at reducing meat cross-contamination were effective. Petting farm and school and nursery outbreaks increased, suggesting the emergence of other modes of transmission and potentially contributing to the sustained incidence over time. Studies assessing interventions and consideration of policies and guidance should be undertaken to reduce Shiga toxin–producing E. coli O157 infections in England and Wales in line with the latest epidemiologic findings. PMID:26982243

  7. Emergence of Escherichia coli Sequence Type 131 Isolates Producing KPC-2 Carbapenemase in China

    PubMed Central

    Cai, Jia Chang; Zhang, Rong; Hu, Yan Yan; Zhou, Hong Wei

    2014-01-01

    Twenty-two KPC-2-producing Escherichia coli isolates were obtained from three hospitals in Hangzhou, China, from 2007 to 2011. One isolate, with OmpC porin deficiency, exhibited high-level carbapenem resistance. Pulsed-field gel electrophoresis showed that few isolates were indistinguishable or closely related. Multilocus sequence typing indicated that sequence type 131 (ST131) was the predominant type (9 isolates, 40.9%), followed by ST648 (5 isolates), ST405 (2 isolates), ST38 (2 isolates), and 4 single STs, ST69, ST2003, ST2179, and ST744. Phylogenetic analysis indicated that 9 group B2 isolates belonged to ST131, and 5 of 11 group D isolates belonged to ST648. Only one group B1 isolate and one group A isolate were identified. A representative plasmid (pE1) was partially sequenced, and a 7,788-bp DNA fragment encoding Tn3 transposase, Tn3 resolvase, ISKpn8 transposase, KPC-2, and ISKpn6-like transposase was obtained. The blaKPC-2-surrounding sequence was amplified by a series of primers. The PCR results showed that 13 isolates were consistent with the genetic environment in pE1. It is the first report of rapid emergence of KPC-2-producing E. coli ST131 in China. The blaKPC-2 gene of most isolates was located on a similar genetic structure. PMID:24323475

  8. Shiga Toxin-Producing Escherichia coli O157, England and Wales, 1983-2012.

    PubMed

    Adams, Natalie L; Byrne, Lisa; Smith, Geraldine A; Elson, Richard; Harris, John P; Salmon, Roland; Smith, Robert; O'Brien, Sarah J; Adak, Goutam K; Jenkins, Claire

    2016-04-01

    We evaluated clinical Shiga toxin-producing Escherichia coli O157 infections in England and Wales during 1983-2012 to describe changes in microbiological and surveillance methods. A strain replacement event was captured; phage type (PT) 2 decreased to account for just 3% of cases by 2012, whereas PT8 and PT21/28 strains concurrently emerged, constituting almost two thirds of cases by 2012. Despite interventions to control and reduce transmission, incidence remained constant. However, sources of infection changed over time; outbreaks caused by contaminated meat and milk declined, suggesting that interventions aimed at reducing meat cross-contamination were effective. Petting farm and school and nursery outbreaks increased, suggesting the emergence of other modes of transmission and potentially contributing to the sustained incidence over time. Studies assessing interventions and consideration of policies and guidance should be undertaken to reduce Shiga toxin-producing E. coli O157 infections in England and Wales in line with the latest epidemiologic findings. PMID:26982243

  9. Emergence of Escherichia coli sequence type 131 isolates producing KPC-2 carbapenemase in China.

    PubMed

    Cai, Jia Chang; Zhang, Rong; Hu, Yan Yan; Zhou, Hong Wei; Chen, Gong-Xiang

    2014-01-01

    Twenty-two KPC-2-producing Escherichia coli isolates were obtained from three hospitals in Hangzhou, China, from 2007 to 2011. One isolate, with OmpC porin deficiency, exhibited high-level carbapenem resistance. Pulsed-field gel electrophoresis showed that few isolates were indistinguishable or closely related. Multilocus sequence typing indicated that sequence type 131 (ST131) was the predominant type (9 isolates, 40.9%), followed by ST648 (5 isolates), ST405 (2 isolates), ST38 (2 isolates), and 4 single STs, ST69, ST2003, ST2179, and ST744. Phylogenetic analysis indicated that 9 group B2 isolates belonged to ST131, and 5 of 11 group D isolates belonged to ST648. Only one group B1 isolate and one group A isolate were identified. A representative plasmid (pE1) was partially sequenced, and a 7,788-bp DNA fragment encoding Tn3 transposase, Tn3 resolvase, ISKpn8 transposase, KPC-2, and ISKpn6-like transposase was obtained. The blaKPC-2-surrounding sequence was amplified by a series of primers. The PCR results showed that 13 isolates were consistent with the genetic environment in pE1. It is the first report of rapid emergence of KPC-2-producing E. coli ST131 in China. The blaKPC-2 gene of most isolates was located on a similar genetic structure.

  10. Investigation of Encephalopathy Caused by Shiga Toxin 2c-Producing Escherichia coli Infection in Mice

    PubMed Central

    Amran, Muhammad Yunus; Fujii, Jun; Suzuki, Satoshi O.; Kolling, Glynis L.; Villanueva, Sharon Y. A. M.; Kainuma, Mosaburo; Kobayashi, Hideyuki; Kameyama, Hideko; Yoshida, Shin-ichi

    2013-01-01

    A large outbreak of Shiga toxin (Stx)-producing enteroaggregative Escherichia coli (EAEC) O104:H4 occurred in northern Germany. From this outbreak, at least 900 patients developed hemolytic uremic syndrome (HUS), resulting in more than 50 deaths. Thirty percent of the HUS patients showed encephalopathy. We previously established a mouse model with encephalopathy associated with blood brain barrier (BBB) damage after oral infection with the Shiga toxin (Stx) 2c-producing Escherichia coli O157: H- strain E32511 (E32511). In this model, we detected high expression of the Stx receptor synthase enzyme, glycosphingolipid globotriaosylceramide (Gb3) synthase, in endothelial cells (ECs) and neurons in the reticular formation of the medulla oblongata by in situ hybridization. Caspase-3 was activated in neurons in the reticular formation of the medulla oblongata and the anterior horn of the spinal cord. Astrocytes (ASTs) were activated in the medulla oblongata and spinal cord, and a decrease in aquaporin 4 around the ECs suggested that BBB integrity was compromised directly by Stx2c or through the activation of ASTs. We also report the effectiveness of azithromycin (AZM) in our model. Moreover, AZM strongly inhibited the release of Stx2c from E32511 in vitro. PMID:23516588

  11. CTX-M-15-type extended-spectrum beta-lactamases-producing Escherichia coli from wild birds in Germany.

    PubMed

    Guenther, Sebastian; Grobbel, Mirjam; Beutlich, Janine; Bethe, Astrid; Friedrich, Nicole D; Goedecke, Andreas; Lübke-Becker, Antina; Guerra, Beatriz; Wieler, Lothar H; Ewers, Christa

    2010-10-01

    The isolation of Escherichia coli from wild birds in Germany revealed the occurrence of four CTX-M-15-producing strains from four different birds (2.3% of 172 isolates). CTX-M producers were recovered from two Eurasian Blackbirds, one Rock Pigeon and a Greater White-fronted Goose. All CTX-M-producing E. coli revealed a clonal relationship as determined by pulsed-field gel electrophoresis (PFGE) and were assigned to multilocus sequence type (ST) 648. Our findings suggest the emergence of a new clone with epidemiological importance and strengthen the role of wild bird species other than waterfowl as possible reservoirs of ESBL-producing Enterobacteriaceae. PMID:23766249

  12. A new immunoassay for detecting all subtypes of Shiga toxins produced by Shiga toxin-producing E. coli in ground beef

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background Shiga toxin (Stx) is a common virulence factor of all Shiga toxin producing E. coli (STEC) that cause a wide spectrum of disease, including hemorrhagic colitis and hemolytic uremic syndrome (HUS). Although several commercial kits are available for detection of Stx produced by STEC, none o...

  13. Selective recovery by different culture methods of Shiga toxin-producing Escherichia coli genotypes from a major produce production region in California(Abstract)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The higher consumption of fresh fruits and vegetables in the United States has corresponded to an increase in the number of outbreaks. More than 45% of the leafy greens-associated outbreaks of Shiga toxin-producing Escherichia coli (STEC) O157:H7 have been linked to produce from California’s central...

  14. [The directed modification of Escherichia coli MG1655 to obtain histidine-producing mutants].

    PubMed

    Doroshenko, V G; Lobanov, A O; Fedorina, E A

    2013-01-01

    Strain MG 1655+hisGr hisL'-Delta, purR, which produces histidine with a weight yield of approximately 12% from glucose, was constructed through directed chromosomal modifications of the laboratory Escherichia coli strain MG 1655+, which has a known genome sequence. A feedback-resistant ATP-phosphoribosyl transferase encoded by the mutant hisGr (E271 K) was the main determinant of histidine production. A further increase in histidine production was achieved by the expression enhance of a mutant his operon containing hisGr through the deleting attenuator region (hisL'-Delta). An increase in the expression of the wildtype his operon did not result in histidine accumulation. Deletion of the transcriptional regulator gene purR increased the biomass produced and maintained the level of histidine production per cell under the fermentation conditions used.

  15. Properties of lactase produced by enteropathogenic Escherichia coli from diarrhoeic children.

    PubMed

    Olusanya, O; Olutiola, P O

    1989-09-01

    The quantity of lactase produced by enteropathogenic Escherichia coli (EPEC) was significantly higher (P less than 0.01) than that produced by non-EPEC. The enzyme production was induced by lactose but repressed by glucose and galactose. The lactase from EPEC which was partially purified by ammonium sulphate precipitation and gel permeation chromatography had a molecular weight of 56 kD and apparent Km of approximately 2.78 mM for lactose. The lactase exhibited optimum activity at pH 7.0 at 40 degree C and was stimulated by Mg2+, Mn2+, Na+ and inhibited by Ba2+, Ca+, Cu2+, EDTA, iodo acetic acid (IAA) and Hg2+ and U2+ ions. The higher production of lactase by EPEC may be linked to its pathogenic role in childhood diarrhoea.

  16. 60Co irradiation of Shiga toxin (Stx)-producing Escherichia coli induces Stx phage.

    PubMed

    Yamamoto, Tatsuo; Kojio, Seiichi; Taneike, Ikue; Nakagawa, Saori; Iwakura, Nobuhiro; Wakisaka-Saito, Noriko

    2003-05-16

    Shiga toxin (Stx)-producing Escherichia coli (STEC), an important cause of hemolytic uremic syndrome, was completely killed by (60)Co irradiation at 1 x l0(3) gray (1 kGy) or higher. However, a low dose of irradiation (0.1-0.3 kGy) markedly induced Stx phage from STEC. Stx production was observed in parallel to the phage induction. Inactivation of Stx phage required a higher irradiation dose than that for bacterial killing. Regarding Stx, cytotoxicity was susceptible to irradiation, but cytokine induction activity was more resistant than Stx phage. The findings suggest that (1). although (60)Co irradiation is an effective means to kill the bacteria, it does induce Stx phage at a lower irradiation dose, with a risk of Stx phage transfer and emergence of new Stx-producing strains, and (2). irradiation differentially inactivates some activities of Stx.

  17. Characterization of Shiga Toxin Subtypes and Virulence Genes in Porcine Shiga Toxin-Producing Escherichia coli.

    PubMed

    Baranzoni, Gian Marco; Fratamico, Pina M; Gangiredla, Jayanthi; Patel, Isha; Bagi, Lori K; Delannoy, Sabine; Fach, Patrick; Boccia, Federica; Anastasio, Aniello; Pepe, Tiziana

    2016-01-01

    Similar to ruminants, swine have been shown to be a reservoir for Shiga toxin-producing Escherichia coli (STEC), and pork products have been linked with outbreaks associated with STEC O157 and O111:H-. STEC strains, isolated in a previous study from fecal samples of late-finisher pigs, belonged to a total of 56 serotypes, including O15:H27, O91:H14, and other serogroups previously associated with human illness. The isolates were tested by polymerase chain reaction (PCR) and a high-throughput real-time PCR system to determine the Shiga toxin (Stx) subtype and virulence-associated and putative virulence-associated genes they carried. Select STEC strains were further analyzed using a Minimal Signature E. coli Array Strip. As expected, stx 2e (81%) was the most common Stx variant, followed by stx 1a (14%), stx 2d (3%), and stx 1c (1%). The STEC serogroups that carried stx 2d were O15:H27, O159:H16 and O159:H-. Similar to stx 2a and stx 2c, the stx 2d variant is associated with development of hemorrhagic colitis and hemolytic uremic syndrome, and reports on the presence of this variant in STEC strains isolated from swine are lacking. Moreover, the genes encoding heat stable toxin (estIa) and enteroaggregative E. coli heat stable enterotoxin-1 (astA) were commonly found in 50 and 44% of isolates, respectively. The hemolysin genes, hlyA and ehxA, were both detected in 7% of the swine STEC strains. Although the eae gene was not found, other genes involved in host cell adhesion, including lpfAO113 and paa were detected in more than 50% of swine STEC strains, and a number of strains also carried iha, lpfAO26, lpfAO157, fedA, orfA, and orfB. The present work provides new insights on the distribution of virulence factors among swine STEC strains and shows that swine may carry Stx1a-, Stx2e-, or Stx2d-producing E. coli with virulence gene profiles associated with human infections. PMID:27148249

  18. Characterization of Shiga toxin subtypes and virulence genes in porcine Shiga toxin-producing Escherichia coli

    DOE PAGES

    Baranzoni, Gian Marco; Fratamico, Pina M.; Gangiredla, Jayanthi; Patel, Isha; Bagi, Lori K.; Delannoy, Sabine; Fach, Patrick; Boccia, Federica; Anastasio, Aniello; Pepe, Tiziana

    2016-04-21

    Similar to ruminants, swine have been shown to be a reservoir for Shiga toxin-producing Escherichia coli (STEC), and pork products have been linked with outbreaks associated with STEC O157 and O111:H-. STEC strains, isolated in a previous study from fecal samples of late-finisher pigs, belonged to a total of 56 serotypes, including O15:H27, O91:H14, and other serogroups previously associated with human illness. The isolates were tested by polymerase chain reaction (PCR) and a high-throughput real-time PCR system to determine the Shiga toxin (Stx) subtype and virulence-associated and putative virulence-associated genes they carried. Select STEC strains were further analyzed using amore » Minimal Signature E. coli Array Strip. As expected, stx2e (81%) was the most common Stx variant, followed by stx1a (14%), stx2d (3%), and stx1c (1%). The STEC serogroups that carried stx2d were O15:H27, O159:H16 and O159:H-. Similar to stx2a and stx2c, the stx2d variant is associated with development of hemorrhagic colitis and hemolytic uremic syndrome, and reports on the presence of this variant in STEC strains isolated from swine are lacking. Moreover, the genes encoding heat stable toxin (estIa) and enteroaggregative E. coli heat stable enterotoxin-1 (astA) were commonly found in 50 and 44% of isolates, respectively. The hemolysin genes, hlyA and ehxA, were both detected in 7% of the swine STEC strains. Although the eae gene was not found, other genes involved in host cell adhesion, including lpfAO113 and paa were detected in more than 50% of swine STEC strains, and a number of strains also carried iha, lpfAO26, lpfAO157, fedA, orfA, and orfB. Furthermore, the present work provides new insights on the distribution of virulence factors among swine STEC strains and shows that swine may carry Stx1a-, Stx2e-, or Stx2d-producing E. coli with virulence gene profiles associated with human infections.« less

  19. Characterization of Shiga Toxin Subtypes and Virulence Genes in Porcine Shiga Toxin-Producing Escherichia coli

    PubMed Central

    Baranzoni, Gian Marco; Fratamico, Pina M.; Gangiredla, Jayanthi; Patel, Isha; Bagi, Lori K.; Delannoy, Sabine; Fach, Patrick; Boccia, Federica; Anastasio, Aniello; Pepe, Tiziana

    2016-01-01

    Similar to ruminants, swine have been shown to be a reservoir for Shiga toxin-producing Escherichia coli (STEC), and pork products have been linked with outbreaks associated with STEC O157 and O111:H-. STEC strains, isolated in a previous study from fecal samples of late-finisher pigs, belonged to a total of 56 serotypes, including O15:H27, O91:H14, and other serogroups previously associated with human illness. The isolates were tested by polymerase chain reaction (PCR) and a high-throughput real-time PCR system to determine the Shiga toxin (Stx) subtype and virulence-associated and putative virulence-associated genes they carried. Select STEC strains were further analyzed using a Minimal Signature E. coli Array Strip. As expected, stx2e (81%) was the most common Stx variant, followed by stx1a (14%), stx2d (3%), and stx1c (1%). The STEC serogroups that carried stx2d were O15:H27, O159:H16 and O159:H-. Similar to stx2a and stx2c, the stx2d variant is associated with development of hemorrhagic colitis and hemolytic uremic syndrome, and reports on the presence of this variant in STEC strains isolated from swine are lacking. Moreover, the genes encoding heat stable toxin (estIa) and enteroaggregative E. coli heat stable enterotoxin-1 (astA) were commonly found in 50 and 44% of isolates, respectively. The hemolysin genes, hlyA and ehxA, were both detected in 7% of the swine STEC strains. Although the eae gene was not found, other genes involved in host cell adhesion, including lpfAO113 and paa were detected in more than 50% of swine STEC strains, and a number of strains also carried iha, lpfAO26, lpfAO157, fedA, orfA, and orfB. The present work provides new insights on the distribution of virulence factors among swine STEC strains and shows that swine may carry Stx1a-, Stx2e-, or Stx2d-producing E. coli with virulence gene profiles associated with human infections. PMID:27148249

  20. Clonal spread and interspecies transmission of clinically relevant ESBL-producing Escherichia coli of ST410--another successful pandemic clone?

    PubMed

    Schaufler, Katharina; Semmler, Torsten; Wieler, Lothar H; Wöhrmann, Michael; Baddam, Ramani; Ahmed, Niyaz; Müller, Kerstin; Kola, Axel; Fruth, Angelika; Ewers, Christa; Guenther, Sebastian

    2016-01-01

    Clinically relevant extended-spectrum beta-lactamase (ESBL)-producing multi-resistant Escherichia coli have been on the rise for years. Initially restricted to mostly a clinical context, recent findings prove their prevalence in extraclinical settings independent of the original occurrence of antimicrobial resistance in the environment. To get further insights into the complex ecology of potentially clinically relevant ESBL-producing E. coli, 24 isolates from wild birds in Berlin, Germany, and 40 ESBL-producing human clinical E. coli isolates were comparatively analyzed. Isolates of ST410 occurred in both sample groups (six). In addition, three ESBL-producing E. coli isolates of ST410 from environmental dog feces and one clinical dog isolate were included. All 10 isolates were clonally analyzed showing almost identical macrorestriction patterns. They were chosen for whole-genome sequencing revealing that the whole-genome content of these 10 E. coli isolates showed a very high genetic similarity, differing by low numbers of single nucleotide polymorphisms only. This study gives initial evidence for a recent interspecies transmission of a new successful clone of ST410 E. coli between wildlife, humans, companion animals and the environment. The results underline the zoonotic potential of clinically relevant multi-resistant bacteria found in the environment as well as the mandatory nature of the 'One Health' approach.

  1. Clonal spread and interspecies transmission of clinically relevant ESBL-producing Escherichia coli of ST410--another successful pandemic clone?

    PubMed

    Schaufler, Katharina; Semmler, Torsten; Wieler, Lothar H; Wöhrmann, Michael; Baddam, Ramani; Ahmed, Niyaz; Müller, Kerstin; Kola, Axel; Fruth, Angelika; Ewers, Christa; Guenther, Sebastian

    2016-01-01

    Clinically relevant extended-spectrum beta-lactamase (ESBL)-producing multi-resistant Escherichia coli have been on the rise for years. Initially restricted to mostly a clinical context, recent findings prove their prevalence in extraclinical settings independent of the original occurrence of antimicrobial resistance in the environment. To get further insights into the complex ecology of potentially clinically relevant ESBL-producing E. coli, 24 isolates from wild birds in Berlin, Germany, and 40 ESBL-producing human clinical E. coli isolates were comparatively analyzed. Isolates of ST410 occurred in both sample groups (six). In addition, three ESBL-producing E. coli isolates of ST410 from environmental dog feces and one clinical dog isolate were included. All 10 isolates were clonally analyzed showing almost identical macrorestriction patterns. They were chosen for whole-genome sequencing revealing that the whole-genome content of these 10 E. coli isolates showed a very high genetic similarity, differing by low numbers of single nucleotide polymorphisms only. This study gives initial evidence for a recent interspecies transmission of a new successful clone of ST410 E. coli between wildlife, humans, companion animals and the environment. The results underline the zoonotic potential of clinically relevant multi-resistant bacteria found in the environment as well as the mandatory nature of the 'One Health' approach. PMID:26656065

  2. Diversity and relatedness of Shiga toxin-producing Escherichia coli and Campylobacter jejuni between farms in a dairy catchment.

    PubMed

    Irshad, H; Cookson, A L; Ross, C M; Jaros, P; Prattley, D J; Donnison, A; McBRIDE, G; Marshall, J; French, N P

    2016-05-01

    The aim of this study was to examine the population structure, transmission and spatial relationship between genotypes of Shiga toxin-producing Escherichia coli (STEC) and Campylobacter jejuni, on 20 dairy farms in a defined catchment. Pooled faecal samples (n = 72) obtained from 288 calves were analysed by real-time polymerase chain reaction (rtPCR) for E. coli serotypes O26, O103, O111, O145 and O157. The number of samples positive for E. coli O26 (30/72) was high compared to E. coli O103 (7/72), O145 (3/72), O157 (2/72) and O111 (0/72). Eighteen E. coli O26 and 53 C. jejuni isolates were recovered from samples by bacterial culture. E. coli O26 and C. jejuni isolates were genotyped using pulsed-field gel electrophoresis and multilocus sequence typing, respectively. All E. coli O26 isolates could be divided into four clusters and the results indicated that E. coli O26 isolates recovered from calves on the same farm were more similar than isolates recovered from different farms in the catchment. There were 11 different sequence types of C. jejuni isolated from the cattle and 22 from water. An analysis of the population structure of C. jejuni isolated from cattle provided evidence of clustering of genotypes within farms, and among groups of farms separated by road boundaries.

  3. Detection of Extended-Spectrum Beta-Lactamase (ESBL)-Producing Escherichia coli on Flies at Poultry Farms

    PubMed Central

    Hamidjaja, Raditijo A.; van Hoek, Angela H. A. M.; de Heer, Lianne; de Roda Husman, Ana Maria; Schets, Franciska M.

    2014-01-01

    In the Netherlands, extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli bacteria are highly prevalent in poultry, and chicken meat has been implicated as a source of ESBL-producing E. coli present in the human population. The current study describes the isolation of ESBL-producing E. coli from house flies and blow flies caught at two poultry farms, offering a potential alternative route of transmission of ESBL-producing E. coli from poultry to humans. Overall, 87 flies were analyzed in 19 pools. ESBL-producing E. coli bacteria were detected in two fly pools (10.5%): a pool of three blow flies from a broiler farm and a pool of eight house flies from a laying-hen farm. From each positive fly pool, six isolates were characterized and compared with isolates obtained from manure (n = 53) sampled at both farms and rinse water (n = 10) from the broiler farm. Among six fly isolates from the broiler farm, four different types were detected with respect to phylogenetic group, sequence type (ST), and ESBL genotype: A0/ST3519/SHV-12, A1/ST10/SHV-12, A1/ST58/SHV-12, and B1/ST448/CTX-M-1. These types, as well as six additional types, were also present in manure and/or rinse water at the same farm. At the laying-hen farm, all fly and manure isolates were identical, carrying blaTEM-52 in an A1/ST48 genetic background. The data imply that flies acquire ESBL-producing E. coli at poultry farms, warranting further evaluation of the contribution of flies to dissemination of ESBL-producing E. coli in the community. PMID:24162567

  4. Occurrence of Escherichia coli, Campylobcter, Salmonella and Shiga-Toxin Producing E. coli in Norwegian Primary Strawberry Production.

    PubMed

    Johannessen, Gro S; Eckner, Karl F; Heiberg, Nina; Monshaugen, Marte; Begum, Mumtaz; Økland, Marianne; Høgåsen, Helga R

    2015-06-01

    The aim of this study was to investigate the bacteriological quality of strawberries at harvest and to study risk factors such as irrigation water, soil and picker's hand cleanliness. Four farms were visited during the harvest season in 2012. Samples of strawberries, irrigation water, soil and hand swabs were collected and analyzed for E. coli, Campylobacter, Salmonella and STEC Although fecal indicators and pathogens were found in environmental samples, only one of 80 samples of strawberries was positive for E. coli (1.0 log10 cfu/g) and pathogens were not detected in any of the strawberry samples. The water samples from all irrigation sources were contaminated with E. coli in numbers ranging from 0 to 3.3 log10 cfu/g. Campylobacter (8/16 samples) and Salmonella (1/16 samples) were isolated from samples with high numbers of E. coli. The water samples collected from a lake had lower numbers of E. coli than the samples from rivers and a stream. The present study indicated continuous background contamination in the primary production environment. Although the background contamination was not reflected on the strawberries tested here, the results must be interpreted with caution due to the limited number of samples. PMID:26090606

  5. Occurrence of Escherichia coli, Campylobcter, Salmonella and Shiga-Toxin Producing E. coli in Norwegian Primary Strawberry Production

    PubMed Central

    Johannessen, Gro S.; Eckner, Karl F.; Heiberg, Nina; Monshaugen, Marte; Begum, Mumtaz; Økland, Marianne; Høgåsen, Helga R.

    2015-01-01

    The aim of this study was to investigate the bacteriological quality of strawberries at harvest and to study risk factors such as irrigation water, soil and picker’s hand cleanliness. Four farms were visited during the harvest season in 2012. Samples of strawberries, irrigation water, soil and hand swabs were collected and analyzed for E. coli, Campylobacter, Salmonella and STEC Although fecal indicators and pathogens were found in environmental samples, only one of 80 samples of strawberries was positive for E. coli (1.0 log10 cfu/g) and pathogens were not detected in any of the strawberry samples. The water samples from all irrigation sources were contaminated with E. coli in numbers ranging from 0 to 3.3 log10 cfu/g. Campylobacter (8/16 samples) and Salmonella (1/16 samples) were isolated from samples with high numbers of E. coli. The water samples collected from a lake had lower numbers of E. coli than the samples from rivers and a stream. The present study indicated continuous background contamination in the primary production environment. Although the background contamination was not reflected on the strawberries tested here, the results must be interpreted with caution due to the limited number of samples. PMID:26090606

  6. Occurrence of Escherichia coli, Campylobcter, Salmonella and Shiga-Toxin Producing E. coli in Norwegian Primary Strawberry Production.

    PubMed

    Johannessen, Gro S; Eckner, Karl F; Heiberg, Nina; Monshaugen, Marte; Begum, Mumtaz; Økland, Marianne; Høgåsen, Helga R

    2015-06-01

    The aim of this study was to investigate the bacteriological quality of strawberries at harvest and to study risk factors such as irrigation water, soil and picker's hand cleanliness. Four farms were visited during the harvest season in 2012. Samples of strawberries, irrigation water, soil and hand swabs were collected and analyzed for E. coli, Campylobacter, Salmonella and STEC Although fecal indicators and pathogens were found in environmental samples, only one of 80 samples of strawberries was positive for E. coli (1.0 log10 cfu/g) and pathogens were not detected in any of the strawberry samples. The water samples from all irrigation sources were contaminated with E. coli in numbers ranging from 0 to 3.3 log10 cfu/g. Campylobacter (8/16 samples) and Salmonella (1/16 samples) were isolated from samples with high numbers of E. coli. The water samples collected from a lake had lower numbers of E. coli than the samples from rivers and a stream. The present study indicated continuous background contamination in the primary production environment. Although the background contamination was not reflected on the strawberries tested here, the results must be interpreted with caution due to the limited number of samples.

  7. Whole genome sequencing of diverse Shiga toxin-producing and non-producing Escherichia coli strains reveals a variety of virulence and novel antibiotic resistance plasmids.

    PubMed

    Losada, Liliana; DebRoy, Chitrita; Radune, Diana; Kim, Maria; Sanka, Ravi; Brinkac, Lauren; Kariyawasam, Subhashinie; Shelton, Daniel; Fratamico, Pina M; Kapur, Vivek; Feng, Peter C H

    2016-01-01

    The genomes of a diverse set of Escherichia coli, including many Shiga toxin-producing strains of various serotypes were determined. A total of 39 plasmids were identified among these strains, and many carried virulence or putative virulence genes of Shiga toxin-producing E. coli strains, virulence genes for other pathogenic E. coli groups, and some had combinations of these genes. Among the novel plasmids identified were eight that carried resistance genes to aminoglycosides, carbapenems, penicillins, cephalosporins, chloramphenicol, dihydrofolate reductase inhibitors, sulfonamides, tetracyclines and resistance to heavy metals. Two of the plasmids carried six of these resistance genes and two novel IncHI2 plasmids were also identified. The results of this study showed that plasmids carrying diverse resistance and virulence genes of various pathogenic E. coli groups can be found in E. coli strains and serotypes regardless of the isolate's source and therefore, is consistent with the premise that these mobile elements carrying these traits may be broadly disseminated among E. coli.

  8. Whole genome sequencing of diverse Shiga toxin-producing and non-producing Escherichia coli strains reveals a variety of virulence and novel antibiotic resistance plasmids.

    PubMed

    Losada, Liliana; DebRoy, Chitrita; Radune, Diana; Kim, Maria; Sanka, Ravi; Brinkac, Lauren; Kariyawasam, Subhashinie; Shelton, Daniel; Fratamico, Pina M; Kapur, Vivek; Feng, Peter C H

    2016-01-01

    The genomes of a diverse set of Escherichia coli, including many Shiga toxin-producing strains of various serotypes were determined. A total of 39 plasmids were identified among these strains, and many carried virulence or putative virulence genes of Shiga toxin-producing E. coli strains, virulence genes for other pathogenic E. coli groups, and some had combinations of these genes. Among the novel plasmids identified were eight that carried resistance genes to aminoglycosides, carbapenems, penicillins, cephalosporins, chloramphenicol, dihydrofolate reductase inhibitors, sulfonamides, tetracyclines and resistance to heavy metals. Two of the plasmids carried six of these resistance genes and two novel IncHI2 plasmids were also identified. The results of this study showed that plasmids carrying diverse resistance and virulence genes of various pathogenic E. coli groups can be found in E. coli strains and serotypes regardless of the isolate's source and therefore, is consistent with the premise that these mobile elements carrying these traits may be broadly disseminated among E. coli. PMID:26746359

  9. A longitudinal study of Vero cytotoxin producing Escherichia coli in cattle calves in Sri Lanka.

    PubMed Central

    Tokhi, A. M.; Peiris, J. S.; Scotland, S. M.; Willshaw, G. A.; Smith, H. R.; Cheasty, T.

    1993-01-01

    Two cohorts of 10 and 16 calves were followed at weekly or fortnightly intervals from 4-28 and 1-9 weeks respectively to determine whether natural infection by Vero cytotoxin (VT) producing Escherichia coli (VTEC) occurred. Ninety-one of 171 (53%) faecal specimens were VTEC positive and 20-80% of animals at any given time excreted VTEC. Of 104 VTEC strains studied further, 6 different serogroups (O 22.H16; O 25.H5; O 49.H-; O 86.H26; O 88.H25; O 153.H12) and an untypable strain (O? .H21) were identified. All strains belonging to the same serotype had identical profiles of reactivity with DNA probes to toxins VT1 or 2, LTI or II and a probe (CVD419) derived from a plasmid carried by enterohaemorrhagic Escherichia coli O 157.H7. Four of these serotypes were found in the faecal flora of the calves, taken as a group, throughout the 4-month study period. Sixty percent of the strains hybridized with the probe for VT1, 4% with the probe for VT2, and 36% with both probes. Faecal VTEC were significantly associated with overt diarrhoeal illness in animals < 10 weeks of age, but no characteristic profile of markers (serotype or hybridization pattern) in E. coli isolates was associated with diarrhoea. A serological response to VT1 was detected in some animals, but faecal VT1 VTEC excretion persisted in spite of seroconversion. VT1 seroconversion was not associated with diarrhoea. A serological response to VT2 was not detected even in those animals excreting VT2 VTEC in the faeces. PMID:8472764

  10. Genotypes and phenotypes of Shiga toxin-producing Escherichia coli (STEC) in Abeokuta, Southwestern Nigeria

    PubMed Central

    Olowe, Olugbenga Adekunle; Aboderin, Bukola W; Idris, Olayinka O; Mabayoje, Victor O; Opaleye, Oluyinka O; Adekunle, O Catherine; Olowe, Rita Ayanbolade; Akinduti, Paul Akinniyi; Ojurongbe, Olusola

    2014-01-01

    Purpose To characterize the prevalence of hemolytic Shiga toxin-producing Escherichia coli (STEC) with a multidrug-resistant pattern in different age groups in Abeokuta, Nigeria. Methods Nonrepetitive E. coli isolates were collected from 202 subjects with or without evidence of diarrhea. Each isolate was biochemically identified and antimicrobial susceptibility testing was performed using the disk diffusion method. A sorbitol fermentation test of all the E. coli isolates was done and the minimum inhibitory concentration of suspected STEC was measured by the standard broth microdilution method to determine antibiotic resistance. The genotypes of stx1, stx2, and hlyA were determined by polymerase chain reaction assay. Results The majority of subjects were aged ≥40 years (41.6%) and were female (61.9%). Of the 202 subjects, 86.1% had STEC isolates (P<0.05). A high rate of STEC isolates resistant to amoxicillin (90.6%), cefotaxime (77.7%), and cefuroxime (75.7%) was observed. Resistance to amoxicillin, gentamicin, and cefotaxime was demonstrated with a minimum inhibitory concentration >16 μg/mL in 13.9%, 11.4%, and 10.4% of the isolates, respectively. The prevalence of stx1, stx2, and hlyA was 13.9%, 6.9%, and 2.0%, respectively; 5.5% of stx1 were in the 0–10-year-old age group, 3.5% of stx2 were aged ≥40 and above, and 1.0% of the hlyA isolates were in the 0–10-year-old age group. Conclusion The prevalence of virulent STEC is a public health concern. The use of polymerase chain reaction assay should aid quick detection of this virulent serotype and help curb the severe epidemic of human diseases associated with STEC infections. PMID:25342913

  11. Phylogenetic and molecular analysis of food-borne shiga toxin-producing Escherichia coli.

    PubMed

    Hauser, Elisabeth; Mellmann, Alexander; Semmler, Torsten; Stoeber, Helen; Wieler, Lothar H; Karch, Helge; Kuebler, Nikole; Fruth, Angelika; Harmsen, Dag; Weniger, Thomas; Tietze, Erhard; Schmidt, Herbert

    2013-04-01

    Seventy-five food-associated Shiga toxin-producing Escherichia coli (STEC) strains were analyzed by molecular and phylogenetic methods to describe their pathogenic potential. The presence of the locus of proteolysis activity (LPA), the chromosomal pathogenicity island (PAI) PAI ICL3, and the autotransporter-encoding gene sabA was examined by PCR. Furthermore, the occupation of the chromosomal integration sites of the locus of enterocyte effacement (LEE), selC, pheU, and pheV, as well as the Stx phage integration sites yehV, yecE, wrbA, z2577, and ssrA, was analyzed. Moreover, the antibiotic resistance phenotypes of all STEC strains were determined. Multilocus sequence typing (MLST) was performed, and sequence types (STs) and sequence type complexes (STCs) were compared with those of 42 hemolytic-uremic syndrome (HUS)-associated enterohemorrhagic E. coli (HUSEC) strains. Besides 59 STs and 4 STCs, three larger clusters were defined in this strain collection. Clusters A and C consist mostly of highly pathogenic eae-positive HUSEC strains and some related food-borne STEC strains. A member of a new O26 HUS-associated clone and the 2011 outbreak strain E. coli O104:H4 were found in cluster A. Cluster B comprises only eae-negative food-borne STEC strains as well as mainly eae-negative HUSEC strains. Although food-borne strains of cluster B were not clearly associated with disease, serotypes of important pathogens, such as O91:H21 and O113:H21, were in this cluster and closely related to the food-borne strains. Clonal analysis demonstrated eight closely related genetic groups of food-borne STEC and HUSEC strains that shared the same ST and were similar in their virulence gene composition. These groups should be considered with respect to their potential for human infection.

  12. Comparative genomics and stx phage characterization of LEE-negative Shiga toxin-producing Escherichia coli.

    PubMed

    Steyert, Susan R; Sahl, Jason W; Fraser, Claire M; Teel, Louise D; Scheutz, Flemming; Rasko, David A

    2012-01-01

    Infection by Escherichia coli and Shigella species are among the leading causes of death due to diarrheal disease in the world. Shiga toxin-producing E. coli (STEC) that do not encode the locus of enterocyte effacement (LEE-negative STEC) often possess Shiga toxin gene variants and have been isolated from humans and a variety of animal sources. In this study, we compare the genomes of nine LEE-negative STEC harboring various stx alleles with four complete reference LEE-positive STEC isolates. Compared to a representative collection of prototype E. coli and Shigella isolates representing each of the pathotypes, the whole genome phylogeny demonstrated that these isolates are diverse. Whole genome comparative analysis of the 13 genomes revealed that in addition to the absence of the LEE pathogenicity island, phage-encoded genes including non-LEE encoded effectors, were absent from all nine LEE-negative STEC genomes. Several plasmid-encoded virulence factors reportedly identified in LEE-negative STEC isolates were identified in only a subset of the nine LEE-negative isolates further confirming the diversity of this group. In combination with whole genome analysis, we characterized the lambdoid phages harboring the various stx alleles and determined their genomic insertion sites. Although the integrase gene sequence corresponded with genomic location, it was not correlated with stx variant, further highlighting the mosaic nature of these phages. The transcription of these phages in different genomic backgrounds was examined. Expression of the Shiga toxin genes, stx(1) and/or stx(2), as well as the Q genes, were examined with quantitative reverse transcriptase polymerase chain reaction assays. A wide range of basal and induced toxin induction was observed. Overall, this is a first significant foray into the genome space of this unexplored group of emerging and divergent pathogens.

  13. 2′-Deoxyribonolactone lesion produces G→A transitions in Escherichia coli

    PubMed Central

    Faure, Virginie; Constant, Jean-François; Dumy, Pascal; Saparbaev, Murat

    2004-01-01

    2′-Deoxyribonolactone (dL) is a C1′-oxidized abasic site damage generated by a radical attack on DNA. Numerous genotoxic agents have been shown to produce dL including UV and γ-irradiation, ene-dye antibiotics etc. At present the biological consequences of dL present in DNA have been poorly documented, mainly due to the lack of method for introducing the lesion in oligonucleotides. We have recently designed a synthesis of dL which allowed investigation of the mutagenicity of dL in Escherichia coli by using a genetic reversion assay. The lesion was site-specifically incorporated in a double-stranded bacteriophage vector M13G*1, which detects single-base-pair substitutions at position 141 of the lacZα gene by a change in plaque color. In E.coli JM105 the dL-induced reversion frequency was 4.7 × 10–5, similar to that of the classic abasic site 2′-deoxyribose (dR). Here we report that a dL residue in a duplex DNA codes mainly for thymidine. The processing of dL in vivo was investigated by measuring lesion-induced mutation frequencies in DNA repair deficient E.coli strains. We showed a 32-fold increase in dL-induced reversion rate in AP endonuclease deficient (xth nfo) mutant compared with wild-type strain, indicating that the Xth and Nfo AP endonucleases participate in dL repair in vivo. PMID:15159441

  14. Shiga Toxin-Producing Escherichia coli O104:H4: a New Challenge for Microbiology

    PubMed Central

    Muniesa, Maite; Hammerl, Jens A.; Hertwig, Stefan; Appel, Bernd

    2012-01-01

    In 2011, Germany experienced the largest outbreak with a Shiga toxin-producing Escherichia coli (STEC) strain ever recorded. A series of environmental and trace-back and trace-forward investigations linked sprout consumption with the disease, but fecal-oral transmission was also documented. The genome sequences of the pathogen revealed a clonal outbreak with enteroaggregative E. coli (EAEC). Some EAEC virulence factors are carried on the virulence plasmid pAA. From an unknown source, the epidemic strains acquired a lambdoid prophage carrying the gene for the Shiga toxin. The resulting strains therefore possess two different mobile elements, a phage and a plasmid, contributing essential virulence genes. Shiga toxin is released by decaying bacteria in the gut, migrates through the intestinal barrier, and is transported via the blood to target organs, like the kidney. In a mouse model, probiotic bifidobacteria interfered with transport of the toxin through the gut mucosa. Researchers explored bacteriophages, bacteriocins, and low-molecular-weight inhibitors against STEC. Randomized controlled clinical trials of enterohemorrhagic E. coli (EHEC)-associated hemolytic uremic syndrome (HUS) patients found none of the interventions superior to supportive therapy alone. Antibodies against one subtype of Shiga toxin protected pigs against fatal neurological infection, while treatment with a toxin receptor decoy showed no effect in a clinical trial. Likewise, a monoclonal antibody directed against a complement protein led to mixed results. Plasma exchange and IgG immunoadsoprtion ameliorated the condition in small uncontrolled trials. The epidemic O104:H4 strains were resistant to all penicillins and cephalosporins but susceptible to carbapenems, which were recommended for treatment. PMID:22504816

  15. Effects of stresses on the growth and Cytotoxicity of Shiga-Toxin producing Escherichia coli in ground beef and spinach

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objectives of this study were to examine the effect of stresses on the growth and cytotoxicity of pathogenic Escherichia coli in beef and spinach. A mixture of three strains of Shiga toxin-producing E. coli (STEC) O157:H7 or four strains of non-O157 STEC O26:H11, O103:H1, O104:H4, and O145:NM wa...

  16. Prevalence of Quinolone Resistance Genes Among Extended-Spectrum B-Lactamase-Producing Escherichia coli in Mashhad, Iran

    PubMed Central

    Harifi Mood, Elnaz; Meshkat, Zahra; Izadi, Nafiseh; Rezaei, Maryam; Amel Jamehdar, Saeid; Naderi Nasab, Mahboubeh

    2015-01-01

    Background: Escherichia coli is an important bacterial species based on incidence and associated infection severity. Some E. coli strains produce extended-spectrum beta lactamase (ESBL) and are called ESBL-producing E. coli. These strains are resistant to most classes of cephalosporin and a number of other classes of antibiotics. Plasmids carrying qnr genes have been found to transmit quinolone resistance. Objectives: The aim of this study was to determine the frequency of qnr genes in ESBL-producing and non-ESBL-producing E. coli isolated from outpatient and hospitalized patient clinical specimens from Imam Reza hospital in Mashhad, Iran. Materials and Methods: Two hundred E. coli strains, isolated from different clinical specimens were used. ESBL-producing E. coli were detected by determining susceptibility to ceftazidime, cefotaxime, and cefpodoxime with the phenotypic confirmatory test (PCT). PCR analysis was employed to detect the qnrA, qnrB, qnrS, blaTEM, and blaSHV genes. Results: Eighty-six (43%) isolates were ciprofloxacin-resistant. The PCT identified 85 (42.5%) of 200 E. coli isolates as ESBL-producing. The blaTEM, blaSHV, qnrA, qnrB, and qnrS gene were found in 65 (76.47%), 23 (27%), 63 (31%), 34 (17%), and 14 (7%) isolates, respectively. Conclusions: The high prevalence of quinolone resistance genes, which indicates antibiotic resistance, in the Imam Reza Hospital of Mashhad is a major concern. Hence, the antibiotics prescription policy should be revised, and infection control measures should be improved. PMID:26870307

  17. Characterization of Shiga toxin – producing Escherichia coli infections in beef feeder calves and the effectiveness of a prebiotic in alleviating Shiga toxin - producing Escherichia coli infections

    PubMed Central

    2013-01-01

    Background In the less-sensitive mouse model, Shiga toxin-producing Escherichia coli (STEC) challenges result in shedding that reflect the amount of infection and the expression of virulence factors such as Shiga toxins (Stx). The purpose of this study was to characterize the contribution of STEC diversity and Stx expression to shedding in beef feeder calves and to evaluate the effectiveness of a prebiotic, Celmanax®, to alleviate STEC shedding. Fecal samples were collected from calves at entry and after 35 days in the feedlot in spring and summer. STECs were evaluated using selective media, biochemical profile, serotyping and Stx detection. Statistical analysis was performed using repeated measures ANOVA and logistic regression. Results At entry, non-O157 STEC were dominant in shedding calves. In spring, 21%, 14% and 14% of calves acquired O157, non-O157 and mixed STEC infections, respectively. In contrast, 45%, 48% and 46% of calves in summer acquired O157, non-O157 and mixed STEC infections, respectively. Treatment with a prebiotic, Celmanax®, in spring significantly reduced 50% of the O157 STEC infections, 50% of the non-O157 STEC infections and 36% of the STEC co-infections (P = 0.037). In summer, there was no significant effect of the prebiotic on STEC infections. The amount of shedding at entry was significantly related to the number and type of STECs present and Stx expression (r2 = 0.82). The same relationship was found for shedding at day 35 (r2 = 0.85), but it was also related to the number and type of STECs present at entry. Stx - producing STEC infections resulted in 100 to 1000 × higher shedding in calves compared with Stx-negative STECs. Conclusions STEC infections in beef feeder calves reflect the number and type of STECs involved in the infection and STEC expression of Stx. Application of Celmanax® reduced O157 and non-O157 STEC shedding by calves but further research is required to determine appropriate dosages to manage STEC

  18. Vero cytotoxin-producing Escherichia coli O157 gastroenteritis in farm visitors, North Wales.

    PubMed

    Payne, Christopher J I; Petrovic, Marko; Roberts, Richard J; Paul, Ashish; Linnane, Eithne; Walker, Mark; Kirby, David; Burgess, Anthony; Smith, Robert M M; Cheasty, Thomas; Willshaw, Geraldine; Salmon, Roland L

    2003-05-01

    An outbreak of Vero cytotoxin-producing Escherichia coli O157 (VTEC O157) gastroenteritis in visitors to an open farm in North Wales resulted in 17 primary and 7 secondary cases of illness. E. coli O157 Vero cytotoxin type 2, phage type 2 was isolated from 23 human cases and environmental animal fecal samples. A case-control study of 16 primary case-patients and 36 controls (all children) showed a significant association with attendance on the 2nd day of a festival, eating ice cream or cotton candy (candy floss), and contact with cows or goats. On multivariable analysis, only the association between illness and ice cream (odds ratio [OR]=11.99, 95% confidence interval [CI] 1.04 to 137.76) and cotton candy (OR=51.90, 95% CI 2.77 to 970.67) remained significant. In addition to supervised handwashing, we recommend that foods on open farms only be eaten in dedicated clean areas and that sticky foods be discouraged. PMID:12737734

  19. Vero Cytotoxin–Producing Escherichia coli O157 Gastroenteritis in Farm Visitors, North Wales

    PubMed Central

    Petrovic, Marko; Roberts, Richard J.; Paul, Ashish; Linnane, Eithne; Walker, Mark; Kirby, David; Burgess, Anthony; Smith, Robert M.M.; Cheasty, Thomas; Willshaw, Geraldine; Salmon, Roland L.

    2003-01-01

    An outbreak of Vero cytotoxin–producing Escherichia coli O157 (VTEC O157) gastroenteritis in visitors to an open farm in North Wales resulted in 17 primary and 7 secondary cases of illness. E. coli O157 Vero cytotoxin type 2, phage type 2 was isolated from 23 human cases and environmental animal fecal samples. A case-control study of 16 primary case-patients and 36 controls (all children) showed a significant association with attendance on the 2nd day of a festival, eating ice cream or cotton candy (candy floss), and contact with cows or goats. On multivariable analysis, only the association between illness and ice cream (odds ratio [OR]=11.99, 95% confidence interval [CI] 1.04 to 137.76) and cotton candy (OR=51.90, 95% CI 2.77 to 970.67) remained significant. In addition to supervised handwashing, we recommend that foods on open farms only be eaten in dedicated clean areas and that sticky foods be discouraged. PMID:12737734

  20. Shiga Toxin-Producing Escherichia Coli Isolated From Lettuce Samples in Tehran, Iran

    PubMed Central

    Mazaheri, Somayeh; Salmanzadeh Ahrabi, Siavosh; Aslani, Mohammad Mahdi

    2014-01-01

    Background: During the last decade, the prevalence of foodborne diseases due to contaminated food as well as the outbreaks of diseases due to Shiga toxin-producing Escherichia coli (STEC) strains has increased. Objectives: The aim of this study was to evaluate the prevalence and antibiotic resistance pattern of STEC strains in lettuce samples. Since lettuce is used as a raw vegetable in salads, the rates of infections caused by this vegetable are high. Materials and Methods: A total of 100 samples collected from Tehran, Iran, were transported to the laboratory, homogenized by a stomacher in E. coli broth containing cefixime, and cultured on MacConkey agar medium. Their DNA was extracted by boiling method and polymerase chain reaction (PCR) was performed, using five primers targeting the stx1, stx2, fliCh7, rbfO157, and eaeA genes. Susceptibility testing against ampicillin, imipenem, cephalosporin, tetracycline, aminoglycosides, chloramphenicol and quinolones was performed using disk diffusion method. Results: Eight samples were positive for presence of STEC strains, three contained stx1, five contained stx2, and one sample was positive for presence of both rbfO157 and fliCh7. They were susceptible to all the antibiotics except for ampicillin and tetracycline. Conclusions: This study indicated the contamination of lettuce by STEC strains and its possible role as the source of infection. Resistance to both tetracycline and ampicillin may be considered as an emergency alarm for a multidrug resistance of STEC strains. PMID:25774272

  1. Extended-Spectrum Beta-Lactamases Producing E. coli in Wildlife, yet Another Form of Environmental Pollution?

    PubMed Central

    Guenther, Sebastian; Ewers, Christa; Wieler, Lothar H.

    2011-01-01

    Wildlife is normally not exposed to clinically used antimicrobial agents but can acquire antimicrobial resistant bacteria through contact with humans, domesticated animals and the environment, where water polluted with feces seems to be the most important vector. Escherichia coli, an ubiquitous commensal bacterial species colonizing the intestinal tract of mammals and birds, is also found in the environment. Extended-spectrum beta-lactamases producing E. coli (ESBL-E. coli) represent a major problem in human and veterinary medicine, particular in nosocomial infections. Additionally an onset of community-acquired ESBL-E. coli infections and an emergence in livestock farming has been observed in recent years, suggesting a successful transmission as well as persistence of ESBL-E. coli strains outside clinical settings. Another parallel worldwide phenomenon is the spread of ESBL-E. coli into the environment beyond human and domesticated animal populations, and this seems to be directly influenced by antibiotic practice. This might be a collateral consequence of the community-onset of ESBL-E. coli infections but can result (a) in a subsequent colonization of wild animal populations which can turn into an infectious source or even a reservoir of ESBL-E. coli, (b) in a contribution of wildlife to the spread and transmission of ESBL-E. coli into fragile environmental niches, (c) in new putative infection cycles between wildlife, domesticated animals and humans, and (d) in problems in the medical treatment of wildlife. This review aims to summarize the current knowledge on ESBL-E. coli in wildlife, in turn underlining the need for more large scale investigations, in particular sentinel studies to monitor the impact of multiresistant bacteria on wildlife. PMID:22203818

  2. Prevalence and characteristics of ESBL-producing E. coli in Dutch recreational waters influenced by wastewater treatment plants.

    PubMed

    Blaak, Hetty; de Kruijf, Patrick; Hamidjaja, Raditijo A; van Hoek, Angela H A M; de Roda Husman, Ana Maria; Schets, Franciska M

    2014-07-16

    Outside health care settings, people may acquire ESBL-producing bacteria through different exposure routes, including contact with human or animal carriers or consumption of contaminated food. However, contact with faecally contaminated surface water may also represent a possible exposure route. The current study investigated the prevalence and characteristics of ESBL-producing Escherichia coli in four Dutch recreational waters and the possible role of nearby waste water treatment plants (WWTP) as contamination source. Isolates from recreational waters were compared with isolates from WWTP effluents, from surface water upstream of the WWTPs, at WWTP discharge points, and in connecting water bodies not influenced by the studied WWTPs. ESBL-producing E. coli were detected in all four recreational waters, with an average concentration of 1.3 colony forming units/100ml, and in 62% of all samples. In surface waters not influenced by the studied WWTPs, ESBL-producing E. coli were detected in similar concentrations, indicating the existence of additional ESBL-E. coli contamination sources. Isolates with identical ESBL-genes, phylogenetic background, antibiotic resistance profiles, and sequence type, were obtained from effluent and different surface water sites in the same watershed, on the same day; occasionally this included isolates from recreational waters. Recreational waters were identified as a potential exposure source of ESBL-producing E. coli. WWTPs were shown to contribute to the presence of these bacteria in surface waters, but other (yet unidentified) sources likely co-contribute.

  3. Extended-Spectrum Beta-Lactamases Producing Escherichia coli and Klebsiella pneumoniae: A Multi-Centric Study Across Karnataka

    PubMed Central

    Rao, Sridhar PN; Rama, Prasad Subba; Gurushanthappa, Vishwanath; Manipura, Radhakrishna; Srinivasan, Krishna

    2014-01-01

    Background: There are sporadic reports on detection of extended-spectrum beta-lactamases (ESBL) producers from Karnataka; hence, this is a first multicentric study across Karnataka state to determine the prevalence of ESBL production among clinical isolates of Escherichia coli and Klebsiella pneumoniae. Aims and objectives: To determine the prevalence of ESBL producing clinical isolates of E. coli and K. pneumoniae from five geographically distributed centers across Karnataka, to study the susceptibility of ESBL producing isolates to other beta-lactam and beta-lactam-beta-lactamase inhibitors and to demonstrate transferability of plasmids coding for ESBL phenotype. Materials and Methods: Two hundred isolates of E. coli and K. pneumoniae each were collected from each of the five centers (Bellary, Dharwad, Davangere, Kolar and Mangalore). They were screened for resistance to screening agents (ceftazidime, cefotaxime, ceftriaxone, aztreonam) and positive isolates were confirmed for ESBL production by test described by Clinical and Laboratory Standards Institute. Co-production of ESBL and AmpC beta-lactamase was identified by using amino-phenylboronic acid disk method. Susceptibility of ESBL producers to beta-lactam antibiotics and beta-lactamase inhibitors was performed. Transferability of plasmids was performed by conjugation experiment. Results: Overall prevalence of ESBL production among E. coli and K. pneumoniae across five centers of the state was 57.5%. ESBL production was found to be 61.4% among E. coli and 46.2% among K. pneumoniae. ESBL production was significantly more among E. coli than K. pneumoniae. Significant variations in distribution of ESBL across the state was observed among E. coli isolates, but not among K. pneumoniae isolates. All ESBL producers demonstrated minimum inhibitory concentration levels ≥2 μg/ml towards cefotaxime, ceftazidime and ceftriaxone. Conclusion: Overall prevalence of ESBL production among clinical isolates of E. coli and K

  4. Use of ramification amplification assay for detection of Escherichia coli O157:H7 and other E. coli Shiga toxin-producing strains.

    PubMed

    Li, Fan; Zhao, Chunyan; Zhang, Wandi; Cui, Shenghui; Meng, Jianghong; Wu, Josephine; Zhang, David Y

    2005-12-01

    Escherichia coli O157:H7 and other Shiga toxin-producing E. coli (STEC) strains are important human pathogens that are mainly transmitted through the food chain. These pathogens have a low infectious dose and may cause life-threatening illnesses. However, detection of this microorganism in contaminated food or a patient's stool specimens presents a diagnostic challenge because of the low copy number in the sample. Often, a more sensitive nucleic acid amplification method, such as PCR, is required for rapid detection of this microorganism. Ramification amplification (RAM) is a recently introduced isothermal DNA amplification technique that utilizes a circular probe for target detection and achieves exponential amplification through the mechanism of primer extension, strand displacement, and ramification. In this study, we synthesized a circular probe specific for the Shiga toxin 2 gene (stx(2)). Our results showed that as few as 10 copies of stx(2) could be detected, indicating that the RAM assay was as sensitive as conventional PCR. We further tested 33 isolates of E coli O157:H7, STEC, Shigella dysenteriae, and nonpathogenic E. coli by RAM assay. Results showed that all 27 STEC isolates containing the stx(2) gene were identified by RAM assay, while S. dysenteriae and nonpathogenic E. coli isolates were undetected. The RAM results were 100% in concordance with those of PCR. Because of its simplicity and isothermal amplification, the RAM assay could be a useful method for detecting STEC in food and human specimens.

  5. Risk factors for sporadic Shiga toxin-producing Escherichia coli infections in children, Argentina.

    PubMed

    Rivas, Marta; Sosa-Estani, Sergio; Rangel, Josefa; Caletti, Maria G; Vallés, Patricia; Roldán, Carlos D; Balbi, Laura; Marsano de Mollar, Maria C; Amoedo, Diego; Miliwebsky, Elizabeth; Chinen, Isabel; Hoekstra, Robert M; Mead, Paul; Griffin, Patricia M

    2008-05-01

    We evaluated risk factors for sporadic Shiga toxin-producing Escherichia coli (STEC) infection among children in Argentina. We conducted a prospective case-control study in 2 sites and enrolled 150 case-patients and 299 controls. The median age of case-patients was 1.8 years; 58% were girls. Serotype O157:H7 was the most commonly isolated STEC. Exposures associated with infection included eating undercooked beef, living in or visiting a place with farm animals, and contact with a child <5 years of age with diarrhea. Protective factors included the respondent reporting that he or she always washed hands after handling raw beef and the child eating more than the median number of fruits and vegetables. Many STEC infections in children could be prevented by avoiding consumption of undercooked beef, limiting exposure to farm animals and their environment, not being exposed to children with diarrhea, and washing hands after handling raw beef.

  6. Risk Factors for Sporadic Shiga Toxin–producing Escherichia coli Infections in Children, Argentina1

    PubMed Central

    Rivas, Marta; Sosa-Estani, Sergio; Rangel, Josefa; Caletti, Maria G.; Vallés, Patricia; Roldán, Carlos D.; Balbi, Laura; Marsano de Mollar, Maria C.; Amoedo, Diego; Miliwebsky, Elizabeth; Chinen, Isabel; Hoekstra, Robert M.; Mead, Paul

    2008-01-01

    We evaluated risk factors for sporadic Shiga toxin–producing Escherichia coli (STEC) infection among children in Argentina. We conducted a prospective case–control study in 2 sites and enrolled 150 case-patients and 299 controls. The median age of case-patients was 1.8 years; 58% were girls. Serotype O157:H7 was the most commonly isolated STEC. Exposures associated with infection included eating undercooked beef, living in or visiting a place with farm animals, and contact with a child <5 years of age with diarrhea. Protective factors included the respondent reporting that he or she always washed hands after handling raw beef and the child eating more than the median number of fruits and vegetables. Many STEC infections in children could be prevented by avoiding consumption of undercooked beef, limiting exposure to farm animals and their environment, not being exposed to children with diarrhea, and washing hands after handling raw beef. PMID:18439359

  7. Molecular mechanisms that mediate colonization of Shiga toxin-producing Escherichia coli strains.

    PubMed

    Farfan, Mauricio J; Torres, Alfredo G

    2012-03-01

    Shiga toxin-producing Escherichia coli (STEC) is a group of pathogens which cause gastrointestinal disease in humans and have been associated with numerous food-borne outbreaks worldwide. The intimin adhesin has been considered for many years to be the only colonization factor in these strains. However, the rapid progress in whole-genome sequencing of different STEC serotypes has accelerated the discovery of other adhesins (fimbrial and afimbrial), which have emerged as important contributors to the intestinal colonization occurring during STEC infection. This review summarizes recent progress to identify and characterize, at the molecular level, novel adhesion and colonization factors in STEC strains, with an emphasis on their contribution to virulence traits, their host-pathogen interactions, the regulatory mechanisms controlling their expression, and their role as targets eliciting immune responses in the host.

  8. Molecular mechanisms that mediate colonization of Shiga toxin-producing Escherichia coli strains.

    PubMed

    Farfan, Mauricio J; Torres, Alfredo G

    2012-03-01

    Shiga toxin-producing Escherichia coli (STEC) is a group of pathogens which cause gastrointestinal disease in humans and have been associated with numerous food-borne outbreaks worldwide. The intimin adhesin has been considered for many years to be the only colonization factor in these strains. However, the rapid progress in whole-genome sequencing of different STEC serotypes has accelerated the discovery of other adhesins (fimbrial and afimbrial), which have emerged as important contributors to the intestinal colonization occurring during STEC infection. This review summarizes recent progress to identify and characterize, at the molecular level, novel adhesion and colonization factors in STEC strains, with an emphasis on their contribution to virulence traits, their host-pathogen interactions, the regulatory mechanisms controlling their expression, and their role as targets eliciting immune responses in the host. PMID:22144484

  9. Molecular Mechanisms That Mediate Colonization of Shiga Toxin-Producing Escherichia coli Strains

    PubMed Central

    Farfan, Mauricio J.

    2012-01-01

    Shiga toxin-producing Escherichia coli (STEC) is a group of pathogens which cause gastrointestinal disease in humans and have been associated with numerous food-borne outbreaks worldwide. The intimin adhesin has been considered for many years to be the only colonization factor in these strains. However, the rapid progress in whole-genome sequencing of different STEC serotypes has accelerated the discovery of other adhesins (fimbrial and afimbrial), which have emerged as important contributors to the intestinal colonization occurring during STEC infection. This review summarizes recent progress to identify and characterize, at the molecular level, novel adhesion and colonization factors in STEC strains, with an emphasis on their contribution to virulence traits, their host-pathogen interactions, the regulatory mechanisms controlling their expression, and their role as targets eliciting immune responses in the host. PMID:22144484

  10. Real-time isothermal detection of Shiga toxin-producing Escherichia coli using recombinase polymerase amplification.

    PubMed

    Murinda, Shelton E; Ibekwe, A Mark; Zulkaffly, Syaizul; Cruz, Andrew; Park, Stanley; Razak, Nur; Paudzai, Farah Md; Ab Samad, Liana; Baquir, Khairul; Muthaiyah, Kokilah; Santiago, Brenna; Rusli, Amirul; Balkcom, Sean

    2014-07-01

    Shiga toxin-producing Escherichia coli (STEC) are a major family of foodborne pathogens of public health, zoonotic, and economic significance in the United States and worldwide. To date, there are no published reports on use of recombinase polymerase amplification (RPA) for STEC detection. The primary goal of this study was to assess the potential application of RPA in detection of STEC. This study focused on designing and evaluating RPA primers and fluorescent probes for isothermal (39°C) detection of STEC. Compatible sets of candidate primers and probes were designed for detection of Shiga toxin 1 and 2 (Stx1 and 2), respectively. The sets were evaluated for specificity and sensitivity against STEC (n=12) of various stx genotypes (stx1/stx2, stx1, or stx2, respectively), including non-Stx-producing E. coli (n=28) and other genera (n=7). The primers and probes that were designed targeted amplification of the subunit A moiety of stx1 and stx2. The assay detected STEC in real time (within 5-10 min at 39°C) with high sensitivity (93.5% vs. 90%; stx1 vs. stx2), specificity (99.1% vs. 100%; stx1 vs. stx2), and predictive value (97.9% for both stx1 vs. stx2). Limits of detection of ∼ 5-50 colony-forming units/mL were achieved in serially diluted cultures grown in brain heart infusion broth. This study successfully demonstrated for the first time that RPA can be used for isothermal real-time detection of STEC.

  11. Incidence of antibiotic-resistant Escherichia coli in milk produced in the west of Scotland.

    PubMed

    Johnston, D W; Bruce, J; Hill, J

    1983-02-01

    Antibiotic-resistant Esch. coli were found in 10.6% of milk samples collected from 998 farms in the west of Scotland. The incidence of both Esch. coli and antibiotic-resistant Esch. coli in milk was higher when the cattle were housed day and night than when they were outdoors. Of the 1125 Esch. coli isolates tested 22.2% were antibiotic resistant and of these 42.4% were resistant to more than one antibiotic. Escherichia coli carrying up to six resistance determinants were isolated. The possible implications to animal and human health are discussed.

  12. Role of cellulose in protecting Shiga toxin-producing Escherichia coli against osmotic and chlorine stress.

    PubMed

    Yoo, Byong K; Chen, Jinru

    2010-11-01

    This study was undertaken to determine the role of cellulose in protecting Shiga toxin-producing Escherichia coli (STEC) against osmotic and chlorine treatments. STEC cells producing cellulose (19B and 49B) and their respective cellulose-deficient counterparts (19D or 49D) were subjected to osmotic (1, 2, and 3 M NaCl) or chlorine (25, 50, and 100 μg/ml sodium hypochlorite) treatments. The survival of STEC cells was determined at different treatment intervals. Populations of 19B cells were significantly higher (P < 0.05) than those of 19D cells at all sampling intervals for the chlorine treatments, at 24- to 48-h intervals for the 1 M NaCl treatment, and at 9- to 48-h intervals for the 2 M NaCl treatment. Significant differences in populations of 49B and 49D cells were observed after 9, 36, and 48 h of treatment with 2 M NaCl and after 3, 12, 36, and 48 h of treatment with 3 M NaCl (P < 0.05). Populations of 49B cells were higher than those of 49D cells (P < 0.05) also after 5 to 10 min of treatment with 50 μg/ml sodium hypochlorite and 3 to 10 min of treatment with 100 μg/ml sodium hypochlorite. The protective effects conferred by cellulose may explain the greater survival of cellulose-producing STEC under adverse environmental conditions. PMID:21219722

  13. EMERGENCY ROOM: AN UNRECOGNIZED SOURCE OF EXTENDED-SPECTRUM β-LACTAMASE PRODUCING ESCHERICHIA COLI AND KLEBSIELLA PNEUMONIAE.

    PubMed

    Pornsinchai, Pornsook; Chongtrakool, Piriyaporn; Diraphat, Pornphan; Siripanichgon, Kanokrat; Malathum, Kumthorn

    2015-01-01

    Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae are the leading causes of hospital-associated infections, but community-acquired cases are increasingly being reported. This study determined the prevalence of ESBL-producing E. coli and K. pneumoniae carriers, their bla genes and risk factors of 452 patients admitted to the emergency room (ER) of Ramathibodi Hospital, Mahidol University, Bangkok, Thailand between April and August 2011. Prevalence of ESBL-producing E. coli and K. pneumoniae from rectal swabs was 16.5% and 1.0%, respectively. Factors associated with ESBL-producing carriers were a previous history of hospital admission (p = 0.001) and visits to health care facilities (p = 0.002) during the previous 3 months. All ESBL-producing isolates were susceptible to imipenem, meropenem and ertapenem. The majority (78%) of ESBL-producing E. coli isolates showed very high resistance to cefotaxime and ceftriaxone (MIC50 and MIC90 > 256 µg/ml). ESBL-producing E. coli harbored chromosomal blaTEM (96%), blaCTX-M (70%) and blaSHV (1%), while 8%, 73% and 3%, respectively, were located on plasmid. The prevalence of these genes in ESBL-producing K. pneumoniae was 75%, 50% and 25%, respectively on chromosome; and 100%, 25% and 50%, respectively on plasmid. Nucleotide sequence analysis revealed that these bla genes were of the type blaTEM-1' blaTEM-116' blaCTX-M-15' blaCTX-M-161' blaSHV-12, blaSHV-28 and blaSHV-148. Detailed epidemiologic and clinical characteristics of ER patients with history of prior hospital visits should be carried out to identify the ESBL-producing organisms they have acquired in order to institute appropriate treatment for these patients as well as control measures against further dissemination of these life-threatening organisms. PMID:26513905

  14. A potential camel reservoir for extended-spectrum β-lactamase-producing Escherichia coli causing human infection in Saudi Arabia.

    PubMed

    Fadlelmula, Ali; Al-Hamam, Naser Abdallah; Al-Dughaym, Abdulla Mohamed

    2016-02-01

    The prevalence of antimicrobial resistance is continuing to increase. Consequently, efficient approaches to identify sources of resistance are required. This study aimed to compare Escherichia coli isolates from the intestinal tract of camels with isolates from human urinary tract infections (UTIs) in Al Ahsa Province, Kingdom of Saudi Arabia (KSA), for antimicrobial resistance and identification of extended-spectrum β-lactamases (ESBLs). A microbiological study was conducted on 100 samples of cecal contents from camels and 100 urine samples from female UTI patients, to isolate and confirm E. coli using the VITEK 2 Automated System. Sensitivity patterns and identification of ESBLs were analyzed using the antimicrobial susceptibility test. Molecular techniques were used to detect E. coli drug-resistant clones. The presence rate of E. coli in camels was 26.0 % (n = 26/100), and in human samples, the rate of E. coli was 33.0 % (n = 33/100). ESBLs were reported for the first time in KSA, in 26.9 % (n = 5/26) of camel samples and 36.4 % (n = 8/33) of human samples. The multi-drug resistance (MDR) index was 0.13 and 0.17, for camels and humans, respectively. Escherichia coli drug-resistant O25b:H4-sequence type 131(ST131) clone was detected in two camel and two human isolates. This study demonstrates a high presence rate of ESBL-producing E. coli (ESBL-EC) in camels for the first time in KSA. Confirmation of MDR strains and E. coli ST131 clone in human and camel isolates suggests that camels could be a potential reservoir for resistant E. coli strains contributing to the increase in antimicrobial resistance in KSA.

  15. Whole genome sequencing of diverse Shiga toxin-producing and non-producing Escherichia coli strains reveals a variety of virulence and novel antibiotic resistance plasmids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genomes of a diverse set of Shiga toxin-producing E. coli strains and the presence of 38 plasmids among all the isolates were determined. Among the novel plasmids found, there were eight that encoded resistance genes to antibiotics, including aminoglycosides, carbapenems, penicillins, cephalosp...

  16. Ciprofloxacin-resistant, CTX-M-15-producing Escherichia coli ST131 clone in extraintestinal infections in Italy.

    PubMed

    Cerquetti, M; Giufrè, M; García-Fernández, A; Accogli, M; Fortini, D; Luzzi, I; Carattoli, A

    2010-10-01

    Quinolone and β-lactam resistance mechanisms and clonal relationships were characterized among Escherichia coli isolates resistant to ciprofloxacin and extended-spectrum cephalosporins associated with human extra-intestinal infections in Rome. The E. coli. ST131 clone was found to be prevalent. This clone invariably carried a specific pattern of substitutions in the topoisomerase genes and all isolates but one produced CTX-M-15. One ST131 isolate produced SHV-12. The new ST131 variant described here is of particular concern because it combines fluoroquinolone resistance and chromosomally encoded CTX-M-15.

  17. Molecular characterization of integrons in clinical isolates of betalactamase-producing Escherichia coli and Klebsiella pneumoniae in Iran.

    PubMed

    Zeighami, Habib; Haghi, Fakhri; Hajiahmadi, Fahimeh

    2015-06-01

    Integrons are considered to play a significant role in the evolution and spread of antibiotic resistance genes. A total of 349 clinical isolates of Escherichia coli and Klebsiella pneumoniae were investigated for molecular characterization of integrons and betalactamases. Antimicrobial susceptibility testing was also performed as the Clinical and Laboratory Standards Institute (CLSI) guidelines. The frequency of extended spectrum betalactamases (ESBL) or metallo-betalactamases (MBL)-producing isolates, patient demographics, and the susceptibility to various antimicrobial agents were described. BlaCTX-M was the most frequently detected betalactamase in all isolates. Moreover, MBL producing K. pneumoniae carried blaIMP and blaVIM at 100 and 41·6%, respectively but no MBL-positive E. coli was detected. Class 1 integrons were more frequent among E. coli and K. pneumoniae isolates in comparison with class 2 integrons and the frequency of intI2 in K. pneumoniae was significantly higher than E. coli isolates. Five different resistance gene arrays were identified among class 1 integrons. Dihydrofolate reductase (dfrA) and aminoglycoside adenyltransferase (aad) gene cassettes were found to be predominant in the class 1 integrons. These results indicate that class 1 integrons are widespread among ESBL-producing isolates of K. pneumoniae and E. coli and appropriate surveillance and control measures are essential to prevent further dissemination of these elements among Enterobacteriaceae in our country.

  18. Metabolic impact of an NADH-producing glucose-6-phosphate dehydrogenase in Escherichia coli.

    PubMed

    Olavarria, K; De Ingeniis, J; Zielinski, D C; Fuentealba, M; Muñoz, R; McCloskey, D; Feist, A M; Cabrera, R

    2014-12-01

    In Escherichia coli, the oxidative branch of the pentose phosphate pathway (oxPPP) is one of the major sources of NADPH when glucose is the sole carbon nutrient. However, unbalanced NADPH production causes growth impairment as observed in a strain lacking phosphoglucoisomerase (Δpgi). In this work, we studied the metabolic response of this bacterium to the replacement of its glucose-6-phosphate dehydrogenase (G6PDH) by an NADH-producing variant. The homologous enzyme from Leuconostoc mesenteroides was studied by molecular dynamics and site-directed mutagenesis to obtain the NAD-preferring LmG6PDH(R46E,Q47E). Through homologous recombination, the zwf loci (encoding G6PDH) in the chromosomes of WT and Δpgi E. coli strains were replaced by DNA encoding LmG6PDH(R46E,Q47E). Contrary to some predictions performed with flux balance analysis, the replacements caused a substantial effect on the growth rates, increasing 59 % in the Δpgi strain, while falling 44 % in the WT. Quantitative PCR (qPCR) analysis of the zwf locus showed that the expression level of the mutant enzyme was similar to the native enzyme and the expression of genes encoding key enzymes of the central pathways also showed moderate changes among the studied strains. The phenotypic and qPCR data were integrated into in silico modelling, showing an operative G6PDH flux contributing to the NADH pool. Our results indicated that, in vivo, the generation of NADH by G6PDH is beneficial or disadvantageous for growth depending on the operation of the upper Embden-Meyerhof pathway. Interestingly, a genomic database search suggested that in bacteria lacking phosphofructokinase, the G6PDHs tend to have similar preferences for NAD and NADP. The importance of the generation of NADPH in a pathway such as the oxPPP is discussed.

  19. Evaluation of eight agar media for the isolation of shiga toxin-Producing Escherichia coli.

    PubMed

    Gill, Alexander; Huszczynski, George; Gauthier, Martine; Blais, Burton

    2014-01-01

    The growth characteristics of 96 shiga toxin-producing Escherichia coli (STEC) strains representing 36 different O-types (including priority O types O26, O45, O103, O111, O121, O145 and O157) on commercial and in-house agar media were studied. The ability of the strains to grow on agar media with varying selective supplement formulations was evaluated using MacConkey Agar (MAC); Rainbow® Agar O157 (RBA); Rainbow® Agar O157 with manufacturer-recommended selective supplements (RBA-NT); Rainbow® Agar O157 with USDA-recommended selective supplements (RBA-USDA); CHROMagar STEC™ (CH STEC); Tryptone Bile agar containing cefixime and tellurite (TBA-CT); Tryptone Bile agar containing cefixime, tellurite, eosin and methylene blue (TBA-EM); and VTEC agar. All of the strains were able to grow on MAC, RBA and VTEC agar, whereas a number of strains (including some non-O157 priority O types) were unable to grow on the highly selective media CH STEC, RBA-NT, RBA-USDA, TBA-EM and TBA-CT. Only RBA-NT and CH STEC exhibited significant inhibition of background flora from ground beef enrichment. Significant inhibition of background flora from beef trim enrichment was observed with RBA-NT, RBA-USDA, CH STEC, TBA-EM and VTEC agar. With exception of E. coli O157, several different colony morphologies were observed on the differential plating media among strains of the same O type, indicating that this colony morphology is not a reliable means of identifying target STEC. These results suggest that an approach to maximize the recovery of target STEC from beef enrichment cultures is dual plating on lesser (RBA, MAC, VTEC agar) and more highly (RBA-NT, CH STEC) selective agars.

  20. Virulence markers in Shiga toxin-producing Escherichia coli isolated from cattle.

    PubMed Central

    Sandhu, K S; Clarke, R C; Gyles, C L

    1999-01-01

    This study identified potential virulence markers in 93 eae-positive and 179 eae-negative Shiga toxin-producing Escherichia coli (STEC), isolated from a random sampling of healthy cattle in southwestern Ontario. PCR amplification was used to identify genes for enterohemorrhagic E. coli (EHEC)-hemolysin, the EAF plasmid, and bundle-forming pili (Bfp); adherence to HEp-2 cells and to bovine colonocytes, and the fluorescent actin staining (FAS) test were used to characterize interaction of the bacteria with epithelial cells. The EHEC-hemolysin sequences were detected in 98% of eae-positive isolates compared with 34% of eae-negative isolates. All isolates were negative for EAF and bfp sequences. There was 100% correlation between localized adherence (LA) to HEp-2 cells and the FAS test. Forty-eight (52%) of the eae-positive isolates were LA/FAS-positive, whereas none of the 179 eae-negative isolates was positive in either test. Among the eae-negative isolates, 20 (11%) showed diffuse adherence and 5 (2.8%) showed enteroaggregative adherence to HEp-2 cells. Seventy-three percent of the eae-positive isolates adhered to bovine colonocytes, whereas only 26% of 120 eae-negative isolates that were tested adhered. All 13 O157:H7 isolates were positive for eae and EHEC-hemolysin gene sequences, LA/FAS, and adherence to bovine colonocytes. It is concluded that possession of genes for eae and EHEC hemolysin is correlated with the serotype of STEC, that production of EHEC hemolysin was highly correlated with serotypes implicated in human disease, and that none of the potential markers that were examined can be used to predict the potential virulence of an isolate. Images Figure 1. Figure 2. Figure 3. PMID:10480459

  1. Comparable high rates of extended-spectrum-beta-lactamase-producing Escherichia coli in birds of prey from Germany and Mongolia.

    PubMed

    Guenther, Sebastian; Aschenbrenner, Katja; Stamm, Ivonne; Bethe, Astrid; Semmler, Torsten; Stubbe, Annegret; Stubbe, Michael; Batsajkhan, Nyamsuren; Glupczynski, Youri; Wieler, Lothar H; Ewers, Christa

    2012-01-01

    Frequent contact with human waste and liquid manure from intensive livestock breeding, and the increased loads of antibiotic-resistant bacteria that result, are believed to be responsible for the high carriage rates of ESBL-producing E. coli found in birds of prey (raptors) in Central Europe. To test this hypothesis against the influence of avian migration, we initiated a comparative analysis of faecal samples from wild birds found in Saxony-Anhalt in Germany and the Gobi-Desert in Mongolia, regions of dissimilar human and livestock population characteristics and agricultural practices. We sampled a total of 281 wild birds, mostly raptors with primarily north-to-south migration routes. We determined antimicrobial resistance, focusing on ESBL production, and unravelled the phylogenetic and clonal relatedness of identified ESBL-producing E. coli isolates using multi-locus sequence typing (MLST) and macrorestriction analyses. Surprisingly, the overall carriage rates (approximately 5%) and the proportion of ESBL-producers among E. coli (Germany: 13.8%, Mongolia: 10.8%) were similar in both regions. Whereas bla(CTX-M-1) predominated among German isolates (100%), bla(CTX-M-9) was the most prevalent in Mongolian isolates (75%). We identified sequence types (STs) that are well known in human and veterinary clinical ESBL-producing E. coli (ST12, ST117, ST167, ST648) and observed clonal relatedness between a Mongolian avian ESBL-E. coli (ST167) and a clinical isolate of the same ST that originated in a hospitalised patient in Europe. Our data suggest the influence of avian migratory species in the transmission of ESBL-producing E. coli and challenge the prevailing assumption that reducing human influence alone invariably leads to lower rates of antimicrobial resistance.

  2. Comparable high rates of extended-spectrum-beta-lactamase-producing Escherichia coli in birds of prey from Germany and Mongolia.

    PubMed

    Guenther, Sebastian; Aschenbrenner, Katja; Stamm, Ivonne; Bethe, Astrid; Semmler, Torsten; Stubbe, Annegret; Stubbe, Michael; Batsajkhan, Nyamsuren; Glupczynski, Youri; Wieler, Lothar H; Ewers, Christa

    2012-01-01

    Frequent contact with human waste and liquid manure from intensive livestock breeding, and the increased loads of antibiotic-resistant bacteria that result, are believed to be responsible for the high carriage rates of ESBL-producing E. coli found in birds of prey (raptors) in Central Europe. To test this hypothesis against the influence of avian migration, we initiated a comparative analysis of faecal samples from wild birds found in Saxony-Anhalt in Germany and the Gobi-Desert in Mongolia, regions of dissimilar human and livestock population characteristics and agricultural practices. We sampled a total of 281 wild birds, mostly raptors with primarily north-to-south migration routes. We determined antimicrobial resistance, focusing on ESBL production, and unravelled the phylogenetic and clonal relatedness of identified ESBL-producing E. coli isolates using multi-locus sequence typing (MLST) and macrorestriction analyses. Surprisingly, the overall carriage rates (approximately 5%) and the proportion of ESBL-producers among E. coli (Germany: 13.8%, Mongolia: 10.8%) were similar in both regions. Whereas bla(CTX-M-1) predominated among German isolates (100%), bla(CTX-M-9) was the most prevalent in Mongolian isolates (75%). We identified sequence types (STs) that are well known in human and veterinary clinical ESBL-producing E. coli (ST12, ST117, ST167, ST648) and observed clonal relatedness between a Mongolian avian ESBL-E. coli (ST167) and a clinical isolate of the same ST that originated in a hospitalised patient in Europe. Our data suggest the influence of avian migratory species in the transmission of ESBL-producing E. coli and challenge the prevailing assumption that reducing human influence alone invariably leads to lower rates of antimicrobial resistance. PMID:23300857

  3. Comparable High Rates of Extended-Spectrum-Beta-Lactamase-Producing Escherichia coli in Birds of Prey from Germany and Mongolia

    PubMed Central

    Guenther, Sebastian; Aschenbrenner, Katja; Stamm, Ivonne; Bethe, Astrid; Semmler, Torsten; Stubbe, Annegret; Stubbe, Michael; Batsajkhan, Nyamsuren; Glupczynski, Youri; Wieler, Lothar H.; Ewers, Christa

    2012-01-01

    Frequent contact with human waste and liquid manure from intensive livestock breeding, and the increased loads of antibiotic-resistant bacteria that result, are believed to be responsible for the high carriage rates of ESBL-producing E. coli found in birds of prey (raptors) in Central Europe. To test this hypothesis against the influence of avian migration, we initiated a comparative analysis of faecal samples from wild birds found in Saxony-Anhalt in Germany and the Gobi-Desert in Mongolia, regions of dissimilar human and livestock population characteristics and agricultural practices. We sampled a total of 281 wild birds, mostly raptors with primarily north-to-south migration routes. We determined antimicrobial resistance, focusing on ESBL production, and unravelled the phylogenetic and clonal relatedness of identified ESBL-producing E. coli isolates using multi-locus sequence typing (MLST) and macrorestriction analyses. Surprisingly, the overall carriage rates (approximately 5%) and the proportion of ESBL-producers among E. coli (Germany: 13.8%, Mongolia: 10.8%) were similar in both regions. Whereas blaCTX-M-1 predominated among German isolates (100%), blaCTX-M-9 was the most prevalent in Mongolian isolates (75%). We identified sequence types (STs) that are well known in human and veterinary clinical ESBL-producing E. coli (ST12, ST117, ST167, ST648) and observed clonal relatedness between a Mongolian avian ESBL-E. coli (ST167) and a clinical isolate of the same ST that originated in a hospitalised patient in Europe. Our data suggest the influence of avian migratory species in the transmission of ESBL-producing E. coli and challenge the prevailing assumption that reducing human influence alone invariably leads to lower rates of antimicrobial resistance. PMID:23300857

  4. Detection of shiga-toxin producing E. coli (STEC) in leafy greens sold at local retail markets in Alexandria, Egypt.

    PubMed

    Khalil, Rowaida K S; Gomaa, Mohamed A E; Khalil, Mahmoud I M

    2015-03-16

    Leafy green vegetables, a popular and an indispensable ingredient of the daily menus of Egyptians' diets, currently presents a great concern in terms of microbiological hazards. To the best of our knowledge, this is the first report that provides scientific evidence for prevalence of shiga-toxigenic Escherichia coli (STEC) in leafy greens sold at open air local retail markets and superstores in the Egyptian environment. A total of 486 conventional and organic leafy green samples that are eaten raw were collected from different areas in Alexandria, evaluated for total E. coli counts (ECCs), and screened for E. coli O157:H7 using conventional and molecular methods. Recovery of E. coli (≥10(2)CFU/g) from all studied types of leafy greens was indicative of fecal contamination. Total ECCs in conventional samples ranged from 5.47 to 2.56 log CFU/g. Based on their inability to ferment sorbitol on CT-SMAC media, 26 presumptive E. coli O157 isolates were detected in 71.4% (270/378) of the studied conventional samples. From all studied organic samples, only 2 types (organic cabbage and parsley, 16.7%) were contaminated with presumptive E. coli O157. All 28 isolates were further serotyped as E. coli O157 by latex agglutination test, and biochemically confirmed as E. coli. Multiplex PCR assays confirmed the ability of 21.4% (6/28) of the E. coli O157 strains to produce shiga-toxins (Stxs), and their virulence markers were as follows: stx1, 66.6% (4/6); stx2, 50% (3/6); stx1/stx2, 16.7% (1/6); eaeA, 83.3% (5/6); and hlyA, 16.7% (1/6). Only 2 strains recovered from conventional and organic parsley could possibly be classified as E. coli O157:H7 based on the presence of stx-genes (either stx1 or stx2 or both). Results of the present research highlight that high E. coli loads, together with recovery of STEC O157 isolates could pose serious health risks to the produce consumers. This emphasizes the urgent need for health authorities to value and utilize the existing knowledge to

  5. Detection of shiga-toxin producing E. coli (STEC) in leafy greens sold at local retail markets in Alexandria, Egypt.

    PubMed

    Khalil, Rowaida K S; Gomaa, Mohamed A E; Khalil, Mahmoud I M

    2015-03-16

    Leafy green vegetables, a popular and an indispensable ingredient of the daily menus of Egyptians' diets, currently presents a great concern in terms of microbiological hazards. To the best of our knowledge, this is the first report that provides scientific evidence for prevalence of shiga-toxigenic Escherichia coli (STEC) in leafy greens sold at open air local retail markets and superstores in the Egyptian environment. A total of 486 conventional and organic leafy green samples that are eaten raw were collected from different areas in Alexandria, evaluated for total E. coli counts (ECCs), and screened for E. coli O157:H7 using conventional and molecular methods. Recovery of E. coli (≥10(2)CFU/g) from all studied types of leafy greens was indicative of fecal contamination. Total ECCs in conventional samples ranged from 5.47 to 2.56 log CFU/g. Based on their inability to ferment sorbitol on CT-SMAC media, 26 presumptive E. coli O157 isolates were detected in 71.4% (270/378) of the studied conventional samples. From all studied organic samples, only 2 types (organic cabbage and parsley, 16.7%) were contaminated with presumptive E. coli O157. All 28 isolates were further serotyped as E. coli O157 by latex agglutination test, and biochemically confirmed as E. coli. Multiplex PCR assays confirmed the ability of 21.4% (6/28) of the E. coli O157 strains to produce shiga-toxins (Stxs), and their virulence markers were as follows: stx1, 66.6% (4/6); stx2, 50% (3/6); stx1/stx2, 16.7% (1/6); eaeA, 83.3% (5/6); and hlyA, 16.7% (1/6). Only 2 strains recovered from conventional and organic parsley could possibly be classified as E. coli O157:H7 based on the presence of stx-genes (either stx1 or stx2 or both). Results of the present research highlight that high E. coli loads, together with recovery of STEC O157 isolates could pose serious health risks to the produce consumers. This emphasizes the urgent need for health authorities to value and utilize the existing knowledge to

  6. Acid Resistance and molecular characterization of Escherichia coli O157:H7 and different non-O157 Shiga toxin-producing E. coli serogroups

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to compare the acid resistance (AR) of seven non-O157 Shiga toxin-producing E. coli (STEC) strains belonging to serogroups O26, O45, O103, O104, O111, O121 and O145 with O157:H7 STEC isolated from various sources in 400 mM acetic acid solutions (AAS) at pH 3.2 and 30°...

  7. Acid resistance and molecular characterization of Escherichia coli O157:H7 and different Non-O157 shiga toxin-producing E. coli serogroups

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to compare the acid resistance (AR) of non-O157 Shiga toxin-producing Escherichia coli (STEC) strains belonging to serogroups O26, O45, O103, O104, O111, O121, and O145 with O157:H7 STEC isolated from various sources in 400 mM acetic acid solutions (AAS) at pH 3.2 and...

  8. Outbreaks of non-O157 Shiga toxin-producing Escherichia coli infection: USA.

    PubMed

    Luna-Gierke, R E; Griffin, P M; Gould, L H; Herman, K; Bopp, C A; Strockbine, N; Mody, R K

    2014-11-01

    Non-O157 Shiga toxin-producing Escherichia coli (STEC) infections are increasingly detected, but sources are not well established. We summarize outbreaks to 2010 in the USA. Single-aetiology outbreaks were defined as ⩾2 epidemiologically linked culture-confirmed non-O157 STEC infections; multiple-aetiology outbreaks also had laboratory evidence of ⩾2 infections caused by another enteric pathogen. Twenty-six states reported 46 outbreaks with 1727 illnesses and 144 hospitalizations. Of 38 single-aetiology outbreaks, 66% were caused by STEC O111 (n = 14) or O26 (n = 11), and 84% were transmitted through food (n = 17) or person-to-person spread (n = 15); food vehicles included dairy products, produce, and meats; childcare centres were the most common setting for person-to-person spread. Of single-aetiology outbreaks, a greater percentage of persons infected by Shiga toxin 2-positive strains had haemolytic uraemic syndrome compared with persons infected by Shiga toxin 1-only positive strains (7% vs. 0·8%). Compared with single-aetiology outbreaks, multiple-aetiology outbreaks were more frequently transmitted through water or animal contact.

  9. Bactericidal Effect of Selected Antidiarrhoeal Medicinal Plants on Intracellular Heat-Stable Enterotoxin-Producing Escherichia coli.

    PubMed

    Birdi, Tannaz J; Brijesh, S; Daswani, Poonam G

    2014-05-01

    Diarrhoeal diseases due to enterotoxigenic Escherichia coli continue to be a cause of global concern. Medicinal plants have been gaining popularity as promising antidiarrhoeal agents. In the present study, four antidiarrhoeal plants, viz. Aegle marmelos, Cyperus rotundus, Psidium guajava and Zingiber officinale were screened against a heat-stable toxin-producing enterotoxigenic E. coli strain. Decoctions of these plants were studied for their effect on intracellular killing of the bacterial strain using murine monocytic cell line, J774. [(3)H] thymidine release assay was used to evaluate the apoptotic/necrotic effect. All plants at concentrations <1% enhanced intracellular killing of the bacteria by J774 cells. However, at higher concentrations, the decoctions induced apoptosis in J774 cells. The study demonstrates that these plants could control diarrhoea caused by heat-stable toxin-producing enterotoxigenic E. coli through their immunomodulatory effect. PMID:25035535

  10. Verocytotoxin-producing Escherichia coli O26 in raw water buffalo (Bubalus bubalis) milk products in Italy.

    PubMed

    Lorusso, Vanessa; Dambrosio, Angela; Quaglia, Nicoletta Cristiana; Parisi, Antonio; La Salandra, Giovanna; Lucifora, Giuseppe; Mula, Giuseppina; Virgilio, Sebastiano; Carosielli, Leonardo; Rella, Addolorata; Dario, Marco; Normanno, Giovanni

    2009-08-01

    Escherichia coli 026 is known as a verocytotoxin-producing E. coli (VTEC) organism that causes severe foodborne diseases such as hemorrhagic colitis and hemolytic uremic syndrome. Although cattle are the most important reservoir of VTEC, only a few reports on the role of water buffalo (Bubalus bubalis) as a reservoir of VTEC and on the presence of these organisms in their milk are available. However, in Southern Italy, where water buffalo are intensively reared, an outbreak of hemolytic uremic syndrome due to E. coli 026 has recently been reported, in which the consumption of typical dairy products was considered to be a common risk factor. The aims of this work were to assess the prevalence of E. coli O26 in raw water buffalo milk, to characterize the virulence gene profiles of the isolates, and to evaluate their phenotypic antimicrobial resistance pattern. Of 160 analyzed samples, 1 (0.6%) tested positive for E. coli O26, and the isolate showed the stx1+/stx2+/eae-/hlyA+ genotypic profile. The strain showed resistance against glycopeptides, macrolides, and penicillins. The presence of VTEC organisms in raw water buffalo milk could be considered to be a potential threat to consumers; however, the strict adherence to the processes used in the preparation of the most common buffalo dairy products could strongly mitigate the foodborne risk. To our knowledge, this article reports the first isolation and characterization of E. coli O26 VTEC in raw water buffalo milk.

  11. Prevalence and pathogenicity of Shiga toxin-producing Escherichia coli in beef cattle and their products.

    PubMed

    Hussein, H S

    2007-03-01

    During the past 23 yr, a large number of human illness outbreaks have been traced worldwide to consumption of undercooked ground beef and other beef products contaminated with Shiga toxin-producing Escherichia coli (STEC). Although several routes exist for human infection with STEC, beef remains a main source. Thus, beef cattle are considered reservoirs of O157 and nonO157 STEC. Because of the global nature of the food supply, safety concerns with beef will continue, and the challenges facing the beef industry will increase at the production and processing levels. To be prepared to address these concerns and challenges, it is critical to assess the beef cattle role in human infection with STEC. Because most STEC outbreaks in the United States were traced to beef containing E. coli O157:H7, the epidemiological studies have focused on the prevalence of this serotype in beef and beef cattle. Worldwide, however, additional STEC serotypes (e.g., members of the O26, O91, O103, O111, O118, O145, and O166 serogroups) have been isolated from beef and caused human illnesses ranging from bloody diarrhea and hemorrhagic colitis to the life-threatening hemolytic uremic syndrome (HUS). To provide a global assessment of the STEC problem, published reports on beef and beef cattle in the past 3 decades were evaluated. The prevalence rates of E. coli O157 ranged from 0.1 to 54.2% in ground beef, from 0.1 to 4.4% in sausage, from 1.1 to 36.0% in various retail cuts, and from 0.01 to 43.4% in whole carcasses. The corresponding prevalence rates of nonO157 STEC were 2.4 to 30.0%, 17.0 to 49.2%, 11.4 to 49.6%, and 1.7 to 58.0%, respectively. Of the 162 STEC serotypes isolated from beef products, 43 were detected in HUS patients and 36 are known to cause other human illnesses. With regard to beef cattle, the prevalence rates of E. coli O157 ranged from 0.3 to 19.7% in feedlots and from 0.7 to 27.3% on pasture. The corresponding prevalence rates of nonO157 STEC were 4.6 to 55.9% and 4.7 to

  12. Low intestinal colonization of Escherichia coli clone ST131 producing CTX-M-15 in Jordanian infants.

    PubMed

    Badran, E F; Qamer Din, R A; Shehabi, A A

    2016-02-01

    Over a period of 3 years' study (2012-2014), a total of 518 faecal samples were collected and cultured to isolate Escherichia coli. Of these, 338 (65.3%) E. coli isolates were recovered from infants, and 142/338 (42%) were multidrug-resistant (MDR) to ≥ 3 drug classes using the antimicrobial susceptibility disc diffusion method. A total of 125/142 (88%) of E. coli isolates were extended-spectrum β-lactamase (ESBL) producers. blaCTX-M-15 types were observed in 80/125 (64%) of the isolates, and 60/80 (75%) were positive for blaCTX-M-15. Out of 338 E. coli isolates, 9 (2.6%) were positive for ST131/O25b clone and each isolate was associated with several plasmids of different sizes (1-21.2 kb). The identities of these nine isolates were confirmed by sequencing for presence of pabB (347 bp) and trpA (427 bp) genes. This study demonstrates low prevalence rate of the highly virulent E. coli ST131 clone producing blaCTX-M-15 in the intestines of Jordanian infants.

  13. Multidrug-Resistant and Extended Spectrum Beta-Lactamase-Producing Escherichia coli in Dutch Surface Water and Wastewater

    PubMed Central

    Blaak, Hetty; Lynch, Gretta; Italiaander, Ronald; Hamidjaja, Raditijo A.; Schets, Franciska M.; de Roda Husman, Ana Maria

    2015-01-01

    Objective The goal of the current study was to gain insight into the prevalence and concentrations of antimicrobial resistant (AMR) Escherichia coli in Dutch surface water, and to explore the role of wastewater as AMR contamination source. Methods The prevalence of AMR E. coli was determined in 113 surface water samples obtained from 30 different water bodies, and in 33 wastewater samples obtained at five health care institutions (HCIs), seven municipal wastewater treatment plants (mWWTPs), and an airport WWTP. Overall, 846 surface water and 313 wastewater E. coli isolates were analysed with respect to susceptibility to eight antimicrobials (representing seven different classes): ampicillin, cefotaxime, tetracycline, ciprofloxacin, streptomycin, sulfamethoxazole, trimethoprim, and chloramphenicol. Results Among surface water isolates, 26% were resistant to at least one class of antimicrobials, and 11% were multidrug-resistant (MDR). In wastewater, the proportions of AMR/MDR E. coli were 76%/62% at HCIs, 69%/19% at the airport WWTP, and 37%/27% and 31%/20% in mWWTP influents and effluents, respectively. Median concentrations of MDR E. coli were 2.2×102, 4.0×104, 1.8×107, and 4.1×107 cfu/l in surface water, WWTP effluents, WWTP influents and HCI wastewater, respectively. The different resistance types occurred with similar frequencies among E. coli from surface water and E. coli from municipal wastewater. By contrast, among E. coli from HCI wastewater, resistance to cefotaxime and resistance to ciprofloxacin were significantly overrepresented compared to E. coli from municipal wastewater and surface water. Most cefotaxime-resistant E. coliisolates produced ESBL. In two of the mWWTP, ESBL-producing variants were detected that were identical with respect to phylogenetic group, sequence type, AMR-profile, and ESBL-genotype to variants from HCI wastewater discharged onto the same sewer and sampled on the same day (A1/ST23/CTX-M-1, B23/ST131/CTX-M-15, D2/ST405/CTX

  14. Effect of curli expression and hydrophobicity of E. coli O157:H7 on attachment to fresh produce surfaces

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aim: To investigate the effect of curli expression on cell hydrophobicity, biofilm formation, and attachment to cut and intact fresh produce surfaces. Methods and Results: Five E. coli O157:H7 strains were evaluated for curli expression, hydrophobicity, biofilm formation, and attachment of E. co...

  15. Virulence gene profiles of shiga toxin-producing Escherichia coli isolated from fecal samples of finishing swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing Escherichia coli (STEC) are important pathogens responsible for food-borne outbreaks and serious illness including hemorrhagic colitis and hemolytic uremic syndrome. Certain STEC serogroups may cause edema disease in swine; and similar to cattle, swine have been shown to be a ...

  16. Characterization of Shiga toxin-producing Escherichia coli associated with two multi-state foodborne outbreaks in 2006

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In the fall of 2006 two multi-state outbreaks of E. coli serotype O157:H7 infection occurred that involved contaminated spinach and contaminated lettuce. In this study, we compare 7 Shiga toxin-producing isolates associated with those two outbreaks to a collection of food, environmental, and animal ...

  17. Mathematical modeling of growth of non-O157 Shiga Toxin-producing Escherichia coli in raw ground beef

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to investigate the growth of Shiga toxin-producing Escherichia coli (STEC, including serogroups O45, O103, O111, O121, and O145) in raw ground beef and to develop mathematical models to describe the bacterial growth under different temperature conditions. Three prima...

  18. Outbreak of non-O157 Shiga toxin-producing Escherichia coli infection from consumption of beef sausage.

    PubMed

    Ethelberg, Steen; Smith, Birgitte; Torpdahl, Mia; Lisby, Morten; Boel, Jeppe; Jensen, Tenna; Nielsen, Eva Møller; Mølbak, Kåre

    2009-04-15

    We describe an outbreak of Shiga toxin-producing Escherichia coli O26:H11 infection in 20 patients (median age, 2 years). The source of the infection was an organic fermented beef sausage. The source was discovered by using credit card information to obtain and compare customer transaction records from the computer systems of supermarkets. PMID:19272017

  19. Immersion in antimicrobial solutions reduces Salmonella enterica and Shiga toxin-producing Escherichia coli on beef cheek meat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to determine the effect of immersing beef cheek meat in antimicrobial solutions on the reduction of O157:H7 Shiga toxin–producing Escherichia coli (STEC), non-O157:H7 STEC, and Salmonella enterica. Beef cheek meat was inoculated with O157:H7 STEC, non-O157:H7 STEC, an...

  20. KPC-2 producing Klebsiella pneumoniae and Escherichia coli co-infection in a catheter-related infection.

    PubMed

    Leão, R S; Carvalho-Assef, A P D' A; Correal, J C D; Silva, R V; Goldemberg, D C; Asensi, M D; Marques, E A

    2011-03-01

    We describe the first report of simultaneous blood infection with KPC-2 producing Klebsiella pneumoniae and Escherichia coli in a Brazilian patient. We highlight the importance of implementing efficient infection control measures to limit the spread of these phenotypes in a hospital setting.

  1. Inactivation of a diverse set of shiga toxin-producing Escherichia coli in ground beef by high pressure processing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga Toxin-Producing Escherichia coli (STEC) are frequently implicated in foodborne illness outbreaks and recalls of ground beef. In this study we determined the High Pressure Processing (HPP) D-10 value (the processing conditions needed to reduce the microbial population by 1 log) of 39 individua...

  2. Genotypic Characterization of Shiga Toxin-Producing Escherichia coli (STEC) Strains Recovered from Farm Animal Feces in Mexico

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Technical Abstract and Interpretive Summary: Provide electronically in Word. Sixty-three strains of Shiga toxin-producing Escherichia coli (STEC) were recovered from farm animal feces in distinct regions in the Culiacan Valley, an important agricultural region in Mexico for horticultural crops that...

  3. Validation of fermentation and cooking parameters for dry-fermented sausage to control Shiga toxin-producing Escherichia coli (STEC)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga-toxin producing Escherichia coli (STEC) have been occasionally associated with causing illness due to under-processed and/or undercooked raw, further processed, and/or fermented meats. Regarding the latter, the United States Department of Agriculture (USDA) Food Safety and Inspection Service (...

  4. Classification of non-O157 shiga toxin-producing escherichia coli(STEC) serotypes with hyperspectral microscope imaging

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Non-O157 Shiga toxin-producing Escherichia coli (STEC) strains such as O26, O45, O103, O111, O121 and O145 are recognized as serious outbreak to cause human illness due to their toxicity. A conventional microbiological method for cell counting is laborious and needs long time for the results. Since ...

  5. Distribution and detection of Shiga toxin-producing Escherichia coli (STEC) during an industrial grinding process of beef trim

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During the grinding and packaging processes, it is important to understand how Shiga toxin-producing Escherichia coli (STEC) would be distributed and how well it could be detected in beef trim. This study is important because it shows what would happen if contaminated meat is allowed into a commerc...

  6. Molecular characterization of shiga toxin-producing E. coli (STEC) from finishing swine in a longitudinal study

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing E. coli (STEC) infections are a critical public health concern because they can cause severe clinical outcomes, such as hemolytic uremic syndrome, in humans. Determining the presence or absence of virulence genes is essential in assessing the potential pathogenicity of STEC str...

  7. Extended spectrum β-lactamase producing Escherichia coli in broiler breeding roosters: Presence in the reproductive tract and effect on sperm motility.

    PubMed

    Mezhoud, Halima; Boyen, Filip; Touazi, Leg-Hel; Garmyn, An; Moula, Nassim; Smet, Annemieke; Haesbrouck, Freddy; Martel, An; Iguer-Ouada, Mokrane; Touati, Abdelaziz

    2015-08-01

    Extended spectrum β-lactamases (ESBL)-producing Escherichia coli have emerged worldwide in animal husbandry and they were reported from different ecosystems. The purpose of this study was firstly, to investigate the presence of ESBL-producing E. coli in the gastrointestinal (GIT) and reproductive (RT) tracts of broiler breeding roosters, and secondly to study the impact of an ESBL-producing E. coli on artificially infected semen. A total of seventeen ESBL-producing E. coli strains were isolated from the gastrointestinal and reproductive tracts of nine broiler breeding roosters. All isolates were identified to the species level by API 20E system and MALDI-TOF, serotyped, and genetically characterized for ESBL production. Semen was artificially infected with E. coli ATCC25922 or with an ESBL-producing E. coli strain recovered from the reproductive tract. A computer aided semen analyzer (CASA) was used to compare different spermatozoa motility parameters in each sample. All ESBL-producing E. coli isolates could not be typed with the currently used sera and they were harboring a blaCTX-M gene alone or in combination with a blaTEM gene. The semen quality was notably less affected in samples infected with ESBL-producing E. coli strain compared to the control and sample infected with E. coli ATCC25922. The present study revealed that ESBL-producing E. coli can be isolated from both reproductive and digestive tracts of broiler breeding roosters. Contamination of the reproductive tract with ESBL-producing E. coli could lead to contamination of semen and could be an important factor in the dissemination of ESBL-producing E. coli in poultry. PMID:26149221

  8. Inactivation of Shiga toxin-producing Escherichia coli O104:H4 using cold atmospheric pressure plasma.

    PubMed

    Baier, Matthias; Janssen, Traute; Wieler, Lothar H; Ehlbeck, Jörg; Knorr, Dietrich; Schlüter, Oliver

    2015-09-01

    From cultivation to the end of the post-harvest chain, heat-sensitive fresh produce is exposed to a variety of sources of pathogenic microorganisms. If contaminated, effective gentle means of sanitation are necessary to reduce bacterial pathogen load below their infective dose. The occurrence of rare or new serotypes raises the question of their tenacity to inactivation processes. In this study the antibacterial efficiency of cold plasma by an atmospheric pressure plasma-jet was examined against the Shiga toxin-producing outbreak strain Escherichia coli O104:H4. Argon was transformed into non-thermal plasma at a power input of 8 W and a gas flow of 5 L min(-1). Basic tests were performed on polysaccharide gel discs, including the more common E. coli O157:H7 and non-pathogenic E. coli DSM 1116. At 5 mm treatment distance and 10(5) cfu cm(-2) initial bacterial count, plasma reduced E. coli O104:H4 after 60 s by 4.6 ± 0.6 log, E. coli O157:H7 after 45 s by 4.5 ± 0.6 log, and E. coli DSM 1116 after 30 s by 4.4 ± 1.1 log. On the surface of corn salad leaves, gentle plasma application at 17 mm reduced 10(4) cfu cm(-2) of E. coli O104:H4 by 3.3 ± 1.1 log after 2 min, whereas E. coli O157:H7 was inactivated by 3.2 ± 1.1 log after 60 s. In conclusion, plasma treatment has the potential to reduce pathogens such as E. coli O104:H4 on the surface of fresh produce. However, a serotype-specific adaptation of the process parameters is required.

  9. Inactivation of Shiga toxin-producing Escherichia coli O104:H4 using cold atmospheric pressure plasma.

    PubMed

    Baier, Matthias; Janssen, Traute; Wieler, Lothar H; Ehlbeck, Jörg; Knorr, Dietrich; Schlüter, Oliver

    2015-09-01

    From cultivation to the end of the post-harvest chain, heat-sensitive fresh produce is exposed to a variety of sources of pathogenic microorganisms. If contaminated, effective gentle means of sanitation are necessary to reduce bacterial pathogen load below their infective dose. The occurrence of rare or new serotypes raises the question of their tenacity to inactivation processes. In this study the antibacterial efficiency of cold plasma by an atmospheric pressure plasma-jet was examined against the Shiga toxin-producing outbreak strain Escherichia coli O104:H4. Argon was transformed into non-thermal plasma at a power input of 8 W and a gas flow of 5 L min(-1). Basic tests were performed on polysaccharide gel discs, including the more common E. coli O157:H7 and non-pathogenic E. coli DSM 1116. At 5 mm treatment distance and 10(5) cfu cm(-2) initial bacterial count, plasma reduced E. coli O104:H4 after 60 s by 4.6 ± 0.6 log, E. coli O157:H7 after 45 s by 4.5 ± 0.6 log, and E. coli DSM 1116 after 30 s by 4.4 ± 1.1 log. On the surface of corn salad leaves, gentle plasma application at 17 mm reduced 10(4) cfu cm(-2) of E. coli O104:H4 by 3.3 ± 1.1 log after 2 min, whereas E. coli O157:H7 was inactivated by 3.2 ± 1.1 log after 60 s. In conclusion, plasma treatment has the potential to reduce pathogens such as E. coli O104:H4 on the surface of fresh produce. However, a serotype-specific adaptation of the process parameters is required. PMID:25782617

  10. Surveillance of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Dairy Cattle Farms in the Nile Delta, Egypt

    PubMed Central

    Braun, Sascha D.; Ahmed, Marwa F. E.; El-Adawy, Hosny; Hotzel, Helmut; Engelmann, Ines; Weiß, Daniel; Monecke, Stefan; Ehricht, Ralf

    2016-01-01

    Introduction: Industrial livestock farming is a possible source of multi-resistant Gram-negative bacteria, including producers of extended spectrum beta-lactamases (ESBLs) conferring resistance to 3rd generation cephalosporins. Limited information is currently available on the situation of ESBL producers in livestock farming outside of Western Europe. A surveillance study was conducted from January to May in 2014 in four dairy cattle farms in different areas of the Nile delta, Egypt. Materials and Methods: In total, 266 samples were collected from 4 dairy farms including rectal swabs from clinically healthy cattle (n = 210), and environmental samples from the stalls (n = 56). After 24 h pre-enrichment in buffered peptone water, all samples were screened for 3rd generation cephalosporin-resistant Escherichia coli using Brilliance™ ESBL agar. Suspected colonies of putatively ESBL-producing E. coli were sub-cultured and subsequently genotypically and phenotypically characterized. Susceptibility testing using the VITEK-2 system was performed. All suspect isolates were genotypically analyzed using two DNA-microarray based assays: CarbDetect AS-1 and E. coli PanType AS-2 kit (ALERE). These tests allow detection of a multitude of genes and their alleles associated with resistance toward carbapenems, cephalosporins, and other frequently used antibiotics. Serotypes were determined using the E. coli SeroGenotyping AS-1 kit (ALERE). Results: Out of 266 samples tested, 114 (42.8%) ESBL-producing E. coli were geno- and phenotypically identified. 113 of 114 phenotypically 3rd generation cephalosporin-resistant isolates harbored at least one of the ESBL resistance genes covered by the applied assays [blaCTX-M15 (n = 105), blaCTX-M9 (n = 1), blaTEM (n = 90), blaSHV (n = 1)]. Alarmingly, the carbapenemase genes blaOXA-48 (n = 5) and blaOXA-181 (n = 1) were found in isolates that also were phenotypically resistant to imipenem and meropenem. Using the array-based serogenotyping

  11. Development of a faster method for detection of Shiga toxin producing E. coli using Tetrahymena thermophila

    Technology Transfer Automated Retrieval System (TEKTRAN)

    While most STEC outbreaks are caused by E. coli O157, non-O157 STECs are increasingly being implicated. Selective agar for E. coli O157 is commercially available but none detect non-O157 STEC. Currently, regulatory agencies screen for non-O157 STECs by enriching foods overnight, spreading aliquots o...

  12. Persistence of Escherichia coli O157:H7 in major leafy green producing soils

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Persistence of Escherichia coli O157:H7 in 32 (16 organically managed and 16 conventionally managed) soils from California (CA) and Arizona (AZ) was investigated. Results showed that the longest survival (ttd, time needed to reach detection limit, 100 CFU/g dry soil) of E. coli O157:H7 was observed ...

  13. Development of biphasic medium for detection of Shiga toxin producing E. coli using Tetrahymena thermophila

    Technology Transfer Automated Retrieval System (TEKTRAN)

    E. coli O157 has long been the leading cause of major foodborne STEC outbreaks but recently non-O157 STECs are increasingly implicated. Selective media for E. coli O157 are commercially available but none detect non-O157 STEC. Currently, regulatory agencies screen for non-O157 STECs by enriching foo...

  14. Comparative pathogenicity of Escherichia coli O157 and intimin-negative non-O157 Shiga toxin-producing E coli strains in neonatal pigs.

    PubMed

    Dean-Nystrom, Evelyn A; Melton-Celsa, Angela R; Pohlenz, Joachim F L; Moon, Harley W; O'Brien, Alison D

    2003-11-01

    We compared the pathogenicity of intimin-negative non-O157:H7 Shiga toxin (Stx)-producing Escherichia coli (STEC) O91:H21 and O104:H21 strains with the pathogenicity of intimin-positive O157:H7 and O157:H(-) strains in neonatal pigs. We also examined the role of Stx2d-activatable genes and the large hemolysin-encoding plasmid of O91:H21 strain B2F1 in the pathogenesis of STEC disease in pigs. We found that all E. coli strains that made wild-type levels of Stx caused systemic illness and histological lesions in the brain and intestinal crypts, whereas none of the control Stx-negative E. coli strains evoked comparable central nervous system signs or intestinal lesions. By contrast, the absence of intimin, hemolysin, or motility had little impact on the overall pathogenesis of systemic disease during STEC infection. The most striking differences between pigs inoculated with non-O157 STEC strains and pigs inoculated with O157 STEC strains were the absence of attaching and effacing intestinal lesions in pigs inoculated with non-O157:H7 strains and the apparent association between the level of Stx2d-activatable toxin produced by an STEC strain and the severity of lesions. PMID:14573674

  15. Control of Shigatoxin-producing Escherichia coli in cheese by dairy bacterial strains.

    PubMed

    Callon, Cécile; Arliguie, Céline; Montel, Marie-Christine

    2016-02-01

    Bio-preservation could be a valuable way to control Shigatoxin-producing Escherichia coli (STEC) in cheese. To this end, 41 strains were screened for their inhibitory potential on model cheese curd and on pasteurized and raw milk uncooked pressed cheeses. Strains of Lactococcus lactis, Lactococcus garvieae, Leuconostoc pseudomesenteroides, Leuconostoc citreum, Lactobacillus sp, Carnobacterium mobile, Enterococcus faecalis, Enterococcus faecium, Macrococcus caseolyticus and Hafnia alvei reduced STEC O26:H11 counts by 1.4-2.5 log cfu g(-1) and to a lesser extent STEC O157:H7 counts in pasteurized milk cheeses. Some strains can act in synergy to inhibit STEC in raw milk uncooked pressed cheeses. Inhibitory associations had no adverse effect on the sensory characteristics of these cheeses. The association of H. alvei, Lactobacillus plantarum and Lc. lactis was the most inhibitory: after inoculation of this consortium into milk, STEC O26:H11 and O157:H7, inoculated at 2 log cfu ml(-1), were reduced by up to 3 log cfu g(-1) in ripened cheese. Inhibition in cheese cannot be predicted from H2O2 production in BHI medium, decreased pH or milk reduction. It is not clear what role the rapid decrease in pH during the first 6 h may play in the inhibition. Further studies will be needed to determine the nature of the inhibition.

  16. Prevalence of verotoxin-producing Escherichia coli (VTEC) 0157 in Swedish dairy herds.

    PubMed Central

    Eriksson, E.; Aspan, A.; Gunnarsson, A.; Vågsholm, I.

    2005-01-01

    A prevalence study of verotoxin-producing Escherichia coli O157 (VTEC O157) was performed in 371 randomly selected dairy herds distributed throughout Sweden. Faecal and manure samples were collected and analysed by immunomagnetic separation and culturing. Data were recorded for each herd regarding herd size, age of sampled animals and whether, in addition to cattle, the farm kept other animals. VTEC O157 was isolated from 33 (8.9%) of the 371 investigated herds. The prevalence was higher (23.3%) in Halland county than in the rest of Sweden (P > 0.01). Halland was also the county in Sweden that during the study period had the highest incidence of human VTEC O157 cases. VTEC O157 was not detected on any farm in northern Sweden. Identified risk factors, in the multivariate analyses, for herds being VTEC O157 positive were herd size, geographical localization, presence of pigs on the farm and median age of sampled animals. PMID:15816162

  17. Classification of Shiga toxin-producing escherichia coli (STEC) serotypes with hyperspectral microscope imagery

    NASA Astrophysics Data System (ADS)

    Park, Bosoon; Windham, William R.; Ladely, Scott R.; Gurram, Prudhvi; Kwon, Heesung; Yoon, Seung-Chul; Lawrence, Kurt C.; Narang, Neelam; Cray, William C.

    2012-05-01

    Non-O157:H7 Shiga toxin-producing Escherichia coli (STEC) strains such as O26, O45, O103, O111, O121 and O145 are recognized as serious outbreak to cause human illness due to their toxicity. A conventional microbiological method for cell counting is laborious and needs long time for the results. Since optical detection method is promising for realtime, in-situ foodborne pathogen detection, acousto-optical tunable filters (AOTF)-based hyperspectral microscopic imaging (HMI) method has been developed for identifying pathogenic bacteria because of its capability to differentiate both spatial and spectral characteristics of each bacterial cell from microcolony samples. Using the AOTF-based HMI method, 89 contiguous spectral images could be acquired within approximately 30 seconds with 250 ms exposure time. From this study, we have successfully developed the protocol for live-cell immobilization on glass slides to acquire quality spectral images from STEC bacterial cells using the modified dry method. Among the contiguous spectral imagery between 450 and 800 nm, the intensity of spectral images at 458, 498, 522, 546, 570, 586, 670 and 690 nm were distinctive for STEC bacteria. With two different classification algorithms, Support Vector Machine (SVM) and Sparse Kernel-based Ensemble Learning (SKEL), a STEC serotype O45 could be classified with 92% detection accuracy.

  18. Shiga toxin-producing Escherichia coli strains negative for locus of enterocyte effacement.

    PubMed

    Newton, Hayley J; Sloan, Joan; Bulach, Dieter M; Seemann, Torsten; Allison, Cody C; Tauschek, Marija; Robins-Browne, Roy M; Paton, James C; Whittam, Thomas S; Paton, Adrienne W; Hartland, Elizabeth L

    2009-03-01

    Most Shiga toxin-producing Escherichia coli (STEC) infections that are associated with severe sequelae such as hemolytic uremic syndrome (HUS) are caused by attaching and effacing pathogens that carry the locus of enterocyte effacement (LEE). However, a proportion of STEC isolates that do not carry LEE have been associated with HUS. To clarify the emergence of LEE-negative STEC, we compared the genetic composition of the virulence plasmids pO113 and pO157 from LEE-negative and LEE-positive STEC, respectively. The complete nucleotide sequence of pO113 showed that several plasmid genes were shared by STEC O157:H7. In addition, allelic profiling of the ehxA gene demonstrated that pO113 belongs to a different evolutionary lineage than pO157 and that the virulence plasmids of LEE-negative STEC strains were highly related. In contrast, multilocus sequence typing of 17 LEE-negative STEC isolates showed several clonal groups, suggesting that pathogenic LEE-negative STEC has emerged several times throughout its evolution.

  19. Relevance of Biofilms in the Pathogenesis of Shiga-Toxin-Producing Escherichia coli Infection

    PubMed Central

    Angel Villegas, Natalia; Baronetti, José; Albesa, Inés; Polifroni, Rosana; Parma, Alberto; Etcheverría, Analía; Becerra, Maria; Padola, Nora; Paraje, Maria

    2013-01-01

    The present study was designed to determine the relationships among biofilm formation, cellular stress and release of Shiga toxin (Stx) by three different clinical Shiga toxin-producing Escherichia coli (STEC) strains. The biofilm formation was determined using crystal violet stain in tryptic soy broth or thioglycollate medium with the addition of sugars (glucose or mannose) or hydrogen peroxide. The reactive oxygen species (ROSs) were detected by the reduction of nitro blue tetrazolium and reactive nitrogen intermediates (RNI) determined by the Griess assay. In addition, the activities of two antioxidant enzymes, superoxide dismutase (SOD) and catalase (CAT), were studied. For the cytotoxicity studies, Vero cells were cultured with Stx released of STEC biofilms. The addition of sugars in both culture mediums resulted in an increase in biofilm biomass, with a decrease in ROS and RNI production, low levels of SOD and CAT activity, and minimal cytotoxic effects. However, under stressful conditions, an important increase in the antioxidant enzyme activity and high level of Stx production were observed. The disturbance in the prooxidant-antioxidant balance and its effect on the production and release of Stx evaluated under different conditions of biofilm formation may contribute to a better understanding of the relevance of biofilms in the pathogenesis of STEC infection. PMID:24324376

  20. Antibiotic resistance and integrons in Shiga toxin-producing Escherichia coli (STEC).

    PubMed

    Colello, Rocío; Etcheverría, Analía I; Di Conza, Jose A; Gutkind, Gabriel O; Padola, Nora L

    2015-03-01

    Shiga toxin-producing Escherichia coli (STEC) cause hemorrhagic colitis (HC) and hemolytic-uremic syndrome in humans (HUS). Cattle are the main reservoir of STEC and transmission to humans occurs through contaminated food and water. Antibiotics are used in pig production systems to combat disease and improve productivity and play a key role in the dissemination of antibiotic resistance genes to the bacteria. Integrons have been identified in resistant bacteria allowing for the acquisition and dissemination of antibiotic resistance genes. STEC strains isolated from humans and animals have developed antibiotic resistance. In our laboratory, 21 non-157 STEC strains isolated from pigs were analyzed to detect class 1 and 2 integrons by PCR. Eight carried integrons, 7 of them harbored intl2. In another study 545 STEC strains were also analyzed for the presence of intl1 and intl2 . Strains carrying intl1 belonged to isolates from environment (n = 1), chicken hamburger (n = 2), dairy calves (n = 4) and pigs (n = 8). Two strains isolated from pigs harbored intl2 and only one intl1 / intl2 , highlighting the presence of intl2 in pigs. The selection for multiresistant strains may contribute to the emergence of antibiotic resistant pathogens and facilitate the spreading of the mobile resistance elements to other bacteria. PMID:26221083

  1. Use of probiotics to reduce faecal shedding of Shiga toxin-producing Escherichia coli in sheep.

    PubMed

    Rigobelo, E E C; Karapetkov, N; Maestá, S A; Avila, F A; McIntosh, D

    2015-03-01

    Shiga toxin-producing Escherichia coli (STEC) are zoonotic, foodborne pathogens of humans. Ruminants, including sheep, are the primary reservoirs of STEC and there is a need to develop intervention strategies to reduce the entry of STEC into the food chain. The initiation of the majority of bacterial, enteric infections involves colonisation of the gut mucosal surface by the pathogen. However, probiotic bacteria can serve to decrease the severity of infection via a number of mechanisms including competition for receptors and nutrients, and/or the synthesis of organic acids and bacteriocins that create an environment unfavourable for pathogen development. The aim of the current study was to determine whether the administration of a probiotic mixture to sheep experimentally infected with a non-O157 STEC strain, carrying stx1, stx2 and eae genes, was able to decrease faecal shedding of the pathogen. The probiotic mixture contained Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus bulgaricus, Lactobacillus lactis, Streptococcus thermophilus and Enterococcus faecium. The numbers of non-O157 STEC in faecal samples collected from sheep receiving daily doses of the probiotic mixture were significantly lower at the 3rd, 5th and 6th week post-inoculation when compared to the levels recorded in untreated animals. It was concluded that administration of the probiotic mixture reduced faecal shedding of non-O157 STEC in sheep, and holds potential as a pre-harvest intervention method to reduce transmission to humans.

  2. Peri- and Postharvest Factors in the Control of Shiga Toxin-Producing Escherichia coli in Beef.

    PubMed

    Moxley, Rodney A; Acuff, Gary R

    2014-12-01

    Certain Shiga toxin-producing Escherichia coli (STEC) strains are important causes of food-borne disease, with hemorrhagic colitis and, in some cases, hemolytic-uremic syndrome as the clinical manifestations of illness. Six serogroups and one serotype of STEC (O26, O45, O103, O111, O121, O145, and O157:H7) are responsible for the vast majority of cases in the United States. Based on recent data for all food commodities combined, 55.3% and 50.0% of the outbreaks of STEC O157 and non-O157 in the United States, respectively, are attributable to beef as a food source. Consequently, the U.S. Department of Agriculture, Food Safety and Inspection Service declared these organisms as adulterants in raw, nonintact beef. In North America, cattle are a major reservoir of STEC strains, with organisms shed in the feces and contaminated hides of the animals being the main vehicle for spread to carcasses at slaughter. A number of peri- and postharvest interventions targeting STEC have been developed, and significant progress has been made in improving the microbiological quality of beef in the past 20 years as a result. However, continued improvements are needed, and accurate assessment of these interventions, especially for non-O157 STEC, would greatly benefit from improvements in detection methods for these organisms. PMID:26104455

  3. Factors influencing the shedding of verocytotoxin-producing Escherichia coli O157 by beef suckler cows.

    PubMed Central

    Synge, B. A.; Chase-Topping, M. E.; Hopkins, G. F.; McKendrick, I. J.; Thomson-Carter, F.; Gray, D.; Rusbridge, S. M.; Munro, F. I.; Foster, G.; Gunn, G. J.

    2003-01-01

    A study was designed to investigate management factors that might influence the shedding of verocytotoxin-producing Escherichia coli (VTEC) O157 by beef cows in Scotland, where there is a particularly high rate of human infection. Thirty-two herds were visited at least monthly over approximately 1 year for collection of fresh faecal pat samples and information on management factors. The faecal pat samples were tested for VTEC O157 by established culture and immunomagnetic separation methods. Questionnaires were completed at the monthly visits to record management factors. Data were analysed using both univariate and multi-factor (GLMM) analysis. Changes in the number of cows in a group, dogs, wild geese, housing, and the feeding of draff (distillers' grains) were statistically significant as risk factors. The event of calving appeared to reduce the likelihood of shedding. Any effects of weaning or turnout were not statistically significant. It appears that the rate of shedding of VTEC O157 is influenced by several factors but possibly the most important of these are the circumstances of animals being housed, or, when outside, the presence of wild geese. PMID:12729199

  4. Antibiotic resistance and integrons in Shiga toxin-producing Escherichia coli (STEC)

    PubMed Central

    Colello, Rocío; Etcheverría, Analía I.; Conza, Jose A. Di; Gutkind, Gabriel O.; Padola, Nora L.

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) cause hemorrhagic colitis (HC) and hemolytic-uremic syndrome in humans (HUS). Cattle are the main reservoir of STEC and transmission to humans occurs through contaminated food and water. Antibiotics are used in pig production systems to combat disease and improve productivity and play a key role in the dissemination of antibiotic resistance genes to the bacteria. Integrons have been identified in resistant bacteria allowing for the acquisition and dissemination of antibiotic resistance genes. STEC strains isolated from humans and animals have developed antibiotic resistance. In our laboratory, 21 non-157 STEC strains isolated from pigs were analyzed to detect class 1 and 2 integrons by PCR. Eight carried integrons, 7 of them harbored intl2. In another study 545 STEC strains were also analyzed for the presence of intl1 and intl2 . Strains carrying intl1 belonged to isolates from environment (n = 1), chicken hamburger (n = 2), dairy calves (n = 4) and pigs (n = 8). Two strains isolated from pigs harbored intl2 and only one intl1 / intl2 , highlighting the presence of intl2 in pigs. The selection for multiresistant strains may contribute to the emergence of antibiotic resistant pathogens and facilitate the spreading of the mobile resistance elements to other bacteria. PMID:26221083

  5. Antibiotic resistance and integrons in Shiga toxin-producing Escherichia coli (STEC).

    PubMed

    Colello, Rocío; Etcheverría, Analía I; Di Conza, Jose A; Gutkind, Gabriel O; Padola, Nora L

    2015-03-01

    Shiga toxin-producing Escherichia coli (STEC) cause hemorrhagic colitis (HC) and hemolytic-uremic syndrome in humans (HUS). Cattle are the main reservoir of STEC and transmission to humans occurs through contaminated food and water. Antibiotics are used in pig production systems to combat disease and improve productivity and play a key role in the dissemination of antibiotic resistance genes to the bacteria. Integrons have been identified in resistant bacteria allowing for the acquisition and dissemination of antibiotic resistance genes. STEC strains isolated from humans and animals have developed antibiotic resistance. In our laboratory, 21 non-157 STEC strains isolated from pigs were analyzed to detect class 1 and 2 integrons by PCR. Eight carried integrons, 7 of them harbored intl2. In another study 545 STEC strains were also analyzed for the presence of intl1 and intl2 . Strains carrying intl1 belonged to isolates from environment (n = 1), chicken hamburger (n = 2), dairy calves (n = 4) and pigs (n = 8). Two strains isolated from pigs harbored intl2 and only one intl1 / intl2 , highlighting the presence of intl2 in pigs. The selection for multiresistant strains may contribute to the emergence of antibiotic resistant pathogens and facilitate the spreading of the mobile resistance elements to other bacteria.

  6. Survival of Shiga toxin-producing Escherichia coli and Stx bacteriophages in moisture enhanced beef.

    PubMed

    Langsrud, Solveig; Heir, Even; Rode, Tone Mari

    2014-07-01

    Moisture enhancement of meat through injection is a technology to improve the sensory properties and the weight of meat. However, the technology may increase the risk of food borne infections. Shiga toxin-producing Escherichia coli (STEC) or bacteriophages carrying cytotoxin genes (Shiga toxin genes, stx), which is normally only present on the surface of intact beef, may be transferred to the inner parts of the muscle during the injection process. Pathogens and bacteriophages surviving the storage period may not be eliminated in the cooking process since many consumers prefer undercooked beef. Measures to increase the microbial food safety of moisture enhanced beef may include sterilization or washing of the outer surface of the meat before injection, avoiding recycling of marinade and addition of antimicrobial agents to the marinade. This paper reviews the literature regarding microbial safety of moisture enhanced beef with special emphasis on STEC and Stx bacteriophages. Also, results from a European Union research project, ProSafeBeef (Food-CT-16 2006-36241) are presented.

  7. Cytotoxic and Apoptotic Effects of Recombinant Subtilase Cytotoxin Variants of Shiga Toxin-Producing Escherichia coli

    PubMed Central

    Funk, J.; Biber, N.; Schneider, M.; Hauser, E.; Enzenmüller, S.; Förtsch, C.

    2015-01-01

    In this study, the cytotoxicity of the recently described subtilase variant SubAB2-2 of Shiga toxin-producing Escherichia coli was determined and compared to the plasmid-encoded SubAB1 and the chromosome-encoded SubAB2-1 variant. The genes for the respective enzymatic active (A) subunits and binding (B) subunits of the subtilase toxins were amplified and cloned. The recombinant toxin subunits were expressed and purified. Their cytotoxicity on Vero cells was measured for the single A and B subunits, as well as for mixtures of both, to analyze whether hybrids with toxic activity can be identified. The results demonstrated that all three SubAB variants are toxic for Vero cells. However, the values for the 50% cytotoxic dose (CD50) differ for the individual variants. Highest cytotoxicity was shown for SubAB1. Moreover, hybrids of subunits from different subtilase toxins can be obtained which cause substantial cytotoxicity to Vero cells after mixing the A and B subunits prior to application to the cells, which is characteristic for binary toxins. Furthermore, higher concentrations of the enzymatic subunit SubA1 exhibited cytotoxic effects in the absence of the respective B1 subunit. A more detailed investigation in the human HeLa cell line revealed that SubA1 alone induced apoptosis, while the B1 subunit alone did not induce cell death. PMID:25824835

  8. Improved traceability of Shiga-toxin-producing Escherichia coli using CRISPRs for detection and typing.

    PubMed

    Delannoy, Sabine; Beutin, Lothar; Fach, Patrick

    2016-05-01

    Among strains of Shiga-toxin-producing Escherichia coli (STEC), seven serogroups (O26, O45, O103, O111, O121, O145, and O157) are frequently associated with severe clinical illness in humans. The development of methods for their reliable detection from complex samples such as food has been challenging thus far, and is currently based on the PCR detection of the major virulence genes stx1, stx2, and eae, and O-serogroup-specific genes. However, this approach lacks resolution. Moreover, new STEC serotypes are continuously emerging worldwide. For example, in May 2011, strains belonging to the hitherto rarely detected STEC serotype O104:H4 were identified as causative agents of one of the world's largest outbreak of disease with a high incidence of hemorrhagic colitis and hemolytic uremic syndrome in the infected patients. Discriminant typing of pathogens is crucial for epidemiological surveillance and investigations of outbreaks, and especially for tracking and tracing in case of accidental and deliberate contamination of food and water samples. Clustered regularly interspaced short palindromic repeats (CRISPRs) are composed of short, highly conserved DNA repeats separated by unique sequences of similar length. This distinctive sequence signature of CRISPRs can be used for strain typing in several bacterial species including STEC. This review discusses how CRISPRs have recently been used for STEC identification and typing. PMID:26449676

  9. Control of Shigatoxin-producing Escherichia coli in cheese by dairy bacterial strains.

    PubMed

    Callon, Cécile; Arliguie, Céline; Montel, Marie-Christine

    2016-02-01

    Bio-preservation could be a valuable way to control Shigatoxin-producing Escherichia coli (STEC) in cheese. To this end, 41 strains were screened for their inhibitory potential on model cheese curd and on pasteurized and raw milk uncooked pressed cheeses. Strains of Lactococcus lactis, Lactococcus garvieae, Leuconostoc pseudomesenteroides, Leuconostoc citreum, Lactobacillus sp, Carnobacterium mobile, Enterococcus faecalis, Enterococcus faecium, Macrococcus caseolyticus and Hafnia alvei reduced STEC O26:H11 counts by 1.4-2.5 log cfu g(-1) and to a lesser extent STEC O157:H7 counts in pasteurized milk cheeses. Some strains can act in synergy to inhibit STEC in raw milk uncooked pressed cheeses. Inhibitory associations had no adverse effect on the sensory characteristics of these cheeses. The association of H. alvei, Lactobacillus plantarum and Lc. lactis was the most inhibitory: after inoculation of this consortium into milk, STEC O26:H11 and O157:H7, inoculated at 2 log cfu ml(-1), were reduced by up to 3 log cfu g(-1) in ripened cheese. Inhibition in cheese cannot be predicted from H2O2 production in BHI medium, decreased pH or milk reduction. It is not clear what role the rapid decrease in pH during the first 6 h may play in the inhibition. Further studies will be needed to determine the nature of the inhibition. PMID:26678131

  10. Fecal carriage of extended-spectrum β-lactamases and AmpC-producing Escherichia coli in a Libyan community

    PubMed Central

    2014-01-01

    Background Extended-spectrum β-lactamases (ESBLs), including the AmpC type, are important mechanisms of resistance among Enterobacteriaeceae. CTX-M type extended-spectrum β- lactamases, of which there are now over 90 variants, are distributed globally, yet appear to vary in regional distribution. AmpC β-lactamases hydrolyze third generation cephalosporins, but are resistant to inhibition by clavulanate or other β-lactamase inhibitors in vitro. Fecal carriage and rates of colonization by bacteria harboring these resistance mechanisms have been reported in patients with community-acquired infections and in healthy members of their households. Expression of these ESBLs compromises the efficacy of current antibacterial therapies, potentially increasing the seriousness of hospital- and community-acquired Escherichia coli (E. coli) infections. To investigate the occurrence of ESBL-producing E. coli in human fecal flora isolated from two pediatric populations residing in the Libyan cities Zleiten and Abou El Khoms. Isolates were further studied to characterize genes encoding β-lactam resistance, and establish genetic relationships. Methods Antibiotic resistance profiles of phenotypically characterized E. coli isolates recovered from the stools of 243 Libyan children during two surveillance periods in 2001 and 2007 were determined by the disk diffusion method. ESBL-screening was performed using the cephalosporin/clavulanate double synergy disc method, and the AmpC-phenotype was confirmed by the aminophenyl-boronic acid test. ESBL genes were molecularly characterized. Phylogenetic group and multilocus sequence typing (MLST) were determined for ESBL-producing isolates and PFGE was performed to compare banding profiles of some dominant strains. Results ESBLs were identified in 13.4% (18/134) of E. coli isolates, and nine isolates (6.7%) demonstrated AmpC activity; all 18 isolates contained a CTX-M gene. Three CTX-M gene families (CTX-M-1, n = 9; CTX-M-15, n = 8

  11. Occurrence of Verocytotoxin-Producing Escherichia coli O157 on Dutch Dairy Farms

    PubMed Central

    Heuvelink, A. E.; van den Biggelaar, F. L. A. M.; Zwartkruis-Nahuis, J. T. M.; Herbes, R. G.; Huyben, R.; Nagelkerke, N.; Melchers, W. J. G.; Monnens, L. A. H.; de Boer, E.

    1998-01-01

    During the period from September 1996 through November 1996, 10 Dutch dairy farms were visited to collect fecal samples from all cattle present. The samples were examined for the presence of verocytotoxin (VT)-producing Escherichia coli (VTEC) of serogroup O157 (O157 VTEC) by immunomagnetic separation following selective enrichment. Cattle on 7 of the 10 dairy farms tested positive for O157 VTEC, with the proportion of cattle infected varying from 0.8 to 22.4%. On the seven farms positive for O157 VTEC, the excretion rate was highest in calves ages 4 to 12 months (21.2%). In a follow-up study, two O157 VTEC-positive farms and two O157 VTEC-negative farms identified in the prevalence study were revisited five times at intervals of approximately 3 months. Cattle on each farm tested positive at least once. The proportion of cattle infected varied from 0 to 61.0%. Excretion rates peaked in summer and were lowest in winter. Again, the highest prevalence was observed in calves ages 4 to 12 months (11.8%). O157 VTEC strains were also isolated from fecal samples from horses, ponies, and sheep and from milk filters and stable flies. O157 VTEC isolates were characterized by VT production and type, the presence of the E. coli attaching-and-effacing gene, phage type, and pulsed-field gel electrophoretic genotype. No overlapping strain types were identified among isolates from different farms except one. The predominance of a single type at each sampling suggests that horizontal transmission is an important factor in dissemination of O157 VTEC within a farm. The presence of more than one strain type, both simultaneously and over time, suggests that there was more than one source of O157 VTEC on the farms. Furthermore, this study demonstrated that the O157 VTEC status of a farm cannot be ascertained from a single visit testing a small number of cattle. PMID:9817858

  12. Occurrence of verocytotoxin-producing Escherichia coli O157 on Dutch dairy farms.

    PubMed

    Heuvelink, A E; van den Biggelaar, F L; Zwartkruis-Nahuis, J; Herbes, R G; Huyben, R; Nagelkerke, N; Melchers, W J; Monnens, L A; de Boer, E

    1998-12-01

    During the period from September 1996 through November 1996, 10 Dutch dairy farms were visited to collect fecal samples from all cattle present. The samples were examined for the presence of verocytotoxin (VT)-producing Escherichia coli (VTEC) of serogroup O157 (O157 VTEC) by immunomagnetic separation following selective enrichment. Cattle on 7 of the 10 dairy farms tested positive for O157 VTEC, with the proportion of cattle infected varying from 0.8 to 22.4%. On the seven farms positive for O157 VTEC, the excretion rate was highest in calves ages 4 to 12 months (21.2%). In a follow-up study, two O157 VTEC-positive farms and two O157 VTEC-negative farms identified in the prevalence study were revisited five times at intervals of approximately 3 months. Cattle on each farm tested positive at least once. The proportion of cattle infected varied from 0 to 61.0%. Excretion rates peaked in summer and were lowest in winter. Again, the highest prevalence was observed in calves ages 4 to 12 months (11.8%). O157 VTEC strains were also isolated from fecal samples from horses, ponies, and sheep and from milk filters and stable flies. O157 VTEC isolates were characterized by VT production and type, the presence of the E. coli attaching-and-effacing gene, phage type, and pulsed-field gel electrophoretic genotype. No overlapping strain types were identified among isolates from different farms except one. The predominance of a single type at each sampling suggests that horizontal transmission is an important factor in dissemination of O157 VTEC within a farm. The presence of more than one strain type, both simultaneously and over time, suggests that there was more than one source of O157 VTEC on the farms. Furthermore, this study demonstrated that the O157 VTEC status of a farm cannot be ascertained from a single visit testing a small number of cattle. PMID:9817858

  13. Sequential necrotizing fasciitis caused by the monomicrobial pathogens Streptococcus equisimilis and extended-spectrum beta-lactamase-producing Escherichia coli.

    PubMed

    Endo, Akiko; Matsuoka, Ryosuke; Mizuno, Yasushi; Doi, Asako; Nishioka, Hiroaki

    2016-08-01

    Necrotizing fasciitis is a rapidly progressing bacterial infection of the superficial fascia and subcutaneous tissue that is associated with a high mortality rate and is caused by a single species of bacteria or polymicrobial organisms. Escherichia coli is rarely isolated from patients with monomicrobial disease. Further, there are few reports of extended-spectrum beta-lactamase (ESBL)-producing E. coli associated with necrotizing fasciitis. We report here our treatment of an 85-year-old man who was admitted because of necrotizing fasciitis of his right thigh. Streptococcus equisimilis was detected as a monomicrobial pathogen, and the infection was cured by amputation of the patient's right leg and the administration of antibiotics. However, 5 days after discontinuing antibiotic therapy, he developed necrotizing fasciitis on his right upper limb and died. ESBL-producing E. coli was the only bacterial species isolated from blood and skin cultures. This case demonstrates that ESBL-producing E. coli can cause monomicrobial necrotizing fasciitis, particularly during hospitalization and that a different bacterial species can cause disease shortly after a previous episode.

  14. Presence of Multidrug-Resistant Shiga Toxin-Producing Escherichia coli, Enteropathogenic E. coli and Enterotoxigenic E. coli, on Raw Nopalitos (Opuntia ficus-indica L.) and in Nopalitos Salads from Local Retail Markets in Mexico.

    PubMed

    Gómez-Aldapa, Carlos A; Cerna-Cortes, Jorge F; Rangel-Vargas, Esmeralda; Torres-Vitela, Mdel Refugio; Villarruel-López, Angelica; Gutiérrez-Alcántara, Eduardo J; Castro-Rosas, Javier

    2016-05-01

    The presence of multidrug-resistant pathogenic bacteria in food is a significant public health concern. Diarrheagenic Escherichia coli pathotypes (DEPs) are foodborne bacteria. In Mexico, DEPs have been associated with diarrheal illness. There is no information about the presence of multidrug-resistant DEPs on fresh vegetables and in cooked vegetable salads in Mexico. "Nopalitos" (Opuntia ficus-indica L.) is a Cactacea extensively used as a fresh green vegetable throughout Mexico. The presence of generic E. coli and multidrug-resistant DEPs on raw whole and cut nopalitos and in nopalitos salad samples was determined. One hundred raw whole nopalitos (without prickles) samples, 100 raw nopalitos cut into small square samples, and 100 cooked nopalitos salad samples were collected from markets. Generic E. coli was determined using the most probable number procedures. DEPs were identified using two multiplex polymerase chain reaction procedures. Susceptibility to 16 antibiotics was tested for the isolated DEP strains by standard test. Of the 100 whole nopalitos samples, 100 cut nopalitos samples, and 100 nopalitos salad samples, generic E. coli and DEPs were identified, respectively, in 80% and 10%, 74% and 10%, and 64% and 8%. Eighty-two DEP strains were isolated from positive nopalitos samples. The identified DEPs included Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli (EPEC), and enterotoxigenic E. coli (ETEC). All isolated strains exhibited resistance to at least six antibiotics. To the best of our knowledge, this is the first report of the presence of multidrug-resistant and antibiotic resistance profiles of STEC, ETEC, and EPEC on raw nopalitos and in nopalitos salads in Mexico.

  15. HlyF Produced by Extraintestinal Pathogenic Escherichia coli Is a Virulence Factor That Regulates Outer Membrane Vesicle Biogenesis.

    PubMed

    Murase, Kazunori; Martin, Patricia; Porcheron, Gaëlle; Houle, Sébastien; Helloin, Emmanuelle; Pénary, Marie; Nougayrède, Jean-Philippe; Dozois, Charles M; Hayashi, Tetsuya; Oswald, Eric

    2016-03-01

    Escherichia coli can cause extraintestinal infections in humans and animals. The hlyF gene is epidemiologically associated with virulent strains of avian pathogenic E. coli and human neonatal meningitis-associated E. coli. We demonstrated that culture supernatants of E. coli expressing HlyF induced autophagy in eukaryotic cells. This phenotype coincided with an enhanced production of outer membrane vesicles (OMVs) by bacteria expressing HlyF. The HlyF protein displays a predicted catalytic domain of the short-chain dehydrogenase/reductase superfamily. This conserved domain was involved the ability of HlyF to promote the production of OMVs. The increased production of OMVs was associated with the release of toxins. hlyF was shown to be expressed during extraintestinal infection and to play a role in the virulence of extraintestinal pathogenic E. coli in a chicken model of colibacillosis. This is the first evidence that pathogenic bacteria produce a virulence factor directly involved in the production of OMVs.

  16. Escherichia coli Common Pilus (ECP) Targets Arabinosyl Residues in Plant Cell Walls to Mediate Adhesion to Fresh Produce Plants*

    PubMed Central

    Rossez, Yannick; Holmes, Ashleigh; Lodberg-Pedersen, Henriette; Birse, Louise; Marshall, Jacqueline; Willats, William G. T.; Toth, Ian K.; Holden, Nicola J.

    2014-01-01

    Outbreaks of verotoxigenic Escherichia coli are often associated with fresh produce. However, the molecular basis to adherence is unknown beyond ionic lipid-flagellum interactions in plant cell membranes. We demonstrate that arabinans present in different constituents of plant cell walls are targeted for adherence by E. coli common pilus (ECP; or meningitis-associated and temperature-regulated (Mat) fimbriae) for E. coli serotypes O157:H7 and O18:K1:H7. l-Arabinose is a common constituent of plant cell wall that is rarely found in other organisms, whereas ECP is widespread in E. coli and other environmental enteric species. ECP bound to oligosaccharides of at least arabinotriose or longer in a glycan array, plant cell wall pectic polysaccharides, and plant glycoproteins. Recognition overlapped with the antibody LM13, which binds arabinanase-sensitive pectic epitopes, and showed a preferential affinity for (1→5)-α-linked l-arabinosyl residues and longer chains of arabinan as demonstrated with the use of arabinan-degrading enzymes. Functional adherence in planta was mediated by the adhesin EcpD in combination with the structural subunit, EcpA, and expression was demonstrated with an ecpR–GFP fusion and ECP antibodies. Spinach was found to be enriched for ECP/LM13 targets compared with lettuce. Specific recognition of arabinosyl residues may help explain the persistence of E. coli in the wider environment and association of verotoxigenic E. coli with some fresh produce plants by exploitation of a glycan found only in plant, not animal, cells. PMID:25320086

  17. Escherichia coli common pilus (ECP) targets arabinosyl residues in plant cell walls to mediate adhesion to fresh produce plants.

    PubMed

    Rossez, Yannick; Holmes, Ashleigh; Lodberg-Pedersen, Henriette; Birse, Louise; Marshall, Jacqueline; Willats, William G T; Toth, Ian K; Holden, Nicola J

    2014-12-01

    Outbreaks of verotoxigenic Escherichia coli are often associated with fresh produce. However, the molecular basis to adherence is unknown beyond ionic lipid-flagellum interactions in plant cell membranes. We demonstrate that arabinans present in different constituents of plant cell walls are targeted for adherence by E. coli common pilus (ECP; or meningitis-associated and temperature-regulated (Mat) fimbriae) for E. coli serotypes O157:H7 and O18:K1:H7. l-Arabinose is a common constituent of plant cell wall that is rarely found in other organisms, whereas ECP is widespread in E. coli and other environmental enteric species. ECP bound to oligosaccharides of at least arabinotriose or longer in a glycan array, plant cell wall pectic polysaccharides, and plant glycoproteins. Recognition overlapped with the antibody LM13, which binds arabinanase-sensitive pectic epitopes, and showed a preferential affinity for (1→5)-α-linked l-arabinosyl residues and longer chains of arabinan as demonstrated with the use of arabinan-degrading enzymes. Functional adherence in planta was mediated by the adhesin EcpD in combination with the structural subunit, EcpA, and expression was demonstrated with an ecpR-GFP fusion and ECP antibodies. Spinach was found to be enriched for ECP/LM13 targets compared with lettuce. Specific recognition of arabinosyl residues may help explain the persistence of E. coli in the wider environment and association of verotoxigenic E. coli with some fresh produce plants by exploitation of a glycan found only in plant, not animal, cells.

  18. Dietary choice affects Shiga toxin-producing Escherichia coli (STEC) O157:H7 colonization and disease.

    PubMed

    Zumbrun, Steven D; Melton-Celsa, Angela R; Smith, Mark A; Gilbreath, Jeremy J; Merrell, D Scott; O'Brien, Alison D

    2013-06-01

    The likelihood that a single individual infected with the Shiga toxin (Stx)-producing, food-borne pathogen Escherichia coli O157:H7 will develop a life-threatening sequela called the hemolytic uremic syndrome is unpredictable. We reasoned that conditions that enhance Stx binding and uptake within the gut after E. coli O157:H7 infection should result in greater disease severity. Because the receptor for Stx, globotriaosylceramide, is up-regulated in the presence of butyrate in vitro, we asked whether a high fiber diet (HFD) that reportedly enhances butyrate production by normal gut flora can influence the outcome of an E. coli O157 infection in mice. To address that question, groups of BALB/c mice were fed high (10%) or low (2%) fiber diets and infected with E. coli O157:H7 strain 86-24 (Stx2+). Mice fed an HFD exhibited a 10- to 100-fold increase in colonization, lost 15% more body weight, exhibited signs of morbidity, and had 25% greater mortality relative to the low fiber diet (LFD)-fed group. Additionally, sections of intestinal tissue from HFD-fed mice bound more Stx1 and expressed more globotriaosylceramide than did such sections from LFD-fed mice. Furthermore, the gut microbiota of HFD-fed mice compared with LFD-fed mice contained reduced levels of native Escherichia species, organisms that might protect the gut from colonization by incoming E. coli O157:H7. Taken together, these results suggest that susceptibility to infection and subsequent disease after ingestion of E. coli O157:H7 may depend, at least in part, on individual diet and/or the capacity of the commensal flora to produce butyrate.

  19. Current status of extended spectrum β-lactamase-producing Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis in Okinawa prefecture, Japan.

    PubMed

    Nakama, Rika; Shingaki, Aoi; Miyazato, Hiroko; Higa, Rikako; Nagamoto, Chota; Hamamoto, Kouta; Ueda, Shuhei; Hachiman, Teruyuki; Touma, Yuki; Miyagi, Kazufumi; Kawahara, Ryuji; Toyosato, Takehiko; Hirai, Itaru

    2016-05-01

    Enterobacteriaceae producing extended spectrum β-lactamase (ESBL) are distributed worldwide. In this study, 114 ESBL-producing Enterobacteriaceae were isolated by analyzing 1672 clinical isolates of Enterobacteriaceae collected from an Okinawa prefectural hospital in Japan between June 2013 and July 2014. The overall prevalence of ESBL-producing Enterobacteriaceae was 6.8%; the prevalence of different bacterial species among the ESBL-producing isolates was as follows: 11.5% Escherichia coli (90 of 783 isolates), 6.2% Klebsiella pneumoniae (19 of 307 isolates), and 11.1% Proteus mirabilis (5 of 45 isolates). The ESBL types blaCTX-M-1, -3, -15, -2, -14, -27, and mutants of blaSHV-1 were detected. Among them, blaCTX-M-15 (33.3%), blaCTX-M-14 (27.8%) and blaCTX-M-27 (33.3%) were dominant in the E. coli isolates, whereas a blaSHV mutant which possessed four mutations (Tyr7Phe, Leu35Gln, Gly238Ser and Glu240Lys) in the amino acid sequence of SHV-1 dominated in the K. pneumoniae isolates (11 of 19, 57.9%). The pandemic E. coli ST131 clone was found to constitute 3.3% of the overall examined isolates and 62.2% of the ESBL-producing E. coli isolates. Our results suggest that the genetic combination of blaCTX-M, and blaSHV and antibiotics-resistant profile were different from that in other regions such as other areas of Japan, Asia, Europe, and North America, especially in the ESBL-producing K. pneumoniae isolates and in the E. coli B2-O25b-ST131 isolates possessing blaCTX-M-15 (40.7% of the E. coli B2-O25b-ST131 isolates). Taken together, our results indicate that the ESBL-producing Enterobacteriaceae in Okinawa, Japan, might be of a unique nature. PMID:26898665

  20. Current status of extended spectrum β-lactamase-producing Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis in Okinawa prefecture, Japan.

    PubMed

    Nakama, Rika; Shingaki, Aoi; Miyazato, Hiroko; Higa, Rikako; Nagamoto, Chota; Hamamoto, Kouta; Ueda, Shuhei; Hachiman, Teruyuki; Touma, Yuki; Miyagi, Kazufumi; Kawahara, Ryuji; Toyosato, Takehiko; Hirai, Itaru

    2016-05-01

    Enterobacteriaceae producing extended spectrum β-lactamase (ESBL) are distributed worldwide. In this study, 114 ESBL-producing Enterobacteriaceae were isolated by analyzing 1672 clinical isolates of Enterobacteriaceae collected from an Okinawa prefectural hospital in Japan between June 2013 and July 2014. The overall prevalence of ESBL-producing Enterobacteriaceae was 6.8%; the prevalence of different bacterial species among the ESBL-producing isolates was as follows: 11.5% Escherichia coli (90 of 783 isolates), 6.2% Klebsiella pneumoniae (19 of 307 isolates), and 11.1% Proteus mirabilis (5 of 45 isolates). The ESBL types blaCTX-M-1, -3, -15, -2, -14, -27, and mutants of blaSHV-1 were detected. Among them, blaCTX-M-15 (33.3%), blaCTX-M-14 (27.8%) and blaCTX-M-27 (33.3%) were dominant in the E. coli isolates, whereas a blaSHV mutant which possessed four mutations (Tyr7Phe, Leu35Gln, Gly238Ser and Glu240Lys) in the amino acid sequence of SHV-1 dominated in the K. pneumoniae isolates (11 of 19, 57.9%). The pandemic E. coli ST131 clone was found to constitute 3.3% of the overall examined isolates and 62.2% of the ESBL-producing E. coli isolates. Our results suggest that the genetic combination of blaCTX-M, and blaSHV and antibiotics-resistant profile were different from that in other regions such as other areas of Japan, Asia, Europe, and North America, especially in the ESBL-producing K. pneumoniae isolates and in the E. coli B2-O25b-ST131 isolates possessing blaCTX-M-15 (40.7% of the E. coli B2-O25b-ST131 isolates). Taken together, our results indicate that the ESBL-producing Enterobacteriaceae in Okinawa, Japan, might be of a unique nature.

  1. Genetic evolution and clinical impact in extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella pneumoniae.

    PubMed

    Chong, Yong; Ito, Yoshikiyo; Kamimura, Tomohiko

    2011-10-01

    The emergence of extended-spectrum β-lactamase (ESBL)-producing bacteria, particularly Escherichia coli and Klebsiella pneumoniae, is now a critical concern for the development of therapies against bacterial infection. ESBLs consist of three major genetic groups: TEM, SHV, and CTX-M types. Nosocomial infections due to TEM and SHV-producing K. pneumoniae strains were frequently documented until the late 1990s. The number of reports on community-acquired infections caused by CTX-M-producing E. coli strains have dramatically increased over the last decade; however, K. pneumoniae strains, of either the TEM or SHV types, are persistent and important ESBL producers. The spread of ESBL genes is associated with various mobile genetic elements, such as transposons, insertion sequences, and integrons. The rapid dissemination of ESBL genes of the CTX-M type may be related to highly complicated genetic structures. These structures harboring ESBL genes and mobile elements are found in a variety of plasmids, which often carry many other antibiotic resistance genes. Multidrug-resistant CTX-M-15-producing E. coli strains disseminate worldwide. Efficient mobile elements and plasmids may have accelerated the genetic diversity and the rapid spread of ESBL genes, and their genetic evolution has caused an emerging threat to the bacteria for which few effective drugs have been identified.

  2. Screening and identification of genetic loci involved in producing more/denser inclusion bodies in Escherichia coli

    PubMed Central

    2013-01-01

    Background Many proteins and peptides have been used in therapeutic or industrial applications. They are often produced in microbial production hosts by fermentation. Robust protein production in the hosts and efficient downstream purification are two critical factors that could significantly reduce cost for microbial protein production by fermentation. Producing proteins/peptides as inclusion bodies in the hosts has the potential to achieve both high titers in fermentation and cost-effective downstream purification. Manipulation of the host cells such as overexpression/deletion of certain genes could lead to producing more and/or denser inclusion bodies. However, there are limited screening methods to help to identify beneficial genetic changes rendering more protein production and/or denser inclusion bodies. Results We report development and optimization of a simple density gradient method that can be used for distinguishing and sorting E. coli cells with different buoyant densities. We demonstrate utilization of the method to screen genetic libraries to identify a) expression of glyQS loci on plasmid that increased expression of a peptide of interest as well as the buoyant density of inclusion body producing E. coli cells; and b) deletion of a host gltA gene that increased the buoyant density of the inclusion body produced in the E. coli cells. Conclusion A novel density gradient sorting method was developed to screen genetic libraries. Beneficial host genetic changes could be exploited to improve recombinant protein expression as well as downstream protein purification. PMID:23638724

  3. Prevalence of ESBL producing Escherichia Coli and Klebsiella species with their co-resistance pattern to antimicrobials.

    PubMed

    Biswas, T; Das, M; Mondal, R; Raj, H J; Mondal, S

    2013-04-01

    Extended spectrum β-lactamase producing bacteria are potential emerging pathogens and continue to be a major challenge in clinical setup worldwide. In the present study an attempt was made to study the prevalence of extended spectrum β-lactamase producing Escherichia coli and Klebsiella species from clinical isolates in a rural tertiary care hospital in West Bengal, India with their antimicrobial susceptibility as well as co-resistance pattern to different antimicrobials. A total of 179 Escherichia coli and 62 Klebsiella isolates recovered from various clinical samples of urine, pus, aural swabs and respiratory secretions (including sputum) for a period of six months were subjected to routine antimicrobial susceptibility testing and also tested for extended spectrum β-lactamase production as per NCCLS recommendations. Extended spectrum β-lactamase was detected in 32.40% of Escherichia coli and 40.32% of Klebsiella species isolates. Urine, pus and respiratory samples were common source of extended spectrum β-lactamase producers and resistance rate of these organisms to third generation cephalosporins were more than 30 to 40%. Co-resistance pattern of these extended spectrum β-lactamase producers to other commonly used antimicrobials were also statistically significant (p≤0.05). From the study it is concluded that indiscriminate use of third generation cephalosporins may be responsible for the selection of extended spectrum β-lactamase producing multidrug resistant strains in hospital setup and amikacin is a reliable drug against them.

  4. Phylogenetic groups and cephalosporin resistance genes of Escherichia coli from diseased food-producing animals in Japan

    PubMed Central

    2011-01-01

    A total of 318 Escherichia coli isolates obtained from different food-producing animals affected with colibacillosis between 2001 and 2006 were subjected to phylogenetic analysis: 72 bovine isolates, 89 poultry isolates and 157 porcine isolates. Overall, the phylogenetic group A was predominant in isolates from cattle (36/72, 50%) and pigs (101/157, 64.3%) whereas groups A (44/89, 49.4%) and D (40/89, 44.9%) were predominant in isolates from poultry. In addition, group B2 was not found among diseased food-producing animals except for a poultry isolate. Thus, the phylogenetic group distribution of E. coli from diseased animals was different by animal species. Among the 318 isolates, cefazolin resistance (minimum inhibitory concentrations: ≥32 μg/ml) was found in six bovine isolates, 29 poultry isolates and three porcine isolates. Of them, 11 isolates (nine from poultry and two from cattle) produced extended spectrum β-lactamase (ESBL). The two bovine isolates produced blaCTX-M-2, while the nine poultry isolates produced blaCTX-M-25 (4), blaSHV-2 (3), blaCTX-M-15 (1) and blaCTX-M-2 (1). Thus, our results showed that several types of ESBL were identified and three types of β-lactamase (SHV-2, CTX-M-25 and CTX-M-15) were observed for the first time in E. coli from diseased animals in Japan. PMID:21989155

  5. First Characterization of CTX-M-15-Producing Escherichia coli ST131 and ST405 Clones Causing Community-Onset Infections in South America▿

    PubMed Central

    Ruiz, Sory J.; Montealegre, Maria Camila; Ruiz-Garbajosa, Patricia; Correa, Adriana; Briceño, David F.; Martinez, Ernesto; Rosso, Fernando; Muñoz, Martin; Quinn, John P.; Cantón, Rafael; Villegas, Maria Virginia

    2011-01-01

    CTX-M-15-producing Escherichia coli has emerged worldwide as an important pathogen associated with community-onset infections, but in South America reports are scarce. We document the presence of CTX-M-15-producing E. coli of the international ST131 and ST405 clones in Colombia and present the first molecular characterization of these isolates in South America. PMID:21325548

  6. O-antigen and virulence profiling of Shiga toxin-producing Escherichia coli by a rapid and cost-effective DNA microarray colorimetric method

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing Escherichia coli (STEC) is a leading cause of foodborne illness worldwide. To evaluate better methods to rapidly detect and genotype Shiga toxin-producing Escherichia coli strains, the present study evaluated the use of the ampliPHOX colorimetric detection technology, based on ...

  7. Effect of ceftriaxone on the outcome of murine pyelonephritis caused by extended-spectrum-β-lactamase-producing Escherichia coli.

    PubMed

    Tratselas, A; Simitsopoulou, M; Giannakopoulou, A; Dori, I; Saoulidis, S; Kollios, K; Papaioannidou, P; Pournaras, S; Roilides, E

    2014-12-01

    Urinary tract infections (UTIs) due to extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae in children are becoming more frequent, and they are commonly treated initially with a second- or third-generation cephalosporin. We developed a murine model of ascending UTI caused by ESBL-producing Escherichia coli. Using this model, we investigated the renal bacterial burden, interleukin-6 (IL-6) expression, and histopathological alterations caused by ESBL- and non-ESBL-producing bacteria after 1, 2, or 6 days with or without ceftriaxone therapy. The renal bacterial burden, IL-6 concentration, and histological inflammatory lesions were not significantly different between mice infected with ESBL- and non-ESBL-producing bacteria without treatment at any of the time points examined. Following ceftriaxone administration, the bacterial burden was eliminated in the kidneys of mice infected with ESBL- and non-ESBL-producing bacteria on the 6th postinfection day. The histological analysis demonstrated that among mice treated with ceftriaxone, those infected with ESBL-producing bacteria had more profound renal alterations than those infected with non-ESBL-producing bacteria on the 6th day (P < 0.001). In comparison, microbiological outcomes did not differ significantly between mice infected with ESBL- and non-ESBL-producing bacteria at any of the time points examined. The effectiveness of ceftriaxone in mice with UTIs due to ESBL-producing E. coli may have therapeutic implications; it is, however, hampered by limited activity on the histopathological lesions, a finding that needs further investigation.

  8. Hygiene quality and presence of ESBL-producing Escherichia coli in raw food diets for dogs

    PubMed Central

    Nilsson, Oskar

    2015-01-01

    Background Raw food diets are popular among some dog owners, even though there are concerns regarding the infectious disease risk and public health implications. Hence, the two aims of this study were to investigate the hygiene quality of raw food diets for dogs in the Swedish market and if Escherichia coli with transferable resistance to extended spectrum cephalosporins (ESC) was present in such products. Methods Samples of raw food diets were suspended and further diluted in 0.9% saline. Appropriate dilutions were 1) cultured on Petrifilm™SEC to quantify the amount of E. coli in the samples and 2) mixed with cefotaxime to a final concentration of 1 mg/L and cultured on Petrifilm™SEC to quantify the amount of ESC-resistant E. coli in the samples. Furthermore, undiluted suspensions were mixed 1:1 with double strength MacConkey broth with cefotaxime, enriched overnight and finally cultured on MacConkey agar with cefotaxime (1 mg/L). Suspected ESC-resistant E. coli were screened by PCR for genes encoding extended spectrum beta lactamases and plasmid-mediated AmpC and their susceptibility to a panel of antimicrobials was performed by broth microdilution using VetMIC GN-mo. Results Escherichia coli was isolated from all samples (n=39) and ESC-resistant E. coli was isolated from nine samples (23%). All ESC-resistant E. coli were PCR-positive for the bla CMY-2 group and only one of them was also resistant to a non-beta-lactam antibiotic. Conclusion The results of this study indicate that raw food diets could be a source of ESC-resistant E. coli to dogs and highlight the need for maintaining good hygiene when handling these products to prevent infection. PMID:26490763

  9. The Serine Protease Motif of EspC from Enteropathogenic Escherichia coli Produces Epithelial Damage by a Mechanism Different from That of Pet Toxin from Enteroaggregative E. coli

    PubMed Central

    Navarro-García, Fernando; Canizalez-Roman, Adrián; Sui, Bao Quan; Nataro, James P.; Azamar, Yenia

    2004-01-01

    EspC (Escherichia coli secreted protein C) of enteropathogenic E. coli (EPEC) shows the three classical domains of the autotransporter proteins and has a conserved serine protease motif belonging to the SPATE (serine protease autotransporters of Enterobacteriaceae) subfamily. EspC and its homolog Pet in enteroaggregative E. coli (EAEC) bear the same sequence within the serine protease motif, and both proteins produce enterotoxic effects, suggesting that like Pet, EspC could be internalized to reach and cleave the calmodulin-binding domain of fodrin, causing actin cytoskeleton disruption. Even though both proteins cause cytoskeleton damage by virtue of their serine protease motifs, the following evidence supports the hypothesis that the mechanisms are different. (i) To obtain similar cytotoxic and cytoskeletal effects, a threefold-higher EspC concentration and a twofold-higher exposure time are needed. (ii) EspC internalization into epithelial cells takes more time (6 h) than Pet internalization (30 min), and the distributions of the two proteins inside the cells are also different. (iii) Both proteins have affinity for fodrin and cleave it, but the cleavage sites are different; EspC produces two cleavages, while Pet produces just one. (iv) EspC does not cause fodrin redistribution within epithelial cells. (v) An EspC serine protease motif mutant, but not a Pet serine protease mutant, competes with EspC by blocking cytoskeletal damage. All these data suggest that the protein conformational structure is very important for the activity of the catalytic site, influencing its interaction with the target protein and its internalization. The differences between these proteins may explain the reduced ability of EspC to cause cytopathic effects. However, these differences may confer a specialized role on EspC in the pathogenesis of EPEC, which is different from that of Pet in EAEC pathogenesis. PMID:15155671

  10. [Cephalosporin-Acid Synthetase of Escherichia coli Strain VKPM B-10182: Genomic Context, Gene Identification, Producer Strain Production].

    PubMed

    Eldarov M, A; Sklyarenko, A V; Mardanov, A V; Beletsky, A V; Zhgun, A A; Dumina, M V; Medvedeva, N V; Satarova, D E; Ravin, N V; Yarockii, S V

    2015-01-01

    An enzyme of cephalosporin-acid synthetase produced by the E. coli strain VKPM B-10182 has specificity for the synthesis of β-lactam antibiotics of the cephalosporin acids class (cefazolin, cefalotin, cefezole etc.). A comparison of the previously determined genomic sequence of E. coli VKPM B-10182 with a genome of the parent E. coli strain ATCC 9637 was performed. Multiple mutations indicating the long selection history of the strain were detected, including mutations in the genes of RNase and β-lactamases that could enhance the level of enzyme synthesis and reduce the degree of degradation of the synthesized cephalosporin acids. The CASA gene--a direct homolog of the penicillin G-acylase gene--was identified by bioinformatics methods. The homology of the gene was confirmed by gene cloning and the expression and determination of its enzymatic activity in the reaction of cefazolin synthesis. The CASA gene was isolated and cloned into the original expression vector, resulting in an effective E. coli BL2l(DE3) pMD0107 strain producing CASA. PMID:26596082

  11. Conventional curing practices reduce generic Escherichia coli and Salmonella spp. on dry bulb onions produced with contaminated irrigation water.

    PubMed

    Emch, Alexander W; Waite-Cusic, Joy G

    2016-02-01

    Food Safety Modernization Act (FSMA) has emphasized microbial risks associated with irrigation water. Treasure Valley (eastern Oregon/western Idaho) has the highest yield of dry bulb onions in the country; however, their irrigation water is often non-compliant with current industry and proposed federal standards for fresh produce. Conventional curing practices may provide a mechanism to mitigate irrigation water quality to comply with FSMA regulations. Dry bulb onions were grown in Owyhee silt loam and Semiahmoo muck soils in greenhouses and irrigated with water containing a cocktail of rifampicin-resistant generic Escherichia coli and Salmonella spp. (4.80 log CFU/ml). To mimic conventional practices, mature onions remained undisturbed in soil without irrigation for 12 days prior to being lifted and cured for 16 additional days. Surviving generic E. coli and Salmonella spp. were selectively enumerated on using standard plating (Hektoen Enteric Agar with rifampicin; HE + rif) or most probable number (lactose broth with rifampicin; HE + rif) methods. Generic E. coli and Salmonella spp. on onions decreased 0.19-0.26 log CFU/g·d during the initial 12 days of finishing. At lifting, generic E. coli and Salmonella spp. had been reduced to <1 CFU/g and persisted through the end of curing. This study demonstrates conventional curing practices as an effective mitigation strategy for dry bulb onions produced with water of poor microbiological quality. PMID:26678128

  12. Conventional curing practices reduce generic Escherichia coli and Salmonella spp. on dry bulb onions produced with contaminated irrigation water.

    PubMed

    Emch, Alexander W; Waite-Cusic, Joy G

    2016-02-01

    Food Safety Modernization Act (FSMA) has emphasized microbial risks associated with irrigation water. Treasure Valley (eastern Oregon/western Idaho) has the highest yield of dry bulb onions in the country; however, their irrigation water is often non-compliant with current industry and proposed federal standards for fresh produce. Conventional curing practices may provide a mechanism to mitigate irrigation water quality to comply with FSMA regulations. Dry bulb onions were grown in Owyhee silt loam and Semiahmoo muck soils in greenhouses and irrigated with water containing a cocktail of rifampicin-resistant generic Escherichia coli and Salmonella spp. (4.80 log CFU/ml). To mimic conventional practices, mature onions remained undisturbed in soil without irrigation for 12 days prior to being lifted and cured for 16 additional days. Surviving generic E. coli and Salmonella spp. were selectively enumerated on using standard plating (Hektoen Enteric Agar with rifampicin; HE + rif) or most probable number (lactose broth with rifampicin; HE + rif) methods. Generic E. coli and Salmonella spp. on onions decreased 0.19-0.26 log CFU/g·d during the initial 12 days of finishing. At lifting, generic E. coli and Salmonella spp. had been reduced to <1 CFU/g and persisted through the end of curing. This study demonstrates conventional curing practices as an effective mitigation strategy for dry bulb onions produced with water of poor microbiological quality.

  13. Virulence repertoire of Shiga toxin-producing Escherichia coli (STEC) and enterotoxigenic Escherichia coli (ETEC) from diarrhoeic lambs of Arunachal Pradesh, India.

    PubMed

    Bandyopadhyay, Samiran; Mahanti, Achintya; Samanta, I; Dutta, T K; Ghosh, Monoj K; Bera, A K; Bandyopadhyay, Subhasis; Bhattacharya, D

    2011-03-01

    A total of 107 faecal samples were collected from diarrhoeic lambs of high altitude terrains (2,000 to 5,000 m above the mean sea level) of Tawang and West Kameng districts of Arunachal Pradesh, India. Total 234 Escherichia coli were isolated and further subjected to PCR for the study of virulence repertoire characteristics of Shiga toxin-producing E. coli (STEC) and enterotoxigenic E. coli (ETEC). Out of the 234 isolated E. coli, 32% were found positive for STEC, and 9% were carrying virulence gene for ETEC. The isolated STEC serogroups were O159, O127, O120, O113, O60, O30, O25, O8 and O2. Of all the 74 STEC strains, PCR showed that 18% isolates carried stx ( 1 ), 26% possessed stx ( 2 ) and 47% produced positive amplicon for both. Other virulent attributes like intimin (eaeA), enterohaemolysin (ehxA) and STEC auto-agglutinating adhesin (saa) were present in 18%, 43% and 44% of the isolates, respectively. The isolated ETEC serogroups were O172, O170, O159, O146, O127, O120, O113, O86, O75, O60, O30, O25, O8, O2, OR and OUT. Of the 22 ETEC-positive isolates, 23%, 18% and 4.5% possessed the gene only for LT, STa and STb, respectively, whereas 54% carried genes for both LT and STb. Some serogroups of E. coli like O159, O127, O120, O113, O60, O30, O25, O8 and O2 possessed genes for both Shiga toxin and enterotoxin. This study is the first report of ETEC isolation from diarrhoeic lambs in India. The moderately high proportion of STEC and ETEC in the diarrhoeic lambs implicated that these animals are important reservoir of STEC and ETEC. This is really a grave concern for the 'brokpas' and nomads (shepherds) who share a close relationship with this animals for their livelihood. This study also indicates that ETEC may be a major cause for frequent diarrhoeal episodes in lambs of this region. PMID:21104315

  14. Shiga toxin-producing Escherichia coli in beef retail markets from Argentina

    PubMed Central

    Brusa, Victoria; Aliverti, Virginia; Aliverti, Florencia; Ortega, Emanuel E.; de la Torre, Julian H.; Linares, Luciano H.; Sanz, Marcelo E.; Etcheverría, Analía I.; Padola, Nora L.; Galli, Lucía; Peral García, Pilar; Copes, Julio; Leotta, Gerardo A.

    2013-01-01

    Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens that cause mild or serious diseases and can lead to people death. This study reports the prevalence and characteristics of STEC O157 and non-O157 in commercial ground beef and environmental samples, including meat table, knife, meat mincing machine, and manipulator hands (n = 450) obtained from 90 retail markets over a nine-month period. The STEC isolates were serotyped and virulence genes as stx (Shiga toxin), rfbO157] (O157 lipopolysaccharide), fliCH7 (H7 flagellin), eae (intimin), ehxA (enterohemolysin) and saa (STEC autoagglutinating adhesin), were determined. STEC O157 were identified in 23 (25.5%) beef samples and 16 (4.4%) environmental samples, while STEC non-O157 were present in 47 (52.2%) and 182 (50.5%), respectively. Among 54 strains isolated, 17 were STEC O157:H7 and 37 were STEC non-O157. The prevalent genotype for O157 was stx2/eae/ehxA/fliCH7 (83.4%), and for STEC non-O157 the most frequent ones were stx1/stx2/saa/ehxA (29.7%); stx2 (29.7%); and stx2/saa/ehxA (27%). None of the STEC non-O157 strains were eae-positive. Besides O157:H7, other 20 different serotypes were identified, being O8:H19, O178:H19, and O174:H28 the prevalent. Strains belonging to the same serotype could be isolated from different sources of the same retail market. Also, the same serotype could be detected in different stores. In conclusion, screening techniques are increasingly sensitive, but the isolation of STEC non-O157 is still a challenge. Moreover, with the results obtained from the present work, although more studies are needed, cross-contamination between meat and the environment could be suspected. PMID:23346554

  15. Whole-Genome Sequencing for National Surveillance of Shiga Toxin–Producing Escherichia coli O157

    PubMed Central

    Dallman, Timothy J.; Byrne, Lisa; Ashton, Philip M.; Cowley, Lauren A.; Perry, Neil T.; Adak, Goutam; Petrovska, Liljana; Ellis, Richard J.; Elson, Richard; Underwood, Anthony; Green, Jonathan; Hanage, William P.; Jenkins, Claire; Grant, Kathie; Wain, John

    2015-01-01

    Background. National surveillance of gastrointestinal pathogens, such as Shiga toxin–producing Escherichia coli O157 (STEC O157), is key to rapidly identifying linked cases in the distributed food network to facilitate public health interventions. In this study, we used whole-genome sequencing (WGS) as a tool to inform national surveillance of STEC O157 in terms of identifying linked cases and clusters and guiding epidemiological investigation. Methods. We retrospectively analyzed 334 isolates randomly sampled from 1002 strains of STEC O157 received by the Gastrointestinal Bacteria Reference Unit at Public Health England, Colindale, in 2012. The genetic distance between each isolate, as estimated by WGS, was calculated and phylogenetic methods were used to place strains in an evolutionary context. Results. Estimates of linked clusters representing STEC O157 outbreaks in England and Wales increased by 2-fold when WGS was used instead of traditional typing techniques. The previously unidentified clusters were often widely geographically distributed and small in size. Phylogenetic analysis facilitated identification of temporally distinct cases sharing common exposures and delineating those that shared epidemiological and temporal links. Comparison with multi locus variable number tandem repeat analysis (MLVA) showed that although MLVA is as sensitive as WGS, WGS provides a more timely resolution to outbreak clustering. Conclusions. WGS has come of age as a molecular typing tool to inform national surveillance of STEC O157; it can be used in real time to provide the highest strain-level resolution for outbreak investigation. WGS allows linked cases to be identified with unprecedented specificity and sensitivity that will facilitate targeted and appropriate public health investigations. PMID:25888672

  16. Prevalence of Shiga toxin-producing Escherichia coli in beef cattle.

    PubMed

    Hussein, Hussein S; Bollinger, Laurie M

    2005-10-01

    A large number of Shiga toxin-producing Escherichia coli (STEC) strains have caused major outbreaks and sporadic cases of human illnesses, including mild diarrhea, bloody diarrhea, hemorrhagic colitis, and the life-threatening hemolytic uremic syndrome. These illnesses have been traced to both O157 and non-O157 STEC. In a large number of STEC-associated outbreaks, the infections were attributed to consumption of ground beef or other beef products contaminated with cattle feces. Thus, beef cattle are considered reservoirs of STEC and can pose significant health risks to humans. The global nature of the human food supply suggests that safety concerns with beef will continue and the challenges facing the beef industry will increase at the production and processing levels. To be prepared to address these concerns and challenges, it is critical to assess the role of beef cattle in human STEC infections. In this review, published reports on STEC in beef cattle were evaluated to achieve the following specific objectives: (i) assess the prevalence of STEC in beef cattle, and (ii) determine the potential health risks of STEC strains from beef cattle. The latter objective is critically important because many beef STEC isolates are highly virulent. Global testing of beef cattle feces revealed wide ranges of prevalence rates for O157 STEC (i.e., 0.2 to 27.8%) and non-O157 STEC (i.e., 2.1 to 70.1%). Of the 261 STEC serotypes found in beef cattle, 44 cause hemolytic uremic syndrome and 37 cause other illnesses.

  17. Prevalence of shiga toxin-producing Escherichia coli in dairy cattle and their products.

    PubMed

    Hussein, H S; Sakuma, T

    2005-02-01

    The main objective of this review was to assess the role of dairy cattle and their products in human infections with Shiga toxin-producing Escherichia coli (STEC). A large number of STEC strains (e.g., members of the serogroups O26, O91, O103, O111, O118, O145, and O166) have caused major outbreaks and sporadic cases of human illnesses that have ranged from mild diarrhea to the life-threatening hemolytic uremic syndrome. These illnesses were traced to O157 and non-O157 STEC. In most cases, STEC infection was attributed to consumption of ground beef or dairy products that were contaminated with cattle feces. Thus, dairy cattle are considered reservoirs of STEC and can impose a significant health risk to humans. The global nature of food supply suggests that safety concerns with beef and dairy foods will continue and the challenges facing the dairy industry will increase at the production and processing levels. In this review, published reports on STEC in dairy cattle and their products were evaluated to achieve the following specific objectives: 1) to assemble a database on human infections with STEC from dairy cattle, 2) to assess prevalence of STEC in dairy cattle, and 3) to determine the health risks associated with STEC strains from dairy cattle. The latter objective is critically important, as many dairy STEC isolates are known to be of high virulence. Fecal testing of dairy cattle worldwide showed wide ranges of prevalence rates for O157 (0.2 to 48.8%) and non-O157 STEC (0.4 to 74.0%). Of the 193 STEC serotypes of dairy cattle origin, 24 have been isolated from patients with hemolytic uremic syndrome. Such risks emphasize the importance and the need to develop long-term strategies to assure safety of foods from dairy cattle.

  18. Detection, Characterization, and Typing of Shiga Toxin-Producing Escherichia coli

    PubMed Central

    Parsons, Brendon D.; Zelyas, Nathan; Berenger, Byron M.; Chui, Linda

    2016-01-01

    Shiga toxin-producing Escherichia coli (STEC) are responsible for gastrointestinal diseases reported in numerous outbreaks around the world. Given the public health importance of STEC, effective detection, characterization and typing is critical to any medical laboratory system. While non-O157 serotypes account for the majority of STEC infections, frontline microbiology laboratories may only screen for STEC using O157-specific agar-based methods. As a result, non-O157 STEC infections are significantly under-reported. This review discusses recent advances on the detection, characterization and typing of STEC with emphasis on work performed at the Alberta Provincial Laboratory for Public Health (ProvLab). Candidates for the detection of all STEC serotypes include chromogenic agars, enzyme immunoassays (EIA) and quantitative real time polymerase chain reaction (qPCR). Culture methods allow further characterization of isolates, whereas qPCR provides the greatest sensitivity and specificity, followed by EIA. The virulence gene profiles using PCR arrays and stx gene subtypes can subsequently be determined. Different non-O157 serotypes exhibit markedly different virulence gene profiles and a greater prevalence of stx1 than stx2 subtypes compared to O157:H7 isolates. Finally, recent innovations in whole genome sequencing (WGS) have allowed it to emerge as a candidate for the characterization and typing of STEC in diagnostic surveillance isolates. Methods of whole genome analysis such as single nucleotide polymorphisms and k-mer analysis are concordant with epidemiological data and standard typing methods, such as pulsed-field gel electrophoresis and multiple-locus variable number tandem repeat analysis while offering additional strain differentiation. Together these findings highlight improved strategies for STEC detection using currently available systems and the development of novel approaches for future surveillance. PMID:27148176

  19. Diverse Virulence Gene Content of Shiga Toxin-Producing Escherichia coli from Finishing Swine

    PubMed Central

    Fratamico, Pina M.; Bagi, Lori; Delannoy, Sabine; Fach, Patrick; Manning, Shannon D.; Funk, Julie A.

    2014-01-01

    Shiga toxin-producing Escherichia coli (STEC) infections are a critical public health concern because they can cause severe clinical outcomes, such as hemolytic uremic syndrome, in humans. Determining the presence or absence of virulence genes is essential in assessing the potential pathogenicity of STEC strains. Currently, there is limited information about the virulence genes carried by swine STEC strains; therefore, this study was conducted to examine the presence and absence of 69 virulence genes in STEC strains recovered previously from finishing swine in a longitudinal study. A subset of STEC strains was analyzed by pulsed-field gel electrophoresis (PFGE) to examine their genetic relatedness. Swine STEC strains (n = 150) were analyzed by the use of a high-throughput real-time PCR array system, which included 69 virulence gene targets. Three major pathotypes consisted of 16 different combinations of virulence gene profiles, and serotypes were determined in the swine STEC strains. The majority of the swine STEC strains (n = 120) belonged to serotype O59:H21 and carried the same virulence gene profile, which consisted of 9 virulence genes: stx2e, iha, ecs1763, lpfAO113, estIa (STa), ehaA, paa, terE, and ureD. The eae, nleF, and nleH1-2 genes were detected in one swine STEC strain (O49:H21). Other genes encoding adhesins, including iha, were identified (n = 149). The PFGE results demonstrated that swine STEC strains from pigs raised in the same finishing barn were closely related. Our results revealed diverse virulence gene contents among the members of the swine STEC population and enhance understanding of the dynamics of transmission of STEC strains among pigs housed in the same barn. PMID:25107960

  20. Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli

    PubMed Central

    Andrade, Fernanda B.; Nepomuceno, Roberto; Silva, Anderson; Munhoz, Danielle D.; Yamamoto, Bruno B.; Luz, Daniela; Abreu, Patrícia A. E.; Horton, Denise S. P. Q.; Elias, Waldir P.; Ramos, Oscar H. P.; Piazza, Roxane M. F.

    2015-01-01

    Background Diarrhea is a prevalent pathological condition frequently associated to the colonization of the small intestine by enterotoxigenic Escherichia coli (ETEC) strains, known to be endemic in developing countries. These strains can produce two enterotoxins associated with the manifestation of clinical symptoms that can be used to detect these pathogens. Although several detection tests have been developed, minimally equipped laboratories are still in need of simple and cost-effective methods. With the aim to contribute to the development of such diagnostic approaches, we describe here two mouse hybridoma-derived single chain fragment variable (scFv) that were produced in E. coli against enterotoxins of ETEC strains. Methods and Findings Recombinant scFv were developed against ETEC heat-labile toxin (LT) and heat-stable toxin (ST), from previously isolated hybridoma clones. This work reports their design, construction, molecular and functional characterization against LT and ST toxins. Both antibody fragments were able to recognize the cell-interacting toxins by immunofluorescence, the purified toxins by ELISA and also LT-, ST- and LT/ST-producing ETEC strains. Conclusion The developed recombinant scFvs against LT and ST constitute promising starting point for simple and cost-effective ETEC diagnosis. PMID:26154103

  1. Prevalence and characterization of extended-spectrum beta-lactamase (ESBL)- and CMY-2-producing Escherichia coli isolates from healthy food-producing animals in Tunisia.

    PubMed

    Ben Sallem, Rym; Ben Slama, Karim; Sáenz, Yolanda; Rojo-Bezares, Beatriz; Estepa, Vanesa; Jouini, Ahlem; Gharsa, Haythem; Klibi, Naouel; Boudabous, Abdellatif; Torres, Carmen

    2012-12-01

    The prevalence of extended-spectrum beta-lactamase (ESBL)- and plasmidic AmpC-beta-lactamase (pAmpC-BL)-producing Escherichia coli isolates has been studied in food-producing animals at the farm level in Tunisia, and recovered isolates were characterized for the presence of other resistance genes and integrons. Eighty fecal samples of food-producing animals (23 sheep, 22 chickens, 22 cattle, six horses, five rabbits, and two dromedaries) were obtained from 35 different farms in Tunisia in 2011. Samples were inoculated onto MacConkey agar plates supplemented with cefotaxime (2 mg/L) for cefotaxime-resistant (CTX(R)) E. coli recovery. CTX(R) E. coli isolates were detected in 11 out of 80 samples (13.8%), and one isolate per sample was further characterized (10 from chickens and one from a dromedary). The 11 CTX(R) isolates were distributed into phylogroups: B1 (five isolates), A (two isolates), D (three isolates), and B2 (one isolate). The following beta-lactamase genes were detected: bla(CTX-M-1) (seven isolates), bla(CTX-M-1)+bla(TEM-135) (one isolate), bla(CTX-M-1)+bla(TEM-1b) (one isolate), and bla(CMY-2) (two isolates). All ESBL- and pAmpC-BL-producing E. coli strains showed unrelated pulsed-field gel electrophoresis patterns. Seven isolates contained class 1 integrons with four gene cassette arrangements: dfrA17-aadA5 (three isolates), dfrA1-aadA1 (two isolates), dfrA15-aadA1 (one isolate), and aadA1 (one isolate). All isolates showed tetracycline resistance and contained the tet(A) +/- tet(B) genes. Virulence genes detected were as follows (number of isolates in parentheses): fimA (10); aer (eight); papC (two); and papGIII, hly, cnf, and bfp (none). Chicken farms constitute a reservoir of ESBL- and pAmpC-BL-producing E. coli isolates of the CTX-M-1 and CMY-2 types that potentially could be transmitted to humans via the food chain or by direct contact.

  2. Epidemiology and risk factors for isolation of Escherichia coli producing CTX-M-type extended-spectrum β-lactamase in a large U.S. Medical Center.

    PubMed

    Hayakawa, Kayoko; Gattu, Sureka; Marchaim, Dror; Bhargava, Ashish; Palla, Mohan; Alshabani, Khaled; Gudur, Uma Mahesh; Pulluru, Harish; Bathina, Pradeep; Sundaragiri, Pranathi Rao; Sarkar, Moumita; Kakarlapudi, Hari; Ramasamy, Balaji; Nanjireddy, Priyanka; Mohin, Shah; Dasagi, Meenakshi; Datla, Satya; Kuchipudi, Vamsi; Reddy, Swetha; Shahani, Shobha; Upputuri, Vijaya; Marrey, Satya; Gannamani, Vedavyas; Madhanagopal, Nandhini; Annangi, Srinadh; Sudha, Busani; Muppavarapu, Kalyan Srinivas; Moshos, Judy A; Lephart, Paul R; Pogue, Jason M; Bush, Karen; Kaye, Keith S

    2013-08-01

    A case-case-control study was conducted to identify independent risk factors for recovery of Escherichia coli strains producing CTX-M-type extended-spectrum β-lactamases (CTX-M E. coli) within a large Southeastern Michigan medical center. Unique cases with isolation of ESBL-producing E. coli from February 2010 through July 2011 were analyzed by PCR for blaCTX-M, blaTEM, and blaSHV genes. Patients with CTX-M E. coli were compared to patients with E. coli strains not producing CTX-M-type ESBLs (non-CTX-M E. coli) and uninfected controls. Of 575 patients with ESBL-producing E. coli, 491 (85.4%) isolates contained a CTX-M ESBL gene. A total of 319 (84.6%) patients with CTX-M E. coli (282 [74.8%] CTX-M-15 type) were compared to 58 (15.4%) non-CTX-M E. coli patients and to uninfected controls. Independent risk factors for CTX-M E. coli isolation compared to non-CTX-M E. coli included male gender, impaired consciousness, H2 blocker use, immunosuppression, and exposure to penicillins and/or trimethoprim-sulfamethoxazole. Compared to uninfected controls, independent risk factors for isolation of CTX-M E. coli included presence of a urinary catheter, previous urinary tract infection, exposure to oxyimino-cephalosporins, dependent functional status, non-home residence, and multiple comorbid conditions. Within 48 h of admission, community-acquired CTX-M E. coli (n = 51 [16%]) and non-CTX-M E coli (n = 11 [19%]) strains were isolated from patients with no recent health care contacts. CTX-M E. coli strains were more resistant to multiple antibiotics than non-CTX-M E. coli strains. CTX-M-encoding genes, especially bla(CTX-M-15) type, represented the most common ESBL determinants from ESBL-producing E. coli, the majority of which were present upon admission. Septic patients with risk factors for isolation of CTX-M E. coli should be empirically treated with appropriate agents. Regional infection control efforts and judicious antibiotic use are needed to control the spread of these

  3. Epidemiology and Risk Factors for Isolation of Escherichia coli Producing CTX-M-Type Extended-Spectrum β-Lactamase in a Large U.S. Medical Center

    PubMed Central

    Gattu, Sureka; Marchaim, Dror; Bhargava, Ashish; Palla, Mohan; Alshabani, Khaled; Gudur, Uma Mahesh; Pulluru, Harish; Bathina, Pradeep; Sundaragiri, Pranathi Rao; Sarkar, Moumita; Kakarlapudi, Hari; Ramasamy, Balaji; Nanjireddy, Priyanka; Mohin, Shah; Dasagi, Meenakshi; Datla, Satya; Kuchipudi, Vamsi; Reddy, Swetha; Shahani, Shobha; Upputuri, Vijaya; Marrey, Satya; Gannamani, Vedavyas; Madhanagopal, Nandhini; Annangi, Srinadh; Sudha, Busani; Muppavarapu, Kalyan Srinivas; Moshos, Judy A.; Lephart, Paul R.; Pogue, Jason M.; Bush, Karen; Kaye, Keith S.

    2013-01-01

    A case-case-control study was conducted to identify independent risk factors for recovery of Escherichia coli strains producing CTX-M-type extended-spectrum β-lactamases (CTX-M E. coli) within a large Southeastern Michigan medical center. Unique cases with isolation of ESBL-producing E. coli from February 2010 through July 2011 were analyzed by PCR for blaCTX-M, blaTEM, and blaSHV genes. Patients with CTX-M E. coli were compared to patients with E. coli strains not producing CTX-M-type ESBLs (non-CTX-M E. coli) and uninfected controls. Of 575 patients with ESBL-producing E. coli, 491 (85.4%) isolates contained a CTX-M ESBL gene. A total of 319 (84.6%) patients with CTX-M E. coli (282 [74.8%] CTX-M-15 type) were compared to 58 (15.4%) non-CTX-M E. coli patients and to uninfected controls. Independent risk factors for CTX-M E. coli isolation compared to non-CTX-M E. coli included male gender, impaired consciousness, H2 blocker use, immunosuppression, and exposure to penicillins and/or trimethoprim-sulfamethoxazole. Compared to uninfected controls, independent risk factors for isolation of CTX-M E. coli included presence of a urinary catheter, previous urinary tract infection, exposure to oxyimino-cephalosporins, dependent functional status, non-home residence, and multiple comorbid conditions. Within 48 h of admission, community-acquired CTX-M E. coli (n = 51 [16%]) and non-CTX-M E coli (n = 11 [19%]) strains were isolated from patients with no recent health care contacts. CTX-M E. coli strains were more resistant to multiple antibiotics than non-CTX-M E. coli strains. CTX-M-encoding genes, especially blaCTX-M-15 type, represented the most common ESBL determinants from ESBL-producing E. coli, the majority of which were present upon admission. Septic patients with risk factors for isolation of CTX-M E. coli should be empirically treated with appropriate agents. Regional infection control efforts and judicious antibiotic use are needed to control the spread of these

  4. Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011–2012, Russia

    PubMed Central

    Kartsev, Nikolay N.; Fursova, Nadezhda K.; Pachkunov, Dmitry M.; Bannov, Vasiliy A.; Eruslanov, Boris V.; Svetoch, Edward A.; Dyatlov, Ivan A.

    2015-01-01

    Enterotoxin-producing Escherichia coli (ETEC) are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST) and/or heat-labile (LT) enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011–2012. It was shown that all ETEC under study (n=18) had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia) and Vladivostok city (Far-East region, Russia). Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6), О6 (n = 4), О25 (n = 5), О26 (n = 2), and O115 (n = 1). Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3) as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates—to two drugs, one isolate—to three drugs, two isolates—to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10), blaCTX-M-15 (n = 1), class 1 integron (n = 3) carrying resistance cassettes to aminoglycosides and

  5. Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011-2012, Russia.

    PubMed

    Kartsev, Nikolay N; Fursova, Nadezhda K; Pachkunov, Dmitry M; Bannov, Vasiliy A; Eruslanov, Boris V; Svetoch, Edward A; Dyatlov, Ivan A

    2015-01-01

    Enterotoxin-producing Escherichia coli (ETEC) are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST) and/or heat-labile (LT) enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011-2012. It was shown that all ETEC under study (n=18) had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia) and Vladivostok city (Far-East region, Russia). Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6), О6 (n = 4), О25 (n = 5), О26 (n = 2), and O115 (n = 1). Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3) as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates-to two drugs, one isolate-to three drugs, two isolates-to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10), blaCTX-M-15 (n = 1), class 1 integron (n = 3) carrying resistance cassettes to aminoglycosides and

  6. Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011-2012, Russia.

    PubMed

    Kartsev, Nikolay N; Fursova, Nadezhda K; Pachkunov, Dmitry M; Bannov, Vasiliy A; Eruslanov, Boris V; Svetoch, Edward A; Dyatlov, Ivan A

    2015-01-01

    Enterotoxin-producing Escherichia coli (ETEC) are one of the main causative agents of diarrhea in children especially in developing countries and travel diarrhoea in adults. Pathogenic properties of ETEC associated with their ability to produce a heat-stable (ST) and/or heat-labile (LT) enterotoxins, as well as adhesins providing bacterial adhesion to intestinal epithelial cells. This study presents the molecular characterization of the ETEC isolates collected from the Central and Far-Eastern regions of Russia in 2011-2012. It was shown that all ETEC under study (n=18) had the heat-labile enterotoxin-coding operon elt, and had no the genes of the heat-stable enterotoxin operon est. DNA sequencing revealed two types of nucleotide exchanges in the eltB gene coding subunit B of LT in isolates collected from Cherepovets city (Central region, Russia) and Vladivostok city (Far-East region, Russia). Only one ETEC strain carried genes cfaA, cfaB, cfaC and cfaD coding adhesion factor CFA/I. Expression of LT in four ETEC isolates in the agglutination reaction was detected using a latex test-system. The isolates were assigned to serogroups O142 (n = 6), О6 (n = 4), О25 (n = 5), О26 (n = 2), and O115 (n = 1). Genotyping showed that they belonged to an earlier described sequence-type ST4 (n = 3) as well as to 11 novel sequence-types ST1043, ST1312, ST3697, ST3707, ST3708, ST3709, ST3710, ST3755, ST3756, ST3757 and ST4509. The ETEC isolates displayed different levels of antimicrobial resistance. Eight isolates were resistant to only one drug, three isolates-to two drugs, one isolate-to three drugs, two isolates-to four antibacterials, and only one isolate to each of the five, six and ten antibacterials simultaneously. Genetic determinants of the resistance to beta-lactams and other classes of antibacterials on the ETEC genomes were identified. There are blaTEM (n = 10), blaCTX-M-15 (n = 1), class 1 integron (n = 3) carrying resistance cassettes to aminoglycosides and

  7. Houseflies (Musca domestica) as Vectors for Extended-Spectrum β-Lactamase-Producing Escherichia coli on Spanish Broiler Farms.

    PubMed

    Solà-Ginés, Marc; González-López, Juan José; Cameron-Veas, Karla; Piedra-Carrasco, Nuria; Cerdà-Cuéllar, Marta; Migura-Garcia, Lourdes

    2015-06-01

    Flies may act as potential vectors for the spread of resistant bacteria to different environments. This study was intended to evaluate the presence of Escherichia coli strains resistant to cephalosporins in flies captured in the areas surrounding five broiler farms. Phenotypic and molecular characterization of the resistant population was performed by different methods: MIC determination, pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and phylotyping. The presence of extended-spectrum beta-lactamase (ESBL) genes, their plasmid location, and the mobile genetic elements involved in their mobilization were studied. Additionally, the presence of 35 genes associated with virulence was evaluated. Out of 682 flies captured, 42 yielded ESBL-producing E. coli. Of these isolates, 23 contained bla(CTX-M-1), 18 contained bla(CTX-M-14), and 1 contained bla(CTX-M-9). ESBL genes were associated mainly with the presence of the IncI1 and IncFIB replicons. Additionally, all the strains were multiresistant, and five of them also harbored qnrS. Identical PFGE profiles were found for E. coli isolates obtained from flies at different sampling times, indicating a persistence of the same clones in the farm environment over months. According to their virulence genes, 81% of the isolates were considered avian-pathogenic E. coli (APEC) and 29% were considered extraintestinal pathogenic E. coli (ExPEC). The entrance of flies into broiler houses constitutes a considerable risk for colonization of broilers with multidrug-resistant E. coli. ESBLs in flies reflect the contamination status of the farm environment. Additionally, this study demonstrates the potential contribution of flies to the dissemination of virulence and resistance genes into different ecological niches.

  8. Isolation of Shiga toxin-producing Escherichia coli from fresh produce using STEC heart infusion washed blood agar with mitomycin-C.

    PubMed

    Lin, Andrew; Nguyen, Lam; Clotilde, Laurie M; Kase, Julie A; Son, Insook; Lauzon, Carol R

    2012-11-01

    The ability to detect and isolate Shiga toxin-producing Escherichia coli (STEC) remains a major challenge for food microbiologists. Although methods based on nucleic acids and antibodies have improved detection of STECs in foods, isolation of these bacteria remains arduous. STEC isolation is necessary for matching food, environmental, and clinical isolates during outbreak investigations and for distinguishing between pathogenic and nonpathogenic organisms. STEC heart infusion washed blood agar with mitomycin-C (SHIBAM) is a modification of washed sheep blood agar prepared by adding mitomycin-C and optimizing both the washed blood and base agar to better isolate STECs. Most STEC isolates produce a zone of hemolysis on SHIBAM plates and are easily distinguishable from background microbiota. Here, we present data supporting the use of SHIBAM to isolate STECs from fresh produce. SHIBAM was tested for accuracy in identifying STECs (365 of 410 STEC strains were hemolytic, and 63 of 73 E. coli strains that did not produce Shiga toxin were not hemolytic) and for recovery from artificially inoculated fresh produce (11 of 24 romaine lettuce samples and 6 of 24 tomato samples). STEC recovery with SHIBAM agar was greatly improved when compared with recovery on Levine's eosin-methylene blue agar as a reference method.

  9. Susceptibility of various oral antibacterial agents against extended spectrum β-lactamase producing Escherichia coli and Klebsiella pneumoniae.

    PubMed

    Nakamura, Tatsuya; Komatsu, Masaru; Yamasaki, Katsutoshi; Fukuda, Saori; Higuchi, Takeshi; Ono, Tamotsu; Nishio, Hisaaki; Sueyoshi, Noriyuki; Kida, Kaneyuki; Satoh, Kaori; Toda, Hirofumi; Toyokawa, Masahiro; Nishi, Isao; Sakamoto, Masako; Akagi, Masahiro; Mizutani, Tetsu; Nakai, Isako; Kofuku, Tomomi; Orita, Tamaki; Zikimoto, Takuya; Natsume, Seiko; Wada, Yasunao

    2014-01-01

    With the increase in extended spectrum β-lactamase (ESBL)-producing bacteria in the community, cases are often seen in which treatment of infectious diseases with oral antimicrobial agents is difficult. Therefore, we measured the antimicrobial activities of 14 currently available oral antimicrobial agents against ESBL-producing Escherichia coli and Klebsiella pneumoniae. Based on the standard of the Clinical and Laboratory Standards Institute (CLSI), E. coli showed high susceptibility rates of 99.4% to faropenem (FRPM). In terms of fluoroquinolones, the susceptibility rate of E. coli to levofloxacin (LVFX) was low at 32.2%, whereas it showed a good susceptibility rate of 93.1% to sitafloxacin (STFX). With respect to other antimicrobial agents, susceptibility rates to fosfomycin (FOM) and colistin (CL) were more than 90% each, whereas rates of the two antimicrobial agents expected as therapeutic agents, minocycline (MINO) and sulfamethoxazole-trimethoprim (ST), were low at 62.4% and 44.3%, respectively. Based on the CLSI standard, K. pneumoniae showed high susceptibility rates to ceftibuten (CETB) (91.89%), LVFX (86.49%), and STFX (94.6%), indicating that K. pneumoniae showed higher rates than those of E. coli, particularly to fluoroquinolones. Comparison of susceptibility rates according to E. coli genotype showed that many antimicrobial agents existed to which the CTX-M-9 group showed high susceptibility rates. However, there were many agents to which the CTX-M-1 group showed low susceptibility rates, particularly to CETB (51.1%) and LVFX (17.0%). Although there was no significant difference by genotype between FRPM, STFX, and FOM, a significant difference was observed between LVFX, MINO, and ST. Antibiotic-resistant bacteria with highly pathogenic strains have spread in the community, appropriate use of oral antimicrobial agents is required. PMID:24462425

  10. Fate of Shiga toxin-producing and generic Escherichia coli during production and ripening of semihard raw milk cheese.

    PubMed

    Peng, S; Hoffmann, W; Bockelmann, W; Hummerjohann, J; Stephan, R; Hammer, P

    2013-02-01

    The fate of 5 different Escherichia coli strains, including 3 Shiga toxin-producing E. coli (STEC) strains, was analyzed during the production and ripening of semihard raw milk cheese. The strains, which were previously isolated from raw milk cheese, were spiked into raw milk before cheese production at 2 different levels (approximately 10(1) and 10(3) cfu/mL, respectively). Two cheese types were produced, which differed in cooking temperatures (40 and 46°C). The cheeses were sampled during manufacture and the 16-wk ripening period. An increase in E. coli counts of approximately 3.5 log(10) cfu/g occurred from raw milk to fresh cheese at d 1, which was attributed to a concentration effect during cheese production and growth of the strains. During ripening over 16 wk, a slow, continuous decrease was observed for all strains. However, significant differences were found between the E. coli strains at the applied spiking levels, whereas the inactivation was similar in the 2 different cheese types. The 2 generic E. coli strains survived at higher counts than did the 3 STEC strains. Nevertheless, only 1 of the 3 STEC strains showed significantly weaker survival at both spiking levels and in both cheese types. Six of 16 cheeses made from raw milk at a low spiking level contained more than 10 cfu/g of STEC at the end of the 16-wk ripening process. After enrichment, STEC were detected in almost all cheeses at both spiking levels. Particularly because of the low infectious dose of highly pathogenic STEC, even low colony counts in raw milk cheese are a matter of concern.

  11. Broad and efficient control of major foodborne pathogenic strains of Escherichia coli by mixtures of plant-produced colicins.

    PubMed

    Schulz, Steve; Stephan, Anett; Hahn, Simone; Bortesi, Luisa; Jarczowski, Franziska; Bettmann, Ulrike; Paschke, Anne-Katrin; Tusé, Daniel; Stahl, Chad H; Giritch, Anatoli; Gleba, Yuri

    2015-10-01

    Enterohemorrhagic Escherichia coli (EHEC) is one of the leading causes of bacterial enteric infections worldwide, causing ∼100,000 illnesses, 3,000 hospitalizations, and 90 deaths annually in the United States alone. These illnesses have been linked to consumption of contaminated animal products and vegetables. Currently, other than thermal inactivation, there are no effective methods to eliminate pathogenic bacteria in food. Colicins are nonantibiotic antimicrobial proteins, produced by E. coli strains that kill or inhibit the growth of other E. coli strains. Several colicins are highly effective against key EHEC strains. Here we demonstrate very high levels of colicin expression (up to 3 g/kg of fresh biomass) in tobacco and edible plants (spinach and leafy beets) at costs that will allow commercialization. Among the colicins examined, plant-expressed colicin M had the broadest antimicrobial activity against EHEC and complemented the potency of other colicins. A mixture of colicin M and colicin E7 showed very high activity against all major EHEC strains, as defined by the US Department of Agriculture/Food and Drug Administration. Treatments with low (less than 10 mg colicins per L) concentrations reduced the pathogenic bacterial load in broth culture by 2 to over 6 logs depending on the strain. In experiments using meats spiked with E. coli O157:H7, colicins efficiently reduced the population of the pathogen by at least 2 logs. Plant-produced colicins could be effectively used for the broad control of pathogenic E. coli in both plant- and animal-based food products and, in the United States, colicins could be approved using the generally recognized as safe (GRAS) regulatory approval pathway.

  12. Broad and efficient control of major foodborne pathogenic strains of Escherichia coli by mixtures of plant-produced colicins

    PubMed Central

    Schulz, Steve; Stephan, Anett; Hahn, Simone; Bortesi, Luisa; Jarczowski, Franziska; Bettmann, Ulrike; Paschke, Anne-Katrin; Tusé, Daniel; Stahl, Chad H.; Giritch, Anatoli; Gleba, Yuri

    2015-01-01

    Enterohemorrhagic Escherichia coli (EHEC) is one of the leading causes of bacterial enteric infections worldwide, causing ∼100,000 illnesses, 3,000 hospitalizations, and 90 deaths annually in the United States alone. These illnesses have been linked to consumption of contaminated animal products and vegetables. Currently, other than thermal inactivation, there are no effective methods to eliminate pathogenic bacteria in food. Colicins are nonantibiotic antimicrobial proteins, produced by E. coli strains that kill or inhibit the growth of other E. coli strains. Several colicins are highly effective against key EHEC strains. Here we demonstrate very high levels of colicin expression (up to 3 g/kg of fresh biomass) in tobacco and edible plants (spinach and leafy beets) at costs that will allow commercialization. Among the colicins examined, plant-expressed colicin M had the broadest antimicrobial activity against EHEC and complemented the potency of other colicins. A mixture of colicin M and colicin E7 showed very high activity against all major EHEC strains, as defined by the US Department of Agriculture/Food and Drug Administration. Treatments with low (less than 10 mg colicins per L) concentrations reduced the pathogenic bacterial load in broth culture by 2 to over 6 logs depending on the strain. In experiments using meats spiked with E. coli O157:H7, colicins efficiently reduced the population of the pathogen by at least 2 logs. Plant-produced colicins could be effectively used for the broad control of pathogenic E. coli in both plant- and animal-based food products and, in the United States, colicins could be approved using the generally recognized as safe (GRAS) regulatory approval pathway. PMID:26351689

  13. Evaluaiton of a novel antimicrobial solution and its potential for control E. coli O157:H7, non-O157:H7 shiga toxin-producing E. coli, Salmononella spp., and Listeria monocytogenes on beef

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The goal of this study was to evaluate the efficacy of a novel antimicrobial solution made with chitosan, lauric arginate ester, and organic acids on Escherichia coli O157:H7, Salmonella spp., Listeria monocytogenes, and non-O157 shiga toxin-producing E. coli cocktails and to test its potential to b...

  14. Water buffaloes (Bubalus bubalis) identified as an important reservoir of Shiga toxin-producing Escherichia coli in Brazil.

    PubMed

    Oliveira, Murilo G; Brito, José R Feitosa; Carvalho, Roberta R; Guth, Beatriz E C; Gomes, Tânia A T; Vieira, Mônica A M; Kato, Maria A M F; Ramos, Isabel I; Vaz, Tânia M I; Irino, Kinue

    2007-09-01

    The presence of Shiga toxin-producing Escherichia coli (STEC) in water buffaloes is reported for the first time in South America. The prevalence of STEC ranged from 0 to 64% depending on the farm. STEC isolates exhibiting the genetic profiles stx(1)stx(2)ehxA iha saa and stx(2)ehxA iha saa predominated. Of the 20 distinct serotypes identified, more than 50% corresponded to serotypes associated with human diseases.

  15. Identification of five Shiga toxin-producing Escherichia coli genes by Luminex microbead-based suspension array.

    PubMed

    Son, Insook; Binet, Rachel; Lin, Andrew; Hammack, Thomas S; Kase, Julie A

    2015-04-01

    To rapidly identify the presence of potentially virulent O157:H7 and non-O157 Shiga toxin-producing Escherichia coli (STEC), a PCR-based Luminex suspension assay was developed to detect the genes coding for four virulence factors (stx1, stx2, eae, and ehxA) plus the O157:H7-specific +93 uidA single nucleotide polymorphism.

  16. Synthetic biology: Tailor-made genetic codes

    NASA Astrophysics Data System (ADS)

    Jewett, Michael C.; Noireaux, Vincent

    2016-04-01

    Expanding the range of amino acids polymerizable by ribosomes could enable new functionalities to be added to polypeptides. Now, the genetic code has been reprogrammed using a reconstituted in vitro translation system to enable synthesis of unnatural peptides with unmatched flexibility.

  17. Detection of verotoxin producing Escherichia coli in marine environments of the Caribbean.

    PubMed

    Walker, Trisha J; Bachoon, D S; Otero, Ernesto; Ramsubhag, Adesh

    2013-11-15

    The goal of this study was to determine the potential for Enterohemorrhagic Escherichia coli O157:H7 (EHEC) contamination in tropical marine waters. Samples were collected from urban, suburban, and rural sites around the islands of Puerto Rico and The Republic of Trinidad and Tobago. Quantification of E. coli and EHEC was evaluated using MI plates and qPCR. EHEC was detected in six sites in Puerto Rico: West of La Parguera Town, Boquilla, Oro Creek, Fishers Association, Joyuda Lagoon, and Boqueron Wetland Creek and in two rural sites in Trinidad: Balandra Bay and Quinam Bay. Plate count enumeration of E. coli was not a reliable indicator for the presence of EHEC. The sites where EHEC was detected on both islands are used for recreational bathing, water sports and recreational/commercial fisheries and therefore pose a public potential health risk.

  18. Molecular epidemiology of carbapenemase-producing Escherichia coli and the prevalence of ST131 subclone H30 in Shanghai, China.

    PubMed

    Zhang, F; Zhu, D; Xie, L; Guo, X; Ni, Y; Sun, J

    2015-06-01

    The molecular characteristics and epidemiology of carbapenemase-producing Escherichia coli (CPEC) isolates from Shanghai, China, were investigated using 21 imipenem-resistant E. coli isolates obtained from a Shanghai teaching hospital from 2011 to 2014. The presence of bla KPC, bla IMP, bla VIM, bla OXA-48, and bla NDM was assessed by polymerase chain reaction (PCR) amplification and sequencing. CPEC isolates were characterized by the Etest®, multilocus sequence typing (MLST), and pulse-field gel electrophoresis (PFGE). Plasmids carrying resistance genes were analyzed by conjugation experiments, replicon typing, plasmid MLST (pMLST), S1 nuclease PFGE (S1-PFGE), and Southern hybridization. The genetic environment of the resistance genes was determined by PCR and sequencing. Among the 21 E. coli isolates, 16 produced carbapenemases; of these, ten isolates transferred carbapenemase-encoding plasmids to recipient bacteria. Nine of the 16 isolates were clonally related, and their PFGE patterns were designated type A. ST131 was the predominant sequence type (11 isolates, 68.8 %); the H30 subclone comprised 81.8 % of the ST131 strains. In all three isolates, bla IMP-4 was located on 50-kb IncN plasmids. All but two bla KPC-2 genes were carried on IncF plasmids of various sizes. Hence, both clone-spread and horizontal transfer mediated the dissemination of carbapenemase-producing genes in the Shanghai isolates.

  19. Characterization of Shiga Toxin-Producing Escherichia coli O157 Isolates from Bovine Carcasses.

    PubMed

    Fontcuberta, M; Planell, R; Torrents, A; Sabaté, S; Gonzalez, R; Ramoneda, M; de Simón, M

    2016-08-01

    The main purpose of this study was to determine the prevalence of Escherichia coli O157 on bovine carcasses before and after chilling at a large slaughterhouse located in the city of Barcelona, Spain, to assess the effectiveness of dry chilling on reducing E. coli O157 contamination of carcasses. In addition, the study characterized the E. coli O157 strains isolated in terms of virulence factors, antibiotic susceptibility, and their genetic diversity. Individual bovine carcasses were sampled before (n = 300) and after (n = 300) chilling over an 8-month period. Positive samples for E. coli O157 were subjected to virulence screening by PCR (stx1, stx2, and eaeA genes and the fliCH7 gene), antimicrobial susceptibility testing, and molecular typing by pulsed-field gel electrophoresis. A total of 9.7% (29 of 300) of the nonrefrigerated carcasses examined and 2.3% (7 of 300) of the refrigerated carcasses were positive for E. coli O157. All the isolates were serotype O157:H7, 92% (33 of 36) carried the stx1, stx2, and eaeA genes, and 8% (3 of 36) carried the stx2 and eaeA genes. Antimicrobial susceptibility testing showed a high degree of resistance: 29 strains (81%) were resistant to at least 1 antimicrobial of the 12 antimicrobials tested; 69% (25 of 36) were resistant to 4 or more antimicrobials. Molecular typing by pulsed-field gel electrophoresis found a high diversity of genetic types, implying little cross-contamination in the slaughterhouse. This study confirms that E. coli O157:H7 is present on the carcasses slaughtered in Spain, although its prevalence is reduced by the dry chilling process used. The recovered isolates showed potential pathogenesis and a high degree of multidrug resistance, confirming the importance of bovine meat monitoring.

  20. Characterization of Shiga Toxin-Producing Escherichia coli O157 Isolates from Bovine Carcasses.

    PubMed

    Fontcuberta, M; Planell, R; Torrents, A; Sabaté, S; Gonzalez, R; Ramoneda, M; de Simón, M

    2016-08-01

    The main purpose of this study was to determine the prevalence of Escherichia coli O157 on bovine carcasses before and after chilling at a large slaughterhouse located in the city of Barcelona, Spain, to assess the effectiveness of dry chilling on reducing E. coli O157 contamination of carcasses. In addition, the study characterized the E. coli O157 strains isolated in terms of virulence factors, antibiotic susceptibility, and their genetic diversity. Individual bovine carcasses were sampled before (n = 300) and after (n = 300) chilling over an 8-month period. Positive samples for E. coli O157 were subjected to virulence screening by PCR (stx1, stx2, and eaeA genes and the fliCH7 gene), antimicrobial susceptibility testing, and molecular typing by pulsed-field gel electrophoresis. A total of 9.7% (29 of 300) of the nonrefrigerated carcasses examined and 2.3% (7 of 300) of the refrigerated carcasses were positive for E. coli O157. All the isolates were serotype O157:H7, 92% (33 of 36) carried the stx1, stx2, and eaeA genes, and 8% (3 of 36) carried the stx2 and eaeA genes. Antimicrobial susceptibility testing showed a high degree of resistance: 29 strains (81%) were resistant to at least 1 antimicrobial of the 12 antimicrobials tested; 69% (25 of 36) were resistant to 4 or more antimicrobials. Molecular typing by pulsed-field gel electrophoresis found a high diversity of genetic types, implying little cross-contamination in the slaughterhouse. This study confirms that E. coli O157:H7 is present on the carcasses slaughtered in Spain, although its prevalence is reduced by the dry chilling process used. The recovered isolates showed potential pathogenesis and a high degree of multidrug resistance, confirming the importance of bovine meat monitoring. PMID:27497130

  1. Verocytotoxin-producing Escherichia coli in chamois (Rupicapra rupicapra) and cattle in Austria.

    PubMed

    Freidl, Gudrun; Stalder, Gabrielle; Kostić, Tanja; Sessitsch, Angela; Beiglböck, Christoph; Walzer, Christian

    2011-07-01

    We assessed the prevalence of verotoxigenic Escherichia coli (VTEC) in chamois (Rupicapra rupicapra) and livestock grazing on a mountain pasture in Austria during June-August 2009. We detected VTEC throughout the sampling period in high numbers in cattle as well as in chamois, leading to the assumption that the degree of contamination of the environment is high. This is the first report of pathogenic E. coli identified in chamois, implicating chamois as a new potential reservoir of these zoonotic pathogens. Because the study area also serves recreational purposes, there is a risk of humans acquiring infection via direct or indirect contact. PMID:21719837

  2. Verocytotoxin-producing Escherichia coli in chamois (Rupicapra rupicapra) and cattle in Austria.

    PubMed

    Freidl, Gudrun; Stalder, Gabrielle; Kostić, Tanja; Sessitsch, Angela; Beiglböck, Christoph; Walzer, Christian

    2011-07-01

    We assessed the prevalence of verotoxigenic Escherichia coli (VTEC) in chamois (Rupicapra rupicapra) and livestock grazing on a mountain pasture in Austria during June-August 2009. We detected VTEC throughout the sampling period in high numbers in cattle as well as in chamois, leading to the assumption that the degree of contamination of the environment is high. This is the first report of pathogenic E. coli identified in chamois, implicating chamois as a new potential reservoir of these zoonotic pathogens. Because the study area also serves recreational purposes, there is a risk of humans acquiring infection via direct or indirect contact.

  3. Phylogenomic approaches to determine the zoonotic potential of Shiga toxin-producing Escherichia coli (STEC) isolated from Zambian dairy cattle

    PubMed Central

    Mainda, Geoffrey; Lupolova, Nadejda; Sikakwa, Linda; Bessell, Paul R.; Muma, John B.; Hoyle, Deborah V.; McAteer, Sean P.; Gibbs, Kirsty; Williams, Nicola J.; Sheppard, Samuel K.; La Ragione, Roberto M.; Cordoni, Guido; Argyle, Sally A.; Wagner, Sam; Chase-Topping, Margo E.; Dallman, Timothy J.; Stevens, Mark P.; Bronsvoort, Barend M. deC.; Gally, David L.

    2016-01-01

    This study assessed the prevalence and zoonotic potential of Shiga toxin-producing Escherichia coli (STEC) sampled from 104 dairy units in the central region of Zambia and compared these with isolates from patients presenting with diarrhoea in the same region. A subset of 297 E. coli strains were sequenced allowing in silico analyses of phylo- and sero-groups. The majority of the bovine strains clustered in the B1 ‘commensal’ phylogroup (67%) and included a diverse array of serogroups. 11% (41/371) of the isolates from Zambian dairy cattle contained Shiga toxin genes (stx) while none (0/73) of the human isolates were positive. While the toxicity of a subset of these isolates was demonstrated, none of the randomly selected STEC belonged to key serogroups associated with human disease and none encoded a type 3 secretion system synonymous with typical enterohaemorrhagic strains. Positive selection for E. coli O157:H7 across the farms identified only one positive isolate again indicating this serotype is rare in these animals. In summary, while Stx-encoding E. coli strains are common in this dairy population, the majority of these strains are unlikely to cause disease in humans. However, the threat remains of the emergence of strains virulent to humans from this reservoir. PMID:27220895

  4. Enhancing isomaltulose production by recombinant Escherichia coli producing sucrose isomerase: culture medium optimization containing agricultural wastes and cell immobilization.

    PubMed

    Li, Sha; Xu, Hong; Yu, Jianguang; Wang, Yanyuan; Feng, Xiaohai; Ouyang, Pingkai

    2013-10-01

    Isomaltulose is a structural isomer of sucrose commercially used in food industries. In this work, recombinant Escherichia coli producing sucrose isomerase (SIase) was used to convert sucrose into isomaltulose. To develop an economical industrial medium, untreated cane molasses (10.63 g l⁻¹), yeast extract (25.93 g l⁻¹), and corn steep liquor (10.45 g l⁻¹) were used as main culture compositions for SIase production. The relatively high SIase activity (14.50 ± 0.11 U mg DCW⁻¹) was obtained by the recombinant cells. To the best of our knowledge, this is the first investigation on SIase production by engineered E. coli using untreated cane molasses. The recombinant E. coli cells expressing the SIase gene were immobilized in calcium alginate gel in order to improve the efficiency of recycling. The immobilization was most effective with 2 % (w/v) sodium alginate and 3 % (w/v) calcium chloride. The optimal initial biomass for immobilization was 20 % (w/v, wet wt.), with a hardening time of 8 h for cell immobilization. The immobilized E. coli cells exhibited good stability for 30 batches with the productivity of 0.45 g isomaltulose g pellet⁻¹ h⁻¹. A continuous isomaltulose formation process using a column reactor remained stable for 40 days with 83 ± 2 % isomaltulose yield, which would be beneficial for economical production of isomaltulose. PMID:23300051

  5. Prevalence of multi-antimicrobial-agent resistant, shiga toxin and enterotoxin producing Escherichia coli in surface waters of river Ganga.

    PubMed

    Ram, Siya; Vajpayee, Poornima; Shanker, Rishi

    2007-11-01

    The consumption of polluted surface water for domestic and recreational purposes by large populations in developing nations is a major cause of diarrheal disease related mortality. The river Ganga and its tributaries meet 40% of the water requirement for drinking and irrigation in India. In this study, Escherichia coli isolates (n=75) of the river Ganga water were investigated for resistance to antimicrobial agents (n=15) and virulence genes specific to shiga toxin (STEC) and enterotoxin producing E. coli (ETEC). E. coli isolates from the river Ganga water exhibit resistance to multiple antimicrobial agents. The distribution of antimicrobial agent resistance in E. colivaries significantly (chi2: 81.28 at df = 24, p < 0.001) between the sites. Both stx1 and stx2 genes were present in 82.3% of STEC (n=17) while remaining isolates possess either stxl (11.8%) or stx2 (5.9%). The presence of eaeA, hlyA, and chuA genes was observed in 70.6, 88.2, and 58.8% of STEC, respectively. Both LT1 and ST1 genes were positive in 66.7% of ETEC (n=15) while 33.3% of isolates harbor only LT1 gene. The prevalence of multi-antimicrobial-agent resistant E. coli in the river Ganga water poses increased risk of infections in the human population.

  6. Prevalence of multi-antimicrobial-agent resistant, shiga toxin and enterotoxin producing Escherichia coli in surface waters of river Ganga.

    PubMed

    Ram, Siya; Vajpayee, Poornima; Shanker, Rishi

    2007-11-01

    The consumption of polluted surface water for domestic and recreational purposes by large populations in developing nations is a major cause of diarrheal disease related mortality. The river Ganga and its tributaries meet 40% of the water requirement for drinking and irrigation in India. In this study, Escherichia coli isolates (n=75) of the river Ganga water were investigated for resistance to antimicrobial agents (n=15) and virulence genes specific to shiga toxin (STEC) and enterotoxin producing E. coli (ETEC). E. coli isolates from the river Ganga water exhibit resistance to multiple antimicrobial agents. The distribution of antimicrobial agent resistance in E. colivaries significantly (chi2: 81.28 at df = 24, p < 0.001) between the sites. Both stx1 and stx2 genes were present in 82.3% of STEC (n=17) while remaining isolates possess either stxl (11.8%) or stx2 (5.9%). The presence of eaeA, hlyA, and chuA genes was observed in 70.6, 88.2, and 58.8% of STEC, respectively. Both LT1 and ST1 genes were positive in 66.7% of ETEC (n=15) while 33.3% of isolates harbor only LT1 gene. The prevalence of multi-antimicrobial-agent resistant E. coli in the river Ganga water poses increased risk of infections in the human population. PMID:18044515

  7. Enhancing isomaltulose production by recombinant Escherichia coli producing sucrose isomerase: culture medium optimization containing agricultural wastes and cell immobilization.

    PubMed

    Li, Sha; Xu, Hong; Yu, Jianguang; Wang, Yanyuan; Feng, Xiaohai; Ouyang, Pingkai

    2013-10-01

    Isomaltulose is a structural isomer of sucrose commercially used in food industries. In this work, recombinant Escherichia coli producing sucrose isomerase (SIase) was used to convert sucrose into isomaltulose. To develop an economical industrial medium, untreated cane molasses (10.63 g l⁻¹), yeast extract (25.93 g l⁻¹), and corn steep liquor (10.45 g l⁻¹) were used as main culture compositions for SIase production. The relatively high SIase activity (14.50 ± 0.11 U mg DCW⁻¹) was obtained by the recombinant cells. To the best of our knowledge, this is the first investigation on SIase production by engineered E. coli using untreated cane molasses. The recombinant E. coli cells expressing the SIase gene were immobilized in calcium alginate gel in order to improve the efficiency of recycling. The immobilization was most effective with 2 % (w/v) sodium alginate and 3 % (w/v) calcium chloride. The optimal initial biomass for immobilization was 20 % (w/v, wet wt.), with a hardening time of 8 h for cell immobilization. The immobilized E. coli cells exhibited good stability for 30 batches with the productivity of 0.45 g isomaltulose g pellet⁻¹ h⁻¹. A continuous isomaltulose formation process using a column reactor remained stable for 40 days with 83 ± 2 % isomaltulose yield, which would be beneficial for economical production of isomaltulose.

  8. Prevalence and characterization of verotoxin-producing Escherichia coli (VTEC) in cattle from an Ontario abattoir

    PubMed Central

    Karama, Musafiri; Johnson, Roger P; Holtslander, Robert; McEwen, Scott A.; Gyles, Carlton L.

    2008-01-01

    This study determined the prevalence of verotoxin (VT)-producing Escherichia coli (VTEC) in Ontario beef cattle at slaughter and characterized the isolates by serotype, virulence factors, virulence markers, and antimicrobial resistance. Cultures of rectal feces from 500 animals were screened for VT by an enzyme-linked immunosorbent assay (ELISA) and by polymerase chain reaction (PCR) for genes vt1, vt2, and eae. The VT-ELISA-positive samples were tested by a VT-immunoblot to isolate VTEC colonies. The prevalence rates of VTEC by VT-ELISA and PCR were 10.2% [95% confidence interval (CI), 7.8% to 13.2%] and 6.2% (95% CI, 4.4% to 8.7%), respectively. Colonies of VTEC were isolated from 27 (53%) of the 51 VT-ELISA-positive samples and belonged to 24 serotypes, which did not include O157:H7. Twelve of the serotypes have been implicated in disease in humans. Virulence profiling of the isolates by PCR revealed that 2 (8%) were eae-positive, 5 (21%) had vt1 only, and 19 (79%) had vt2, of which 3 had vt2 only, 7 had vt1 + vt2, 4 had vt2 + vt2c, 2 had vt2 + vt2c + vt2d, 2 had vt1 + vt2 + vt2c, and 1 had vt1 + vt2 + vt2c + vt2d. The distribution of selected plasmid-encoded putative virulence genes was as follows: ehxA, 63%; espP, 46%; saa, 67%; and subA, 54%. Nine of the 24 isolates were resistant to 1 or more antimicrobials. Major conclusions are that the VTEC prevalence of 10.2% was among the lower rates reported for beef cattle, a high proportion of the isolates had vt2 genes, the subA gene was reported for the 1st time in Canadian VTEC, and the absence of O157 VTEC likely reflects the use of a technique that detected all VTEC. PMID:18783017

  9. Shiga toxin-producing Escherichia coli in beef retail markets from Argentina.

    PubMed

    Brusa, Victoria; Aliverti, Virginia; Aliverti, Florencia; Ortega, Emanuel E; de la Torre, Julian H; Linares, Luciano H; Sanz, Marcelo E; Etcheverría, Analía I; Padola, Nora L; Galli, Lucía; Peral García, Pilar; Copes, Julio; Leotta, Gerardo A

    2012-01-01

    Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens that cause mild or serious diseases and can lead to people death. This study reports the prevalence and characteristics of STEC O157 and non-O157 in commercial ground beef and environmental samples, including meat table, knife, meat mincing machine, and manipulator hands (n = 450) obtained from 90 retail markets over a nine-month period. The STEC isolates were serotyped and virulence genes as stx (Shiga toxin), rfb(O157)] (O157 lipopolysaccharide), fliC(H7) (H7 flagellin), eae (intimin), ehxA (enterohemolysin) and saa (STEC autoagglutinating adhesin), were determined. STEC O157 were identified in 23 (25.5%) beef samples and 16 (4.4%) environmental samples, while STEC non-O157 were present in 47 (52.2%) and 182 (50.5%), respectively. Among 54 strains isolated, 17 were STEC O157:H7 and 37 were STEC non-O157. The prevalent genotype for O157 was stx(2)/eae/ehxA/fliC(H7) (83.4%), and for STEC non-O157 the most frequent ones were stx(1)/stx(2)/saa/ehxA (29.7%); stx(2) (29.7%); and stx(2)/saa/ehxA (27%). None of the STEC non-O157 strains were eae-positive. Besides O157:H7, other 20 different serotypes were identified, being O8:H19, O178:H19, and O174:H28 the prevalent. Strains belonging to the same serotype could be isolated from different sources of the same retail market. Also, the same serotype could be detected in different stores. In conclusion, screening techniques are increasingly sensitive, but the isolation of STEC non-O157 is still a challenge. Moreover, with the results obtained from the present work, although more studies are needed, cross-contamination between meat and the environment could be suspected. PMID:23346554

  10. Non-O157 Shiga toxin-producing Escherichia coli in U. S. retail ground beef.

    PubMed

    Liao, Yen-Te; Miller, Markus F; Loneragan, Guy H; Brooks, J Chance; Echeverry, Alejandro; Brashears, Mindy M

    2014-07-01

    Shiga toxin-producing Escherichia coli (STEC) serotype O157:H7 and serogroups O26, O45, O103, O111, O121, and O145 are the leading cause of STEC-associated infections in humans in the United States. In the United States, these organisms are considered adulterants in raw nonintact beef products and in intact beef destined to be made into or used in nonintact raw beef products. The objective of this study was to provide an estimate of the burden of the six serogroups of non-O157 STEC in ground beef obtained from retail stores across the United States. A convenience sample of commercial ground beef products (n = 1,129) were purchased from retail stores in 24 states from October 2011 to May 2012. The samples had various lean/fat proportions, muscle group of origin (chuck, round, sirloin, or not specified), and packaging types. For each ground beef sample, 25 g was inoculated in 225 ml of modified tryptic soy broth, stomached for 1 min, and then incubated at 41°C for 18 ± 2 h. These enrichment cultures were then screened for stx, eae, and O group genes using a commercially available, closed-platform PCR-based method. The potential positive samples were subjected to immunomagnetic separation and plated on modified Rainbow agar. Morphologically typical colonies were subjected to latex agglutination and PCR determination of stx and eae genes. Nine (0.8%) of the ground beef samples were potentially positive for at least one STEC serogroup after PCR screening. The serogroups detected by PCR assay were O26 (four samples), O103 (four samples), O145 (three samples), O45 (two samples), and O121 (one sample). No STEC isolates belonging to these serogroups were recovered from the sample cultures. The current research provides updated surveillance data for non-O157 STEC isolates among commercial ground beef products and information regarding the potential sources of contamination from different parts of beef trims destined for ground beef production. PMID:24988027

  11. Non-O157 Shiga toxin-producing Escherichia coli in U. S. retail ground beef.

    PubMed

    Liao, Yen-Te; Miller, Markus F; Loneragan, Guy H; Brooks, J Chance; Echeverry, Alejandro; Brashears, Mindy M

    2014-07-01

    Shiga toxin-producing Escherichia coli (STEC) serotype O157:H7 and serogroups O26, O45, O103, O111, O121, and O145 are the leading cause of STEC-associated infections in humans in the United States. In the United States, these organisms are considered adulterants in raw nonintact beef products and in intact beef destined to be made into or used in nonintact raw beef products. The objective of this study was to provide an estimate of the burden of the six serogroups of non-O157 STEC in ground beef obtained from retail stores across the United States. A convenience sample of commercial ground beef products (n = 1,129) were purchased from retail stores in 24 states from October 2011 to May 2012. The samples had various lean/fat proportions, muscle group of origin (chuck, round, sirloin, or not specified), and packaging types. For each ground beef sample, 25 g was inoculated in 225 ml of modified tryptic soy broth, stomached for 1 min, and then incubated at 41°C for 18 ± 2 h. These enrichment cultures were then screened for stx, eae, and O group genes using a commercially available, closed-platform PCR-based method. The potential positive samples were subjected to immunomagnetic separation and plated on modified Rainbow agar. Morphologically typical colonies were subjected to latex agglutination and PCR determination of stx and eae genes. Nine (0.8%) of the ground beef samples were potentially positive for at least one STEC serogroup after PCR screening. The serogroups detected by PCR assay were O26 (four samples), O103 (four samples), O145 (three samples), O45 (two samples), and O121 (one sample). No STEC isolates belonging to these serogroups were recovered from the sample cultures. The current research provides updated surveillance data for non-O157 STEC isolates among commercial ground beef products and information regarding the potential sources of contamination from different parts of beef trims destined for ground beef production.

  12. Prevalence of Class D Carbapenemases among Extended-Spectrum β-Lactamases Producing Escherichia coli Isolates from Educational Hospitals in Shahrekord

    PubMed Central

    Damavandi, Mohammad-Sadegh; Latif Pour, Mohammad

    2016-01-01

    Introduction Extended-spectrum β-lactamases (ESBLs) are a set of plasmid-borne, various and quickly evolving enzymes that are a main therapeutic issue now-a-days for inpatient and outpatient treatment. Aim The aim of this study was to determine multi-drug resistance (MDR) and ESBLs producing E. coli strains, prevalence of class D Carbapenemases among ESBLs producing Escherichia coli isolates from educational hospitals in Shahrekord, Iran. Materials and Methods Uropathogenic Escherichia coli strains were isolated from patients with Urinary Tract Infections (UTIs). The agar disc diffusion test was used to characterize the antimicrobial sensitivity of the E. coli isolates. The ESBL positive strains were identified by phenotypic double-disk synergy test, by third-generation cephalosporin in combination with or without clavulanic acid. Multiplex PCR was carried out for detection of the three families of OXA-type carbapenamases including OXA-23, OXA-24, and OXA-48 in E. coli strains. Results All bacterial isolates were susceptible to meropenem. Ninety isolates produced ESBL, 55 E. coli isolates from inpatients, and 35 isolates from outpatients, with a significant association (p< 0.05). The prevalence of OXA-23, OXA-24, and OXA-48 in the ESBLs producing isolates was respectively 21%, 18%, and 11% for inpatients, and 10%, 8%, and 6% for outpatients. Conclusion ESBL-producing E. coli isolates are also a major threat in the clinical setting. The findings of this study indicated the high occurrence of ESBLs and multiple antibiotic resistance in E. coli isolates. PMID:27462579

  13. Production and regulation of functional amyloid curli fimbriae by Shiga toxin-producing Escherichia coli

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Functional amyloid, in the form of adhesive fimbrial proteins termed curli, was first described in Salmonella and Escherichia coli. Curli fibers adhere to various host cells and structural proteins, interact with components of the host immune system, and participate in biofilm formation. Shiga toxin...

  14. Characterization of enterotoxigenic E. coli (ETEC), Shiga-toxin producing E. coli (STEC) and necrotoxigenic E. coli (NTEC) isolated from diarrhoeic Mediterranean water buffalo calves (Bubalus bubalis).

    PubMed

    Borriello, G; Lucibelli, M G; De Carlo, E; Auriemma, C; Cozza, D; Ascione, G; Scognamiglio, F; Iovane, G; Galiero, G

    2012-08-01

    Two hundred and twenty Escherichia coli isolates from 314 Mediterranean water buffalo calves less than 4 weeks old affected by severe diarrhoea with a lethal outcome were characterized for the presence of the virulence factors LT, ST, Stx1, Stx2, haemolysins, intimin, CNF1, CNF2, CDT-I, CDT-II, CDT-III, CDT-IV, and F17-related fimbriae (F17a, F17b, F17c, F17d). The prevalence of ETEC, STEC and NTEC were 1.8%, 6.8% and 20.9%, respectively. The ETEC isolates were all LT-positive and ST-negative. The STEC isolates were all Stx and intimin-positive, with Stx1 (80%) more frequent than Stx2 (27%). The NTEC isolates were all CNF and Hly-positive, with CNF2 (83%) more frequent than CNF1 (22%). Susceptibility assays to 11 antimicrobials displayed high rates of resistance (>30%) to antimicrobials tested. These data show that the most prevalent strains in diarrhoeic water buffalo calves were NTEC, mostly CNF2 and HlyA-positive, with strong associations CNF2/CDT-III and CNF2/F17c. PMID:21658736

  15. Characterization of enterotoxigenic E. coli (ETEC), Shiga-toxin producing E. coli (STEC) and necrotoxigenic E. coli (NTEC) isolated from diarrhoeic Mediterranean water buffalo calves (Bubalus bubalis).

    PubMed

    Borriello, G; Lucibelli, M G; De Carlo, E; Auriemma, C; Cozza, D; Ascione, G; Scognamiglio, F; Iovane, G; Galiero, G

    2012-08-01

    Two hundred and twenty Escherichia coli isolates from 314 Mediterranean water buffalo calves less than 4 weeks old affected by severe diarrhoea with a lethal outcome were characterized for the presence of the virulence factors LT, ST, Stx1, Stx2, haemolysins, intimin, CNF1, CNF2, CDT-I, CDT-II, CDT-III, CDT-IV, and F17-related fimbriae (F17a, F17b, F17c, F17d). The prevalence of ETEC, STEC and NTEC were 1.8%, 6.8% and 20.9%, respectively. The ETEC isolates were all LT-positive and ST-negative. The STEC isolates were all Stx and intimin-positive, with Stx1 (80%) more frequent than Stx2 (27%). The NTEC isolates were all CNF and Hly-positive, with CNF2 (83%) more frequent than CNF1 (22%). Susceptibility assays to 11 antimicrobials displayed high rates of resistance (>30%) to antimicrobials tested. These data show that the most prevalent strains in diarrhoeic water buffalo calves were NTEC, mostly CNF2 and HlyA-positive, with strong associations CNF2/CDT-III and CNF2/F17c.

  16. Occurrence of ESBL-Producing Escherichia coli in Livestock and Farm Workers in Mecklenburg-Western Pomerania, Germany

    PubMed Central

    Dahms, Carmen; Hübner, Nils-Olaf; Kossow, Annelene; Mellmann, Alexander; Dittmann, Kathleen; Kramer, Axel

    2015-01-01

    In recent years, extended-spectrum β-lactamases (ESBL) producing bacteria have been found in livestock, mainly as asymptomatic colonizers. The zoonotic risk for people working in close contact to animal husbandry has still not been completely assessed. Therefore, we investigated the prevalence of ESBL-producing Escherichia spp. in livestock animals and workers to determine the potential risk for an animal-human cross-transmission.In Mecklenburg-Western Pomerania, northeast Germany, inguinal swabs of 73 individuals with livestock contact from 23 different farms were tested for ESBL-producing Escherichia spp. Two pooled fecal samples per farm of animal origin from 34 different farms (17 pig farms, 11 cattle farms, 6 poultry farms) as well as cloacal swabs of 10 randomly selected broilers or turkeys were taken at each poultry farm. For identification, selective chromogenic agar was used after an enrichment step. Phenotypically ESBL-producing isolates (n = 99) were tested for CTX-M, OXA, SHV and TEM using PCR, and isolates were further characterized using multilocus sequence typing (MLST). In total, 61 diverse isolates from different sources and/or different MLST/PCR results were acquired. Five farm workers (three from cattle farms and two from pig farms) harbored ESBL-producing E. coli. All human isolates harbored the CTX-M β-lactamase; TEM and OXA β-lactamases were additionally detected in two, resp. one, isolates. ESBL-producing Escherichia spp. were found in fecal samples at pig (15/17), cattle (6/11) and poultry farms (3/6). In total, 70.6% (24/36) of the tested farms were ESBL positive. Furthermore, 9 out of 60 cloacal swabs turned out to be ESBL positive. All isolated ESBL-producing bacteria from animal sources were E. coli, except for one E. hermanii isolate. CTX-M was the most prevalent β-lactamase at cattle and pig farms, while SHV predominated in poultry. One human isolate shared an identical MLST sequence type (ST) 3891 and CTX-M allele to the isolate

  17. Long-Term Sentinel Surveillance for Enterotoxigenic Escherichia coli and Non-O157 Shiga Toxin-Producing E. coli in Minnesota

    PubMed Central

    Medus, Carlota; Besser, John M.; Juni, Billie A.; Koziol, Bonnie; Lappi, Victoria; Smith, Kirk E.; Hedberg, Craig W.

    2016-01-01

    Background. Enterotoxigenic Escherichia coli (ETEC) and non-O157 Shiga toxin-producing E. coli (STEC) are not detected by conventional culture methods. The prevalence of ETEC infections in the United States is unknown, and recognized cases are primarily associated with foreign travel. Gaps remain in our understanding of STEC epidemiology. Methods. Two sentinel surveillance sites were enrolled: an urban health maintenance organization laboratory (Laboratory A) and a rural hospital laboratory (Laboratory B). Residual sorbitol MacConkey (SMAC) plates from stool cultures performed at Laboratory A (1996–2006) and Laboratory B (2000–2008) were collected. Colony sweeps from SMAC plates were tested for genes encoding STEC toxins stx1 and stx2 (1996–2008) and ETEC heat-labile and heat-stable toxins eltB, estA 1, 2 and 3 (2000–2008) by polymerase chain reaction (PCR)-based assays. Results. In Laboratory A, a bacterial pathogen was identified in 7.0% of 21 970 specimens. During 1996–2006, Campylobacter was the most common bacterial pathogen (2.7% of cultures), followed by Salmonella (1.2%), Shigella (1.0%), and STEC (0.9%). Among STEC (n = 196), O157 was the most common serogroup (31%). During 2000–2006, ETEC (1.9%) was the second most common bacterial pathogen after Campylobacter (2.6%). In Laboratory B, of 19 293 specimens tested, a bacterial pathogen was identified for 5.5%, including Campylobacter (2.1%), STEC (1.3%), Salmonella (1.0%), and ETEC (0.8%). Among STEC (n = 253), O157 was the leading serogroup (35%). Among ETEC cases, 61% traveled internationally. Conclusions. Enterotoxigenic E. coli and STEC infections were as common as most other enteric bacterial pathogens, and ETEC may be detected more frequently by culture-independent multiplex PCR diagnostic methods. A high proportion of ETEC cases were domestically acquired. PMID:26913288

  18. Characteristics of CTX-M Extended-Spectrum β-Lactamase-Producing Escherichia coli Strains Isolated from Multiple Rivers in Southern Taiwan

    PubMed Central

    Chen, Po-An; Hung, Chih-Hsin; Huang, Ping-Chih; Chen, Jung-Ren; Huang, I-Fei; Chen, Wan-Ling; Chiou, Yee-Hsuan; Hung, Wan-Yu

    2016-01-01

    Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli sequence type ST131 has emerged as the leading cause of community-acquired urinary tract infections and bacteremia worldwide. Whether environmental water is a potential reservoir of these strains remains unclear. River water samples were collected from 40 stations in southern Taiwan from February to August 2014. PCR assay and multilocus sequence typing (MLST) analysis were conducted to determine the CTX-M group and sequence type, respectively. In addition, we identified the seasonal frequency of ESBL-producing E. coli strains and their geographical relationship with runoffs from livestock and poultry farms between February and August 2014. ESBL-producing E. coli accounted for 30% of the 621 E. coli strains isolated from river water in southern Taiwan. ESBL-producing E. coli ST131 was not detected among the isolates. The most commonly detected strain was E. coli CTX-M group 9. Among the 92 isolates selected for MLST analysis, the most common ESBL-producing clonal complexes were ST10 and ST58. The proportion of ESBL-producing E. coli was significantly higher in areas with a lower river pollution index (P = 0.025) and regions with a large number of chickens being raised (P = 0.013). ESBL-producing E. coli strains were commonly isolated from river waters in southern Taiwan. The most commonly isolated ESBL-producing clonal complexes were ST10 and ST58, which were geographically related to chicken farms. ESBL-producing E. coli ST131, the major clone causing community-acquired infections in Taiwan and worldwide, was not detected in river waters. PMID:26773082

  19. Surveillance of ESBL producing multidrug resistant Escherichia coli in a teaching hospital in India

    PubMed Central

    Rath, Shakti; Dubey, Debasmita; Sahu, Mahesh C.; Padhy, Rabindra N

    2014-01-01

    Objective To record nosocomial and community-acquired accounts of antibiotic resistance in Escherichia coli (E. coli) strains, isolated from clinical samples of a teaching hospital by surveillance, over a period of 39 months (November 2009-January 2013). Methods Clinical samples from nosocomial sources, i.e., wards and cabins, intensive care unit (ICU) and neonatal intensive care unit (NICU), and community (outpatient department, OPD) sources of the hospital, were used for isolating strains of E. coli, which were subjected for testing for production of ‘extended spectrum beta-lactamase’-(ESBL) enzyme as well as determining antibiotic sensitivity pattern with 23 antibiotics. Results Of the total 1642 (100%) isolates, 810 (49.33%) strains were from OPD and 832 (50.66%) were from hospital settings. Occurrence of infectious E. coli strains increased in a mathematical progression in community sources, but in nosocomial infections, such values remained almost constant in each quarter. A total of 395 (24.05%) ESBL strains were isolated from the total 810 isolates of community; of the total of 464 (28.25%) isolates of wards and cabins, 199 (12.11%) were ESBL strains; and among the total of 368 (22.41%) isolates of ICU and NICU, ESBLs were 170 (10.35%); the total nosocomial ESBL isolates, 369 (22.47%) were from the nosocomial total of 832 (50.66%) isolates. Statistically, it was confirmed that ESBL strains were equally distributed in community or hospital units. Antibiogram of 23 antibiotics revealed progressive increases of drug-resistance against each antibiotic with the maximum resistance values were recorded against gentamicin: 92% and 79%, oxacillin: 94% and 69%, ceftriaxone: 85% and 58%, and norfloxacin 97% and 69% resistance, in nosocomial and community isolates, respectively. Conclusions This study revealed the daunting state of occurrence of multidrug resistant E. coli and its infection dynamics in both community and hospital settings.

  20. Feed Fermentation with Reuteran- and Levan-Producing Lactobacillus reuteri Reduces Colonization of Weanling Pigs by Enterotoxigenic Escherichia coli

    PubMed Central

    Yang, Yan; Galle, Sandra; Le, Minh Hong Anh; Zijlstra, Ruurd T.

    2015-01-01

    This study determined the effect of feed fermentation with Lactobacillus reuteri on growth performance and the abundance of enterotoxigenic Escherichia coli (ETEC) in weanling piglets. L. reuteri strains produce reuteran or levan, exopolysaccharides that inhibit ETEC adhesion to the mucosa, and feed fermentation was conducted under conditions supporting exopolysaccharide formation and under conditions not supporting exopolysaccharide formation. Diets were chosen to assess the impact of organic acids and the impact of viable L. reuteri bacteria. Fecal samples were taken throughout 3 weeks of feeding; at the end of the 21-day feeding period, animals were euthanized to sample the gut digesta. The feed intake was reduced in pigs fed diets containing exopolysaccharides; however, feed efficiencies did not differ among the diets. Quantification of L. reuteri by quantitative PCR (qPCR) detected the two strains used for feed fermentation throughout the intestinal tract. Quantification of E. coli and ETEC virulence factors by qPCR demonstrated that fermented diets containing reuteran significantly (P < 0.05) reduced the copy numbers of genes for E. coli and the heat-stable enterotoxin in feces compared to those achieved with the control diet. Any fermented feed significantly (P < 0.05) reduced the abundance of E. coli and the heat-stable enterotoxin in colonic digesta at 21 days; reuteran-containing diets reduced the copy numbers of the genes for E. coli and the heat-stable enterotoxin below the detection limit in samples from the ileum, the cecum, and the colon. In conclusion, feed fermentation with L. reuteri reduced the level of colonization of weaning piglets with ETEC, and feed fermentation supplied concentrations of reuteran that may specifically contribute to the effect on ETEC. PMID:26070673

  1. Feed Fermentation with Reuteran- and Levan-Producing Lactobacillus reuteri Reduces Colonization of Weanling Pigs by Enterotoxigenic Escherichia coli.

    PubMed

    Yang, Yan; Galle, Sandra; Le, Minh Hong Anh; Zijlstra, Ruurd T; Gänzle, Michael G

    2015-09-01

    This study determined the effect of feed fermentation with Lactobacillus reuteri on growth performance and the abundance of enterotoxigenic Escherichia coli (ETEC) in weanling piglets. L. reuteri strains produce reuteran or levan, exopolysaccharides that inhibit ETEC adhesion to the mucosa, and feed fermentation was conducted under conditions supporting exopolysaccharide formation and under conditions not supporting exopolysaccharide formation. Diets were chosen to assess the impact of organic acids and the impact of viable L. reuteri bacteria. Fecal samples were taken throughout 3 weeks of feeding; at the end of the 21-day feeding period, animals were euthanized to sample the gut digesta. The feed intake was reduced in pigs fed diets containing exopolysaccharides; however, feed efficiencies did not differ among the diets. Quantification of L. reuteri by quantitative PCR (qPCR) detected the two strains used for feed fermentation throughout the intestinal tract. Quantification of E. coli and ETEC virulence factors by qPCR demonstrated that fermented diets containing reuteran significantly (P < 0.05) reduced the copy numbers of genes for E. coli and the heat-stable enterotoxin in feces compared to those achieved with the control diet. Any fermented feed significantly (P < 0.05) reduced the abundance of E. coli and the heat-stable enterotoxin in colonic digesta at 21 days; reuteran-containing diets reduced the copy numbers of the genes for E. coli and the heat-stable enterotoxin below the detection limit in samples from the ileum, the cecum, and the colon. In conclusion, feed fermentation with L. reuteri reduced the level of colonization of weaning piglets with ETEC, and feed fermentation supplied concentrations of reuteran that may specifically contribute to the effect on ETEC. PMID:26070673

  2. Feed Fermentation with Reuteran- and Levan-Producing Lactobacillus reuteri Reduces Colonization of Weanling Pigs by Enterotoxigenic Escherichia coli.

    PubMed

    Yang, Yan; Galle, Sandra; Le, Minh Hong Anh; Zijlstra, Ruurd T; Gänzle, Michael G

    2015-09-01

    This study determined the effect of feed fermentation with Lactobacillus reuteri on growth performance and the abundance of enterotoxigenic Escherichia coli (ETEC) in weanling piglets. L. reuteri strains produce reuteran or levan, exopolysaccharides that inhibit ETEC adhesion to the mucosa, and feed fermentation was conducted under conditions supporting exopolysaccharide formation and under conditions not supporting exopolysaccharide formation. Diets were chosen to assess the impact of organic acids and the impact of viable L. reuteri bacteria. Fecal samples were taken throughout 3 weeks of feeding; at the end of the 21-day feeding period, animals were euthanized to sample the gut digesta. The feed intake was reduced in pigs fed diets containing exopolysaccharides; however, feed efficiencies did not differ among the diets. Quantification of L. reuteri by quantitative PCR (qPCR) detected the two strains used for feed fermentation throughout the intestinal tract. Quantification of E. coli and ETEC virulence factors by qPCR demonstrated that fermented diets containing reuteran significantly (P < 0.05) reduced the copy numbers of genes for E. coli and the heat-stable enterotoxin in feces compared to those achieved with the control diet. Any fermented feed significantly (P < 0.05) reduced the abundance of E. coli and the heat-stable enterotoxin in colonic digesta at 21 days; reuteran-containing diets reduced the copy numbers of the genes for E. coli and the heat-stable enterotoxin below the detection limit in samples from the ileum, the cecum, and the colon. In conclusion, feed fermentation with L. reuteri reduced the level of colonization of weaning piglets with ETEC, and feed fermentation supplied concentrations of reuteran that may specifically contribute to the effect on ETEC.

  3. Isolation and Characterization of Verocytotoxin-Producing Escherichia coli O157 Strains from Dutch Cattle and Sheep

    PubMed Central

    Heuvelink, A. E.; van den Biggelaar, F. L. A. M.; de Boer, E.; Herbes, R. G.; Melchers, W. J. G.; Huis In ’t Veld, J. H. J.; Monnens, L. A. H.

    1998-01-01

    In the periods from July to November 1995 and 1996, fecal samples from Dutch cattle and sheep were collected at the main slaughterhouses of The Netherlands, located at different geographic sites. The samples were examined for the presence of verocytotoxin (VT)-producing Escherichia coli (VTEC) of serogroup O157. E. coli O157 strains could be isolated from 57 (10.6%) of 540 adult cattle, 2 (0.5%) of 397 veal calves, 2 (3.8%) of 52 ewes, and 2 (4.1%) of 49 lambs. Immunomagnetic separation with O157-specific-antibody-coated beads appeared to be significantly more sensitive than conventional plating for detection of the organism in feces. With the exception of two isolates from adult cattle which appeared to be negative for VT genes, all animal isolates were positive for both VT (VT1 and/or VT2) and E. coli attaching-and-effacing gene sequences, and therefore, they were regarded as potential human pathogens. Although genomic typing by pulsed-field gel electrophoresis revealed a wide variety of distinct restriction patterns, comparison of the 63 animal isolates with 33 fecal O157 VTEC strains previously isolated from humans with the diarrhea-associated form of the hemolytic-uremic syndrome by their phage types and VT genotypes showed a marked similarity between animal and human isolates: 30 (90.9%) of the 33 human isolates appeared to be of E. coli O157 strain types also isolated from cattle and sheep. It was concluded that Dutch cattle and sheep are an important reservoir of E. coli O157 strains that are potentially pathogenic for humans. PMID:9542902

  4. Isolation and characterization of verocytotoxin-producing Escherichia coli O157 strains from Dutch cattle and sheep.

    PubMed

    Heuvelink, A E; van den Biggelaar, F L; de Boer, E; Herbes, R G; Melchers, W J; Huis in 't Veld, J H; Monnens, L A

    1998-04-01

    In the periods from July to November 1995 and 1996, fecal samples from Dutch cattle and sheep were collected at the main slaughterhouses of The Netherlands, located at different geographic sites. The samples were examined for the presence of verocytotoxin (VT)-producing Escherichia coli (VTEC) of serogroup 0157. E. coli O157 strains could be isolated from 57 (10.6%) of 540 adult cattle, 2 (0.5%) of 397 veal calves, 2 (3.8%) of 52 ewes, and 2 (4.1%) of 49 lambs. Immunomagnetic separation with O157-specific-antibody-coated beads appeared to be significantly more sensitive than conventional plating for detection of the organism in feces. With the exception of two isolates from adult cattle which appeared to be negative for VT genes, all animal isolates were positive for both VT (VT1 and/or VT2) and E. coli attaching-and-effacing gene sequences, and therefore, they were regarded as potential human pathogens. Although genomic typing by pulsed-field gel electrophoresis revealed a wide variety of distinct restriction patterns, comparison of the 63 animal isolates with 33 fecal O157 VTEC strains previously isolated from humans with the diarrhea-associated form of the hemolytic-uremic syndrome by their phage types and VT genotypes showed a marked similarity between animal and human isolates: 30 (90.9%) of the 33 human isolates appeared to be of E. coli O157 strain types also isolated from cattle and sheep. It was concluded that Dutch cattle and sheep are an important reservoir of E. coli O157 strains that are potentially pathogenic for humans. PMID:9542902

  5. Updated molecular epidemiology of carbapenem-non-susceptible Escherichia coli in Taiwan: first identification of KPC-2 or NDM-1-producing E. coli in Taiwan

    PubMed Central

    2013-01-01

    Background The global spread and increasing incidence of carbapenem-resistant Enterobacteriaceae have resulted in treatment and public health concerns. Here, we present an investigation of the molecular mechanisms and clonality of carbapenem-non-susceptible Escherichia coli (CnSEC) based on a nationwide survey in Taiwan. Methods We collected 32 and 43 carbapenem-non-susceptible E. coli isolates in 2010 and 2012, respectively. The genes encoding cabapenemases and plasmidic AmpC-type and extended-spectrum β-lactamases (EBSLs) were analyzed by polymerase chain reaction (PCR). The major porin channels OmpF and OmpC were evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Molecular typing was performed with pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). Results The resistance rates of CnSEC isolates to cefazolin, cefotaxime, cefoxitin, ceftazidime, and ertapenem were all 100%, and most (94.7%) isolates were CMY producers. The main mechanism of CnSEC in Taiwan is via plasmidic AmpC β-lactamase CMY-2 and DHA-1 in combination with the loss of OmpC/F. In 2010, one isolate was confirmed to harbor blaIMP-8; a KPC-2 producer and an NDM-1 producer were detected in 2012. No isolate had VIM- or OXA-carbapenemases. ST131 was the predominant ST type (33.3%). PFGE revealed no large cluster in CnSEC isolates in Taiwan. Conclusions The co-existence of plasmidic AmpC β-lactamase and outer membrane protein loss is the main mechanism for CnSEC in Taiwan. The emergence of KPC-2 and NDM-1 in 2012 and the predominance of ST131 warrant close monitoring and infection control. PMID:24354657

  6. An optical biosensor for detection of pathogen biomarkers from Shiga toxin-producing Escherichia coli in ground beef samples

    NASA Astrophysics Data System (ADS)

    Lamoureux, Loreen; Adams, Peter; Banisadr, Afsheen; Stromberg, Zachary; Graves, Steven; Montano, Gabriel; Moxley, Rodney; Mukundan, Harshini

    2014-03-01

    Shiga toxin-producing Escherichia coli (STEC) poses a serious threat to human health through the consumption of contaminated food products, particularly beef and produce. Early detection in the food chain, and discrimination from other non-pathogenic Escherichia coli (E. coli), is critical to preventing human outbreaks, and meeting current agricultural screening standards. These pathogens often present in low concentrations in contaminated samples, making discriminatory detection difficult without the use of costly, time-consuming methods (e.g. culture). Using multiple signal transduction schemes (including novel optical methods designed for amphiphiles), specific recognition antibodies, and a waveguide-based optical biosensor developed at Los Alamos National Laboratory, we have developed ultrasensitive detection methods for lipopolysaccharides (LPS), and protein biomarkers (Shiga toxin) of STEC in complex samples (e.g. beef lysates). Waveguides functionalized with phospholipid bilayers were used to pull down amphiphilic LPS, using methods (membrane insertion) developed by our team. The assay format exploits the amphiphilic biochemistry of lipoglycans, and allows for rapid, sensitive detection with a single fluorescent reporter. We have used a combination of biophysical methods (atomic force and fluorescence microscopy) to characterize the interaction of amphiphiles with lipid bilayers, to efficiently design these assays. Sandwich immunoassays were used for detection of protein toxins. Biomarkers were spiked into homogenated ground beef samples to determine performance and limit of detection. Future work will focus on the development of discriminatory antibodies for STEC serotypes, and using quantum dots as the fluorescence reporter to enable multiplex screening of biomarkers.

  7. Aggregative adherence fimbriae I (AAF/I) mediate colonization of fresh produce and abiotic surface by Shiga toxigenic enteroaggregative Escherichia coli O104:H4

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Shiga toxigenic Escherichia coli O104:H4 bares the characteristics of both enterohemorrhagic (EHEC) and enteroaggregative (EAEC) E. coli. It produces plasmid encoded aggregative adherence fimbriae I (AAF/I) which mediate cell aggregation and biofilm formation in human intestine and promote Shiga...

  8. Use of photopolymerization for the rapid and cost-effective identification of Shiga toxin-producing Escherichia coli on DNA microarrays

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing Escherichia coli O157:H7 is a leading cause of foodborne illness worldwide. To evaluate better methods to rapidly detect and genotype E. coli O157 virulent strains, the present study explored the use of photopolymerization, a colorimetric and photoinduced signal amplification d...

  9. High prevalence of CTX-M-15-producing O25b-ST131 Escherichia coli clone in Bulgarian hospitals.

    PubMed

    Markovska, Rumyana; Schneider, Ines; Ivanova, Dobrinka; Keuleyan, Emma; Stoeva, Temenuga; Sredkova, Mariya; Markova, Boyka; Bojkova, Kalina; Gergova, Raina; Bauernfeind, Adolf; Mitov, Ivan

    2012-08-01

    According to the European Antimicrobial Resistance Surveillance System project results, Bulgaria has become one of the European countries with dramatically increasing rates of extended-spectrum beta-lactamase (ESBL) producers. The aim of this work was to investigate the epidemiology of ESBL-producing Escherichia coli clinical isolates in Bulgaria, collected from seven clinical centers in three towns, during two study periods: 2002-2003 and 2006-2009. For 193 ESBL-producing E. coli isolates random amplified polymorphic DNA (RAPD) analyses, phylogenetic typing, and screening for O25b-ST131 isolates were carried out. Antimicrobial susceptibility, ESBL-type and transferability of resistance determinants were analyzed. Four different ESBL-types, namely TEM-139, SHV-12, CTX-M-3, and CTX-M-15 were found. CTX-M-15 dominated, being found in 88% of the isolates. RAPD-typing revealed 35 types, among which type A dominated, comprising 65% of the isolates. Sixty-eight percent of the 193 isolates belonged to the O25b-ST131 clone, to the phylogenetic group B2, mostly showed RAPD-type A (92%) and were found in all participating hospitals. O25b-ST131 isolates predominantly produced CTX-M-15 (96%), and less SHV-12 (n=3) or TEM-139 (n=2). In conclusion, this study demonstrated for the first time the country-wide dissemination of a highly resistant B2 O25b-ST131 CTX-M-15 producing E. coli clone in Bulgaria.

  10. Prevalence and Antibiotic Susceptibility Patterns of Extended-Spectrum ß-Lactamase and Metallo-ß-Lactamase-Producing Uropathogenic Escherichia coli Isolates.

    PubMed

    Ghadiri, Hamed; Vaez, Hamid; Razavi-Azarkhiavi, Kamal; Rezaee, Ramin; Haji-Noormohammadi, Mehdi; Rahimi, Ali Asghar; Vaez, Vahid; Kalantar, Enayatollah

    2014-01-01

    Healthcare professionals worldwide have expressed concern over infections by extended-spectrum ß-lactamase (ESBL) and metallo-ß-lactamase (MBL)-producing bacteria. We evaluated the prevalence of ESBL- and MBL-producing Escherichia coli (E. coli) isolated from community-acquired urinary tract infections (UTIs) and their antibiotic-resistance profiles at 3 private laboratories in Tehran, Iran. E. coli isolates were mostly susceptible to meropenem (90.4%) and imipenem (90.0%), followed by amikacin (89.0%) and gentamicin (84.7%). Moreover, we detected that, of the E. coli isolates, 67 (22.3%) were ESBL producers and 21 (7.0%) of E. coli isolates were MBL positive via the imipenem-ethylenediaminetetraacetic acid (EDTA) combined disc test. This report is the first, to our knowledge, on the prevalence of MBL-producing uropathogenic E. coli (UPEC) strains in Iran. The antibiotic resistance of E. coli isolates revealed that 122 (40.7%) were multidrug resistant. The high number of antibiotic-resistant and ß-lactamase-producing UPEC strains necessitates further attention and consideration, particularly MBL-producing strains.

  11. Prevalence and Risk Factors associated with Extended Spectrum Beta Lactamase Producing Escherichia coli and Klebsiella pneumoniae Isolates in Hospitalized Patients in Kashan (Iran)

    PubMed Central

    Sharif, Mohammad Reza; Soltani, Babak; Moravveji, Alireza; Erami, Mahzad; Soltani, Nika

    2016-01-01

    Introduction Production of extended spectrum beta lactamase (ESBL) is an important mechanism of antimicrobial resistance in Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae) isolates. This study was performed to determine the prevalence and risk factors associated with ESBL producing strains of E. coli and K. pneumoniae. Methods In this cross-sectional study, 250 strains (134 E. coli and 116 K. pneumoniae) were obtained, and ESBL producing isolates were detected by the combination disk test in Shahid Beheshti Hospital in Kashan, Iran, from February 2012 to June 2013. Antimicrobial resistance was screened by the disk diffusion method and was confirmed by E-test. Furthermore, risk factors of ESBL producing E. coli and K. pneumoniae microorganisms were determined. Data were analyzed by SPSS version 16, using descriptive statistics, chi-squared, independent-samples t-test, and logistic regression analysis. Results One hundred and two (40.8%) of all strains were ESBL producers, of which 54 (52.9%) were E. coli and 48 (47.1%) were K. pneumoniae (p = 0.86). Furthermore, 40.3% of E. coli and 41.4% of K. pneumoniae isolates were ESBL producers (p = 0.86). The most antimicrobial resistance was to ampicillin, and no imipenem resistance was detected. Risk factors for ESBL producing E. coli included admission duration exceeding 7 days (p = 0.011) and antibiotic use in the last month (p < 0.001), and the associated risk factor for ESBL producing K. pneumoniae was antibiotic use during the recent month (p = 0.002). Conclusion This study identified a relatively high prevalence of ESBL production among E. coli and K. pneumoniae strains. Furthermore, anti-bimicrobial use and admission duration were risk factors for ESBL producing isolates. Therefore, more comprehensive investigations are needed for the development of new strategies to control the dissemination of these microbes. PMID:27123215

  12. Hydrogen-producing Escherichia coli strains overexpressing lactose permease: FT-IR analysis of the lactose-induced stress.

    PubMed

    Grube, Mara; Dimanta, Ilze; Gavare, Marita; Strazdina, Inese; Liepins, Janis; Juhna, Talis; Kalnenieks, Uldis

    2014-01-01

    The lactose permease gene (lacY) was overexpressed in the septuple knockout mutant of Escherichia coli, previously engineered for hydrogen production from glucose. It was expected that raising the lactose transporter activity would elevate the intracellular lactose concentration, inactivate the lactose repressor, induce the lactose operon, and as a result stimulate overall lactose consumption and conversion. However, overexpression of the lactose transporter caused a considerable growth delay in the recombinant strain on lactose, resembling to some extent the "lactose killing" phenomenon. Therefore, the recombinant strain was subjected to selection on lactose-containing media. Selection on plates with 3% lactose yielded a strain with a decreased content of the recombinant plasmid but with an improved ability to grow and produce hydrogen on lactose. Macromolecular analysis of its biomass by means of Fourier transform-infrared spectroscopy demonstrated that increase of the cellular polysaccharide content might contribute to the adaptation of E. coli to lactose stress. PMID:23725289

  13. Co-expression of ferrochelatase allows for complete heme incorporation into recombinant proteins produced in E. coli

    PubMed Central

    Sudhamsu, Jawahar; Kabir, Mariam; Airola, Michael V.; Patel, Bhumit A.; Yeh, Syun-Ru; Rousseau, Dennis L.; Crane, Brian R.

    2010-01-01

    Over-expression of heme binding proteins in E. coli often results in sub-optimal heme incorporation and the amount of heme-bound protein produced usually varies with the protein of interest. Complete heme incorporation is important for biochemical characterization, spectroscopy, structural studies, and for the production of homogeneous commercial proteins with high activity. We have determined that recombinant proteins expressed in E. coli often contain less than a full complement of heme because they rather are partially incorporated with free-base porphyrin. Porphyrin-incorporated proteins have similar spectral characteristics as the desired heme-loaded targets, and thus are difficult to detect, even in purified samples. We present a straightforward and inexpensive solution to this problem that involves the co-expression of native ferrochelatase with the protein of interest. The method is shown to be effective for proteins that contain either Cys- or His- ligated hemes. PMID:20303407

  14. Shiga toxin-producing Escherichia coli isolated from chicken meat in Iran: serogroups, virulence factors, and antimicrobial resistance properties.

    PubMed

    Momtaz, Hassan; Jamshidi, Alireza

    2013-05-01

    The aim of the current study was to determine the virulence factors, serogroups, and antibiotic resistance properties of Shiga toxin-producing Escherichia coli isolated from chicken meat samples. A total of 422 chicken meat samples were collected from 5 townships of Iran. Specimens were immediately transferred to the laboratory in a cooler with an ice pack. Samples were cultured, and the positive culture samples were analyzed by PCR assays. Finally, the antimicrobial susceptibility test was performed using the disk diffusion method in Mueller-Hinton agar. According to the results, out of 422 samples, 146 (34.59%) were confirmed to be E. coli positive and among E. coli-positive samples, 51 (34.93%) and 31 (21.23%) were from attaching and effacing E. coli (AEEC) and enterohemorrhagic E. coli (EHEC) subgroups, respectively. All of the EHEC-positive samples had all stx1, eaeA, and ehly virulence genes, whereas only 5 (9.80%) of AEEC subgroup had all stx1, stx2, and eaeA genes. As the data revealed, O157 was the most prevalent and O111 was the least prevalent strains in the Shiga toxin-producing E. coli (STEC) population. Among STEC strains, sulI and blaSHV had the highest and lowest incidence rate, respectively. There was a high resistance to tetracycline (76.82%), followed by chloramphenicol (73.17%) and nitrofurantoin (63.41%), but there was low resistance to cephalotine (7.31%) antibiotics in isolated strains. Results shows that the PCR technique has a high performance for detection of serogroups, virulence genes, and antibiotic resistance genes in STEC strains. This study is the first prevalence report of detection of virulence genes, serogroups, and antibiotic resistance properties of STEC strains isolated from chicken meat samples in Iran. Based on the results, chicken meat is one of the main sources of STEC strains and its virulence factors in Iran, so an accurate meat inspection would reduce disease outbreaks.

  15. Direct ertapenem disk screening method for identification of KPC-producing Klebsiella pneumoniae and Escherichia coli in surveillance swab specimens.

    PubMed

    Lolans, Karen; Calvert, Karen; Won, Sarah; Clark, James; Hayden, Mary K

    2010-03-01

    Klebsiella pneumoniae carbapenemase (KPC) production in Gram-negative bacilli is an increasing problem worldwide. Rectal swab surveillance is recommended as a component of infection prevention programs, yet few screening methods are published. We compared detection of KPC-producing Klebsiella pneumoniae and Escherichia coli in surveillance specimens by 2 methods: (i) inoculation of swabs in tryptic soy broth containing 2 microg/ml imipenem followed by plating to MacConkey agar (MAC) (method 1) and (ii) streaking swabs on MAC onto which a 10-microg ertapenem disk was then placed (method 2). Simulated rectal swab specimens of challenge isolates from a collection of well-characterized K. pneumoniae and E. coli strains and salvage rectal swab specimens collected from patients at 4 different health care facilities over a 7-month period were tested. The gold-standard comparator was bla(KPC) PCR testing of isolates. Method 1 detected 4/9 (44%) KPC-positive challenge isolates. By method 2, 9/9 KPC-positive challenge isolates exhibited zones of inhibition of < or = 27 mm; all KPC-negative isolates exhibited zones of inhibition greater than 27 mm. The sensitivity and specificity of method 1 for detection of KPC-positive K. pneumoniae and E. coli in 149 rectal swab specimens were 65.6% (95% confidence interval [CI], 46.8% to 80.8%) and 49.6% (95% CI, 40.3% to 58.9%), respectively. With method 2, a zone diameter of < or = 27 mm had a sensitivity of 97.0% (95% CI, 82.5% to 99.8%) and specificity of 90.5% (95% CI, 83.3% to 94.9%) for detection of KPC in rectal swab specimens. Direct ertapenem disk testing is simpler, more sensitive, and more specific than selective broth enrichment with imipenem for detection of KPC-producing K. pneumoniae and E. coli in surveillance specimens.

  16. Anti-bacterial effect of essential oil from Xanthium strumarium against shiga toxin-producing Escherichia coli.

    PubMed

    Sharifi-Rad, J; Soufi, L; Ayatollahi, S A M; Iriti, M; Sharifi-Rad, M; Varoni, E M; Shahri, F; Esposito, S; Kuhestani, K; Sharifi-Rad, M

    2016-01-01

    Shiga toxin-producing Escherichia coli (STEC) serotype O157:H7 is one of the most important human pathogenic microorganisms, which can cause life-threatening infections. Xanthium strumarium L. is a plant with anti-bacterial activity against gram-negative and gram-positive bacteria. This study aims to demonstrate in vitro efficacy of the essential oil (EO) extracted from Xanthium strumarium L. against E. coli O157:H7. Using the agar test diffusion, the effect of Xanthium strumarium L. EO (5, 10, 15, 30, 60, and 120 mg/mL) was verified at each of the four different growth phases of E. coli O157:H7. Cell counts of viable cells and colony forming unit (CFU) were determined at regular time points using Breed's method and colony counting method, respectively. No viable cell was detectable after the 1 hour-exposure to X. strumarium EO at 30, 60, and 120 mg/mL concentrations. No bacterial colony was formed after 1 h until the end of the incubation period at 24 h. At lower concentrations, the number of bacteria cells decreased and colonies could be observed only after incubation. At the exponential phase, the EO at 15 mg/mL was only bacteriostatic, while from 30 mg/mL started to be bactericidal. X. strumarium EO antibacterial activity against Shiga toxin-producing E. coli O157:H7 is dependent on EO concentration and physiological state of the microorganisms tested. The best inhibitory activity was achieved during the late exponential and the stationary phases. PMID:27650979

  17. FACTORS ASSOCIATED WITH EXTENDED SPECTRUM β-LACTAMASE PRODUCING ESCHERICHIA COLI IN COMMUNITY-ACQUIRED URINARY TRACT INFECTION AT HOSPITAL EMERGENCY DEPARTMENT, BANGKOK, THAILAND.

    PubMed

    Savatmorigkorngul, Sorravit; Poowarattanawiwit, Pongsuree; Sawanyawisuth, Kittisak; Sittichanbuncha, Yuwares

    2016-03-01

    Urinary tract infection or UTI is most commonly caused by Escherichia coli. This study investigated the prevalence of and risk factors for extended spectrum β-lactamase-producing (ESBL) E. coli in community-acquired UTI presenting at the Emergency Department, Faculty of Medicine Ramathibodi Hospital, Mahidol University, Bangkok, Thailand. A retrospective review was conducted over a one-year period (2014) of case histories of patients over 15 years of age diagnosed with (n = 159) and without culture-positive (n = 249) ESBL E. coli. Backward stepwise multivariate logistic regression analysis revealed four independent risk factors for UTI caused by ESBL E. coli, namely, urinary catheter use, previous UTI in which ESBL E. coli was present, and previous use of antibiotics cephalosporin and penicillin. This information should be useful in devising future public health prevention and control programs for ESBL E. coli-associated community-acquired UTI.

  18. Quantified high-throughput screening of Escherichia coli producing poly(3-hydroxybutyrate) based on FACS.

    PubMed

    Lee, Jae Hyung; Lee, Seung Hwan; Yim, Sung Sun; Kang, Kyoung-Hee; Lee, Sang Yup; Park, Si Jae; Jeong, Ki Jun

    2013-08-01

    Here, we report on a highly sensitive method for the detection of P(3HB) accumulation in Escherichia coli cells based on the automated flow cytometry system using fluorescent dyes. E. coli containing P(3HB) were stained with either BODIPY or Nile red fluorescent dye, and their staining properties were analyzed under a variety of conditions. Compared with Nile red, BODIPY was much more sensitive in staining P(3HB) and overall demonstrated a more rapid staining of cells, a greater resistance to photobleaching, and greater cell viability. In addition, we also successfully monitored heterogeneity in P(3HB) accumulation within a cell population using BODIPY staining and flow cytometry. We believe this optimized staining method using BODIPY in combination with screening by high-speed flow cytometer will be helpful in the engineering of host cells toward an enhanced production of bioplastics. PMID:23740474

  19. Use of Escherichia coli Nissle 1917 producing recombinant colicins for treatment of IBD patients.

    PubMed

    Kotłowski, Roman

    2016-08-01

    Patients with Crohn's Disease and Ulcerative Colitis infected with Adherent-Invasive Escherichia coli strains constitute the largest group among Inflammatory Bowel Disease subjects, when taking into account all known etiological agents of the disease. A possible link between these pathogenic bacteria and inflammation process has gained the confidence in recently published papers. Observed enteric neuroglial cells apoptosis and epithelial gaps of ileum are probably the key manifestations of inflammation, which has been shown in IBD patients in contrary to the samples taken from healthy individuals. The cascade of consecutive events from bacterial infection via inflammation to excessive apoptosis in IBD patients leads up to the aim of our hypothesis about designing of new therapeutic strategy directed to Adherent-Invasive E. coli strains. The main advantage of biological method, which will rely on application of E. coli Nissle 1917 strain as a carrier for specific recombinant colicins against AIEC strains, could probably cause a long-lasting remission of inflammation in CD and UC patients. PMID:27372848

  20. Complete solubilization and purification of recombinant human growth hormone produced in Escherichia coli.

    PubMed

    Kim, Min-Ji; Park, Hyun Soo; Seo, Kyung Hye; Yang, Hyo-Jin; Kim, Sook-Kyung; Choi, Jun-Hyuk

    2013-01-01

    High-level expression of recombinant human growth hormone (hGH) in Escherichia coli (E. coli) leads to the formation of insoluble aggregates as inclusion bodies devoid of biological activity. Until recently, significant efforts have been made to improve the recovery of active hGH from inclusion bodies. Here, we developed an efficient procedure for the production of completely soluble hGH by minimizing the formation of inclusion bodies and optimizing protein purification conditions. Under the newly established conditions we were able to obtain most of the total hGH in the soluble fraction. We show that the soluble protein can be efficiently purified in high yield by a series of chromatographic procedures. We analyzed the resulting hGH using various analytical techniques such as reversed-phase high-performance liquid chromatography (RP-HPLC), size-exclusion chromatography (SEC), matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry, and circular dichroism (CD). These multiple analyses support the conclusion that we obtained highly pure hGH with the expected molecular mass and intact secondary structure. The biological activity of purified hGH was also confirmed by evaluating its growth-promoting effect using a cell proliferation assay. Taken together, we describe a straightforward strategy for the production of completely soluble and biologically active hGH in E. coli.

  1. Life on the outside: role of biofilms in environmental persistence of Shiga-toxin producing Escherichia coli

    PubMed Central

    Vogeleer, Philippe; Tremblay, Yannick D. N.; Mafu, Akier A.; Jacques, Mario; Harel, Josée

    2014-01-01

    Escherichia coli is a heterogeneous species that can be part of the normal flora of humans but also include strains of medical importance. Among pathogenic members, Shiga-toxin producing E. coli (STEC) are some of the more prominent pathogenic E. coli within the public sphere. STEC disease outbreaks are typically associated with contaminated beef, contaminated drinking water, and contaminated fresh produce. These water- and food-borne pathogens usually colonize cattle asymptomatically; cows will shed STEC in their feces and the subsequent fecal contamination of the environment and processing plants is a major concern for food and public safety. This is especially important because STEC can survive for prolonged periods of time outside its host in environments such as water, produce, and farm soil. Biofilms are hypothesized to be important for survival in the environment especially on produce, in rivers, and in processing plants. Several factors involved in biofilm formation such as curli, cellulose, poly-N-acetyl glucosamine, and colanic acid are involved in plant colonization and adherence to different surfaces often found in meat processing plants. In food processing plants, contamination of beef carcasses occurs at different stages of processing and this is often caused by the formation of STEC biofilms on the surface of several pieces of equipment associated with slaughtering and processing. Biofilms protect bacteria against several challenges, including biocides used in industrial processes. STEC biofilms are less sensitive than planktonic cells to several chemical sanitizers such as quaternary ammonium compounds, peroxyacetic acid, and chlorine compounds. Increased resistance to sanitizers by STEC growing in a biofilm is likely to be a source of contamination in the processing plant. This review focuses on the role of biofilm formation by STEC as a means of persistence outside their animal host and factors associated with biofilm formation. PMID:25071733

  2. Understanding the role of agricultural practices in the potential colonization and contamination by Escherichia coli in the rhizospheres of fresh produce.

    PubMed

    Habteselassie, Mussie Y; Bischoff, Marianne; Applegate, Bruce; Reuhs, Bradley; Turco, Ronald F

    2010-11-01

    To better protect consumers from exposure to produce contaminated with Escherichia coli, the potential transfer of E. coli from manure or irrigation water to plants must be better understood. We used E. coli strains expressing bioluminescence (E. coli O157:H7 lux) or multiantibiotic resistance (E. coli²(+)) in this study. These marked strains enabled us to visualize in situ rhizosphere colonization and metabolic activity and to track the occurrence and survival of E. coli in soil, rhizosphere, and phyllosphere. When radish and lettuce seeds were treated with E. coli O157:H7 lux and grown in an agar-based growth system, rapid bacterial colonization of the germinating seedlings and high levels of microbial activity were seen. Introduction of E. coli²(+) to soil via manure or via manure in irrigation water showed that E. coli could establish itself in the lettuce rhizosphere. Regardless of introduction method, 15 days subsequent to its establishment in the rhizosphere, E. coli²(+) was detected on the phyllosphere of lettuce at an average number of 2.5 log CFU/g. When E. coli²(+) was introduced 17 and 32 days postseeding to untreated soil (rather than the plant surface) via irrigation, it was detected at low levels (1.4 log CFU/g) on the lettuce phyllosphere 10 days later. While E. coli²(+) persisted in the bulk and rhizosphere soil throughout the study period (day 41), it was not detected on the external portions of the phyllosphere after 27 days. Overall, we find that E. coli is mobile in the plant system and responds to the rhizosphere like other bacteria.

  3. Thermal inactivation of Escherichia coli O157:H7 and non-O157 shiga toxin-producing Escherichia coli cells in mechanically tenderized veal.

    PubMed

    Luchansky, John B; Porto-Fett, Anna C S; Shoyer, Bradley A; Thippareddi, Harshavardhan; Amaya, Jesus R; Lemler, Michael

    2014-07-01

    Preflattened veal cutlets (ca. 71.5 g, ca. 0.32 cm thick) were surface inoculated with ca. 6.8 log CFU/g of a multistrain cocktail of Escherichia coli O157:H7 (ECOH) or a cocktail made of single strains of serogroups O26, O45, O103, O104, O111, O121, and O145 of Shiga toxin-producing E. coli (STEC) cells and then were mechanically tenderized by passing once through a "Sir Steak" tenderizer. For each cooking time, in each of at least three trials, three inoculated and tenderized cutlets, with and without breading, were individually cooked in 15 or 30 ml of canola oil for 0.0, 0.75, 1.0, 1.25, 1.5, 1.75, or 2.25 min per side on an electric skillet set at 191.5°C. The temperatures of the meat and of the skillet were monitored and recorded using a type J thermocouple. Regardless of the breading or volume of oil used to cook the meat, the longer the cooking times, the higher was the internal temperature of the meat, along with a greater reduction of both ECOH and STEC. The average final internal temperature of the meat at the approximate geometric center ranged from 56.8 to 93.1°C. Microbial reductions of ca. 2.0 to 6.7 log CFU/g and ca. 2.6 to 6.2 log CFU/g were achieved for ECOH and STEC, respectively. Our data also revealed no differences in thermal inactivation of ECOH relative to the volume of oil used to cook nonbreaded cutlets. However, when cooking breaded cutlets, the use of more (30 ml) compared with less (15 ml) cooking oil resulted in greater reductions in pathogen numbers. To deliver about a 5.0-log reduction of ECOH and STEC, and to achieve the recommended internal temperature of 71.1°C, it was necessary to cook mechanically tenderized veal cutlets for at least 1.5 min per side on a preheated electric skillet set at 191.5°C and containing 15 ml of cooking oil. These data also established that cooking times and temperatures effective for inactivating serotype O157:H7 strains of E. coli in tenderized veal are equally effective against the additional six

  4. 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.

    PubMed

    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

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

    PubMed

    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. PMID:27217241

  6. Prevalence and Molecular Characterisation of Shiga Toxin-Producing Escherichia Coli in Raw Milk Cheeses from Lazio Region, Italy

    PubMed Central

    De Santis, Paola; Lovari, Sarah; Condoleo, Roberto; Bilei, Stefano; Marcianò, Rita; Mezher, Ziad

    2016-01-01

    In recent years, the incidence of foodborne diseases caused by Shiga toxin-producing Escherichia coli (STEC) has increased globally. For this reason, within the specific regional control plan for the detection of STEC in food products in Italy, the presence of STEC in unpasteurised milk cheeses was investigated. In total, 203 samples obtained from March 2011 to December 2013 were analysed, with two standard methods (ISO 16654:2001 and ISO 13136:2012). Two strains of E. coli O157 were isolated (2/161, 1.2%) but did not carry any virulence-associated genes and 22 stx-positive samples (22/146, 15.1%) were detected in enrichment cultures, mostly from ovine cheeses. Only two strains isolated from different ovine cheeses carried stx gene and none of these was eae-positive. This study confirms the presence of stx-positive E. coli and suggests that this type of food cannot be excluded as a potential vehicle of STEC. PMID:27800426

  7. Atmospheric cold plasma inactivation of Escherichia coli, Salmonella enterica serovar Typhimurium and Listeria monocytogenes inoculated on fresh produce.

    PubMed

    Ziuzina, D; Patil, S; Cullen, P J; Keener, K M; Bourke, P

    2014-09-01

    Atmospheric cold plasma (ACP) represents a potential alternative to traditional methods for non-thermal decontamination of foods. In this study, the antimicrobial efficacy of a novel dielectric barrier discharge ACP device against Escherichia coli, Salmonella enterica Typhimurium and Listeria monocytogenes inoculated on cherry tomatoes and strawberries, was examined. Bacteria were spot inoculated on the produce surface, air dried and sealed inside a rigid polypropylene container. Samples were indirectly exposed (i.e. placed outside plasma discharge) to a high voltage (70 kVRMS) air ACP and subsequently stored at room temperature for 24 h. ACP treatment for 10, 60 and 120 s resulted in reduction of Salmonella, E. coli and L. monocytogenes populations on tomato to undetectable levels from initial populations of 3.1, 6.3, and 6.7 log10 CFU/sample, respectively. However, an extended ACP treatment time was necessary to reduce bacterial populations attached on the more complex surface of strawberries. Treatment time for 300 s resulted in reduction of E. coli, Salmonella and L. monocytogenes populations by 3.5, 3.8 and 4.2 log10 CFU/sample, respectively, and also effectively reduced the background microflora of tomatoes. PMID:24929725

  8. Flagella interact with ionic plant lipids to mediate adherence of pathogenic Escherichia coli to fresh produce plants.

    PubMed

    Rossez, Yannick; Holmes, Ashleigh; Wolfson, Eliza B; Gally, David L; Mahajan, Arvind; Pedersen, Henriette L; Willats, William G T; Toth, Ian K; Holden, Nicola J

    2014-07-01

    Bacterial attachment to plant and animal surfaces is generally thought to constitute the initial step in colonization, requiring adherence factors such as flagella and fimbriae. We describe the molecular mechanism underpinning flagella-mediated adherence to plant tissue for the foodborne pathogen, enterohaemorrhagic Escherichia coli. Escherichia coli H7 flagella interacted with a sulphated carbohydrate (carrageenan) on a glycan array, which occurred in a dose-dependent manner. Adherence of E. coli O157 : H-expressing flagella of serotype H7, H6 or H48 to plants associated with outbreaks from fresh produce and to Arabidopsis thaliana, was dependent on flagella interactions with phospholipids and sulpholipids in plasma membranes. Adherence of purified H7 and H48 flagella to carrageenan was reduced at higher concentrations of KH2 PO4 or KCl, showing an ionic basis to the interactions. Purified H7 flagella were observed to physically interact with plasma membranes in spinach plants and in A.thaliana. The results show a specific interaction between E. coli H7, H6 and H48 flagella and ionic lipids in plant plasma membranes. The work extends our understanding of the molecular mechanisms underpinning E.coli flagella targeting of plant hosts and suggests a generic mechanism of recognition common in eukaryotic hosts belonging to different biological kingdoms.

  9. Atmospheric cold plasma inactivation of Escherichia coli, Salmonella enterica serovar Typhimurium and Listeria monocytogenes inoculated on fresh produce.

    PubMed

    Ziuzina, D; Patil, S; Cullen, P J; Keener, K M; Bourke, P

    2014-09-01

    Atmospheric cold plasma (ACP) represents a potential alternative to traditional methods for non-thermal decontamination of foods. In this study, the antimicrobial efficacy of a novel dielectric barrier discharge ACP device against Escherichia coli, Salmonella enterica Typhimurium and Listeria monocytogenes inoculated on cherry tomatoes and strawberries, was examined. Bacteria were spot inoculated on the produce surface, air dried and sealed inside a rigid polypropylene container. Samples were indirectly exposed (i.e. placed outside plasma discharge) to a high voltage (70 kVRMS) air ACP and subsequently stored at room temperature for 24 h. ACP treatment for 10, 60 and 120 s resulted in reduction of Salmonella, E. coli and L. monocytogenes populations on tomato to undetectable levels from initial populations of 3.1, 6.3, and 6.7 log10 CFU/sample, respectively. However, an extended ACP treatment time was necessary to reduce bacterial populations attached on the more complex surface of strawberries. Treatment time for 300 s resulted in reduction of E. coli, Salmonella and L. monocytogenes populations by 3.5, 3.8 and 4.2 log10 CFU/sample, respectively, and also effectively reduced the background microflora of tomatoes.

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

    PubMed

    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.

  11. Clinical Epidemiology and Molecular Analysis of Extended-Spectrum-β-Lactamase-Producing Escherichia coli in Nepal: Characteristics of Sequence Types 131 and 648

    PubMed Central

    Sherchan, Jatan Bahadur; Miyoshi-Akiyama, Tohru; Ohmagari, Norio; Kirikae, Teruo; Nagamatsu, Maki; Tojo, Masayoshi; Ohara, Hiroshi; Sherchand, Jeevan B.; Tandukar, Sarmila

    2015-01-01

    Recently, CTX-M-type extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli strains have emerged worldwide. In particular, E. coli with O antigen type 25 (O25) and sequence type 131 (ST131), which is often associated with the CTX-M-15 ESBL, has been increasingly reported globally; however, epidemiology reports on ESBL-producing E. coli in Asia are limited. Patients with clinical isolates of ESBL-producing E. coli in the Tribhuvan University teaching hospital in Kathmandu, Nepal, were included in this study. Whole-genome sequencing of the isolates was conducted to analyze multilocus sequence types, phylotypes, virulence genotypes, O25b-ST131 clones, and distribution of acquired drug resistance genes. During the study period, 105 patients with ESBL-producing E. coli isolation were identified, and the majority (90%) of these isolates were CTX-M-15 positive. The most dominant ST was ST131 (n = 54; 51.4%), followed by ST648 (n = 15; 14.3%). All ST131 isolates were identified as O25b-ST131 clones, subclone H30-Rx. Three ST groups (ST131, ST648, and non-ST131/648) were compared in further analyses. ST648 isolates had a proportionally higher resistance to non-β-lactam antibiotics and featured drug-resistant genes more frequently than ST131 or non-ST131/648 isolates. ST131 possessed the most virulence genes, followed by ST648. The clinical characteristics were similar among groups. More than 38% of ESBL-producing E. coli isolates were from the outpatient clinic, and pregnant patients comprised 24% of ESBL-producing E. coli cases. We revealed that the high resistance of ESBL-producing E. coli to multiple classes of antibiotics in Nepal is driven mainly by CTX-M-producing ST131 and ST648. Their immense prevalence in the communities is a matter of great concern. PMID:25824221

  12. Effect of high pressure treatment on the survival of Shiga toxin-producing Escherichia coli in strawberry puree.

    PubMed

    Hsu, HsinYun; Sheen, Shiowshuh; Sites, Joseph; Huang, Lihan; Wu, James Swi-Bea

    2014-06-01

    Most fresh produce, such as strawberries, receives minimal processing and is often eaten raw. Contamination of produce with pathogenic bacteria may occur during growth, harvest, processing, transportation, and storage (abuse temperature) and presents a serious public health risk. Strawberries have been implicated in an outbreak of Escherichia coli O157:H7 infection that sickened 15 people, including one death. Strawberries may also be contaminated by other serogroups of non-O157 Shiga toxin-producing E. coli (STEC), including O26, O45, O103, O111, O121 and O145, which have become known as the "Big Six" or "Top Six" non-O157 STECs. The objective of this research was to explore the potential application of high pressure processing (HPP) treatment to reduce or eliminate STECs in fresh strawberry puree (FSP). FSP, inoculated with a six-strain cocktail of the "Big Six" non-O157 STEC strains or a five-strain cocktail of E. coli O157:H7 in vacuum-sealed packages, were pressure-treated at 150, 250, 350, 450, 550, and 650 MPa (1 MPa = 10(6) N/m(2)) for 5, 15, and 30 min. HPP treatment, at 350 MPa for ≥5 min, significantly reduced STECs in FSP by about 6-log CFU/g from the initial cell population of ca. 8-log CFU/g. Cell rupture, observed by scanning electron microscopy (SEM), demonstrated that the HPP treatments can be potentially used to control both non-O157 and O157:H7 STECs in heat sensitive products.

  13. Shiga Toxin-Producing Escherichia coli in Meat and Vegetable Products in Emilia Romagna Region, Years 2012-2013

    PubMed Central

    Taddei, Roberta; Nocera, Lucia; Ricchi, Matteo; Merialdi, Giuseppe

    2015-01-01

    In 2012-2013 Emilia-Romagna Region introduced a monitoring plan for Shiga toxin-producing Escherichia coli (STEC) in foodstuff. Six hundred eighty-nine meat samples and 273 fruit and vegetable products were analyzed according to ISOTS13136. Pre-enriched samples were tested by multiplex real time PCR targeting the virulence genes eae, stx1 and stx2. Stx2 positive samples were investigated for the presence of serogroup O104 associated gene. O103, O111, O145, O157, O26 associated genes were tested on samples positive for stx in association with eae gene. Isolation of E. coli strains was attempted from samples positive for serogroup-associated genes. Thirty-four meat products (4.9%) resulted positive for stx1 and/or stx2 genes and 46 (6.7%) for stx1 and/or stx2 genes in association with eae gene. Forty-five (6.5%) samples resulted positive at least at one serogroup. Serogroup O103, O104, O111, O145, O157 and O26 genes were detected respectively in 1.3, 0.3, 0.1, 3.9, 2.9 and 2.5% samples; 0.6% samples resulted positive for STEC isolation (2 E. coli O103 and 2 E. coli O157). It is worth noting that STEC virulence genes were detected at high frequency (19%) in fresh pork meat sausages. Four (1.5%) vegetable samples were positive for stx1 and/or stx2 genes and 1 (0.4%) for stx1 and/or stx2 genes in association with eae gene; none resulted positive for the tested serogroups. Only a low number of samples positive by molecular methods were confirmed by cultural isolation. It is therefore of the uttermost importance for appropriate risk management, to be fully aware of the meaning of the analytical result. PMID:27800377

  14. Comparison of MALDI-TOF MS and AFLP for strain typing of ESBL-producing Escherichia coli.

    PubMed

    Veenemans, J; Welker, M; van Belkum, A; Saccomani, M C; Girard, V; Pettersson, A; Verhulst, C; Kluytmans-Vandenbergh, M; Kluytmans, J

    2016-05-01

    Typing of bacterial isolates using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) potentially provides an efficient on-site method to monitor the spread of antibiotic-resistant bacteria and rapidly detect outbreaks. We compared MALDI-MS typing results to those of amplified fragment length polymorphism (AFLP) in a collection of 52 ESBL-producing Escherichia coli, isolated in a Dutch nursing home with an on-going outbreak of ST131 E. coli. Specific MALDI types were defined based on spectral data from four replicate colony samples of isolates grown on Columbia agar using multivariate statistical procedures. Type-specific superspectra were computed for four E .coli MALDI-types and tested for the potential of rapid and automated typing. The effect of different incubation conditions on typing performance was tested by analysing five isolates incubated for 24 h and 48 h on five different media. Types defined based on MALDI spectra were largely in agreement with the AFLP results, although some MALDI types comprised of more than one AFLP type. In particular, isolates belonging to ST131 showed distinct mass patterns. The proportion of isolates correctly assigned was substantially lower for isolates incubated on Sabouraud-dextrose and Drigalski agars for 24 h, and for those incubated for 48 h (all media). Our results show that the identification of type-specific peaks potentially allows direct typing of isolates belonging to specific clonal lineages. Both incubation time and media affected type assignment, suggesting that there is a need for a careful standardization of incubation time and culturing conditions when developing MALDI-typing schemes for E. coli.

  15. Short communication: Isolation of Shiga toxin-producing Escherichia coli in raw milk and mozzarella cheese in southern Italy.

    PubMed

    Nobili, G; Franconieri, I; Basanisi, M G; La Bella, G; Tozzoli, R; Caprioli, A; La Salandra, G

    2016-10-01

    Shiga toxin-producing Escherichia coli (STEC) are a significant food-borne public health hazard in Europe, where most human infections are associated with 5 serogroups (O157, O26, O103, O145, and O111). In 2015, 95 food and environmental samples were examined for the presence of Shiga toxin genes (stx1 and stx2). The STEC were isolated from 2 raw milk and 1 mozzarella cheese samples that were collected in the period between June and September. To the best of our knowledge, this finding represents the first report of STEC isolation from mozzarella cheese produced in Italy, and it suggests that both the quality of raw milk used to produce mozzarella and the thermal inactivation treatment associated with the curd-stretching step should be carefully monitored. PMID:27522413

  16. Short communication: Isolation of Shiga toxin-producing Escherichia coli in raw milk and mozzarella cheese in southern Italy.

    PubMed

    Nobili, G; Franconieri, I; Basanisi, M G; La Bella, G; Tozzoli, R; Caprioli, A; La Salandra, G

    2016-10-01

    Shiga toxin-producing Escherichia coli (STEC) are a significant food-borne public health hazard in Europe, where most human infections are associated with 5 serogroups (O157, O26, O103, O145, and O111). In 2015, 95 food and environmental samples were examined for the presence of Shiga toxin genes (stx1 and stx2). The STEC were isolated from 2 raw milk and 1 mozzarella cheese samples that were collected in the period between June and September. To the best of our knowledge, this finding represents the first report of STEC isolation from mozzarella cheese produced in Italy, and it suggests that both the quality of raw milk used to produce mozzarella and the thermal inactivation treatment associated with the curd-stretching step should be carefully monitored.

  17. Renal damage and death in weaned mice after oral infection with Shiga toxin 2-producing Escherichia coli strains

    PubMed Central

    Brando, R J F; Miliwebsky, E; Bentancor, L; Deza, N; Baschkier, A; Ramos, M V; Fernández, G C; Meiss, R; Rivas, M; Palermo, M S

    2008-01-01

    Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 infections are considered a public health problem in both developed and developing countries because of their increasing incidence and the severity of clinical presentation. Approximately 10% of infected patients develop complications such as haemolytic uraemic syndrome (HUS) characterized by acute renal failure, thrombocytopenia and haemolytic anaemia. The precise sequence of events leading to HUS is still understood incompletely. Because of the lack of a reproducible small animal model for EHEC infections, in vivo studies examining EHEC–host early interactions are limited and insufficient. The aim of this study was to characterize the weaned BALB/c mouse as a model of E. coli O157:H7 infection. In this paper we report that human Shiga toxin 2 (Stx2)-producing EHEC strains can adhere to the intestinal epithelium of weaned BALB/c mice, and produce local damage which leads to systemic disease and death in a percentage of infected mice. The lethality of the EHEC strain is closely age-dependent, and is related to the bacterial ability to colonize intestine and to produce Stx2. It can be concluded that the weaned BALB/c mouse can be used as a small animal model to study host early responses, and the role of bacterial pathogenic factors in the induction of systemic disease, thus providing a useful tool for the evaluation of therapeutic or vaccine approaches. PMID:18549440

  18. Prevalence of sorbitol non-fermenting Shiga toxin-producing Escherichia coli in Black Bengal goats on smallholdings.

    PubMed

    Gupta, M DAS; DAS, A; Islam, M Z; Biswas, P K

    2016-09-01

    A cross-sectional survey was carried out in Bangladesh with the sampling of 514 Black Bengal goats on smallholdings to determine the presence of sorbitol non-fermenting (SNF) Shiga toxin-producing E. coli (STEC). Swab samples collected from the recto-anal junction were plated onto cefixime and potassium tellurite added sorbitol MacConkey (CT-SMAC) agar, a selective medium for STEC O157 serogroup, where this serogroup and other SNF STEC produce colourless colonies. The SNF E. coli (SNF EC) isolates obtained from the survey were investigated by PCR for the presence of Shiga toxin-producing genes, stx1 and stx2, and two other virulence genes, eae and hlyA that code for adherence factor (intimin protein) and pore-forming cytolysin, respectively. The SNF EC isolates were also assessed for the presence of the rfbO157 gene to verify their identity to O157 serogroup. The results revealed that the proportions of goats carrying SNF EC isolates and stx1 and stx2 genes were 6·2% (32/514) [95% confidence interval (CI) 4·4-8·7)], 1·2% (95% CI 0·5-2·6) and 1·2% (95% CI 0·5-2·6), respectively. All the SNF STEC tested negative for rfbO157, hlyA and eae genes. The risk for transmission of STEC from Black Bengal goats to humans is low. PMID:27267779

  19. Longitudinal Study of Shiga Toxin-Producing Escherichia coli Shedding in Sheep Feces: Persistence of Specific Clones in Sheep Flocks▿

    PubMed Central

    Sánchez, Sergio; Martínez, Remigio; García, Alfredo; Blanco, Jorge; Blanco, Jesús E.; Blanco, Miguel; Dahbi, Ghizlane; López, Cecilia; Mora, Azucena; Rey, Joaquín; Alonso, Juan M.

    2009-01-01

    To provide information on the persistence and maintenance of colonization with Shiga toxin-producing Escherichia coli (STEC) in sheep, pulsed-field gel electrophoresis analysis of STEC isolates (n = 145) belonging to serogroups O5, O91, and O146 from 39 healthy animals was performed in a 12-month longitudinal study carried out with four sheep flocks. At the flock level as well as the individual-animal level, the same clones were obtained on sampling occasions separated by as much as 11 months. PMID:19168649

  20. Structural insight in the inhibition of adherence of F4 fimbriae producing enterotoxigenic Escherichia coli by llama single domain antibodies.

    PubMed

    Moonens, Kristof; Van den Broeck, Imke; Okello, Emmanuel; Pardon, Els; De Kerpel, Maia; Remaut, Han; De Greve, Henri

    2015-01-01

    Enterotoxigenic Escherichia coli that cause neonatal and post-weaning diarrhea in piglets express F4 fimbriae to mediate attachment towards host receptors. Recently we described how llama single domain antibodies (VHHs) fused to IgA, produced in Arabidopsis thaliana seeds and fed to piglets resulted in a progressive decline in shedding of F4 positive ETEC bacteria. Here we present the structures of these inhibiting VHHs in complex with the major adhesive subunit FaeG. A conserved surface, distant from the lactose binding pocket, is targeted by these VHHs, highlighting the possibility of targeting epitopes on single-domain adhesins that are non-involved in receptor binding. PMID:25828907

  1. NDM-1-Producing Citrobacter freundii, Escherichia coli, and Acinetobacter baumannii Identified from a Single Patient in China.

    PubMed

    Huang, Ying-Min; Zhong, Lan-Lan; Zhang, Xue-Fei; Hu, Hang-Tong; Li, Yu-Qi; Yang, Xiao-Rong; Feng, Lian-Qiang; Huang, Xi; Tian, Guo-Bao

    2015-08-01

    We identified New Delhi metallo-β-lactamase (NDM-1)-producing Citrobacter freundii GB032, Escherichia coli GB102, and Acinetobacter baumannii GB661 in urine and stool samples from a single patient in China. Plasmid profiling and Southern blotting indicated that blaNDM-1 from GB032 and that from GB102 were likely located on the same plasmid, while blaNDM-1 from GB661 was located on a very large (>400-kb) plasmid. This case underscores the broad host range of blaNDM-1 and its potential to spread between members of the family Enterobacteriaceae and A. baumannii.

  2. Novel method for the rapid and specific extraction of multiple β2 -agonist residues in food by tailor-made Monolith-MIPs extraction disks and detection by gas chromatography with mass spectrometry.

    PubMed

    Liu, Haibo; Gan, Ning; Chen, Yinji; Ding, Qingqing; Huang, Jie; Lin, Saichai; Cao, Yuting; Li, Tianhua

    2016-09-01

    A quick and specific pretreatment method based on a series of extraction clean-up disks, consisting of molecularly imprinted polymer monoliths and C18 adsorbent, was developed for the specific enrichment of salbutamol and clenbuterol residues in food. The molecularly imprinted monolithic polymer disk was synthesized using salbutamol as a template through a one-step synthesis process. It can simultaneously and specifically recognize salbutamol and clenbuterol. The monolithic polymer disk and series of C18 disks were assembled with a syringe to form a set of tailor-made devices for the extraction of target molecules. In a single run, salbutamol and clenbuterol can be specifically extracted, cleaned, and eluted by methanol/acetic acid/H2 O. The target molecules, after a silylation derivatization reaction were detected by gas chromatography-mass spectrometry. The parameters including solvent desorption, sample pH, and the cycles of reloading were investigated and discussed. Under the optimized extraction and clean-up conditions, the limits of detection and quantitation were determined as 0.018-0.022 and 0.042-0.049 ng/g for salbutamol and clenbuterol, respectively. The assay described was convenient, rapid, and specific; thereby potentially efficient in the high-throughput analysis of β2 -agonists residues in real food samples.

  3. Characterization of Multidrug Resistant Extended-Spectrum Beta-Lactamase-Producing Escherichia coli among Uropathogens of Pediatrics in North of Iran

    PubMed Central

    Rezai, Mohammad Sadegh; Salehifar, Ebrahim; Rafiei, Alireza; Rafati, Mohammadreza; Shafahi, Kheironesa

    2015-01-01

    Escherichia coli remains as one of the most important bacteria causing infections in pediatrics and producing extended-spectrum beta-lactamases (ESBLs) making them resistant to beta-lactam antibiotics. In this study we aimed to genotype ESBL-producing E. coli isolates from pediatric patients for ESBL genes and determine their association with antimicrobial resistance. One hundred of the E. coli isolates were initially considered ESBL producing based on their MIC results. These isolates were then tested by polymerase chain reaction (PCR) for the presence or absence of CTX, TEM, SHV, GES, and VEB beta-lactamase genes. About 30.5% of isolated E. coli was ESBL-producing strain. The TEM gene was the most prevalent (49%) followed by SHV (44%), CTX (28%), VEB (8%), and GES (0%) genes. The ESBL-producing E. coli isolates were susceptible to carbapenems (66%) and amikacin (58%) and showed high resistance to cefixime (99%), colistin (82%), and ciprofloxacin (76%). In conclusion, carbapenems were the most effective antibiotics against ESBl-producing E. coli in urinary tract infection in North of Iran. The most prevalent gene is the TEM-type, but the other resistant genes and their antimicrobial resistance are on the rise. PMID:26064896

  4. Prevalence and Characteristics of the Epidemic Multiresistant Escherichia coli ST131 Clonal Group among Extended-Spectrum Beta-Lactamase-Producing E. coli Isolates in Copenhagen, Denmark

    PubMed Central

    Hansen, Dennis S.; Nilsson, Frida; Frimodt-Møller, Jakob; Leihof, Rikke Fleron; Struve, Carsten; Scheutz, Flemming; Johnston, Brian; Krogfelt, Karen A.; Johnson, James R.

    2013-01-01

    We report the characteristics of 115 extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli clinical isolates, from 115 unique Danish patients, over a 1-year study interval (1 October 2008 to 30 September 2009). Forty-four (38%) of the ESBL isolates represented sequence type 131 (ST13)1, from phylogenetic group B2. The remaining 71 isolates were from phylogenetic groups D (27%), A (22%), B1 (10%), and B2 (3%). Serogroup O25 ST131 isolates (n = 42; 95% of ST131) comprised 7 different K antigens, whereas two ST131 isolates were O16:K100:H5. Compared to non-ST131 isolates, ST131 isolates were associated positively with CTX-M-15 and negatively with CTX-M-1 and CTX-M-14. They also were associated positively with 11 virulence genes, including afa and dra (Dr family adhesins), the F10 papA allele (P fimbria variant), fimH (type 1 fimbriae), fyuA (yersiniabactin receptor), iha (adhesin siderophore), iutA (aerobactin receptor), kpsM II (group 2 capsules), malX (pathogenicity island marker), ompT (outer membrane protease), sat (secreted autotransporter toxin), and usp (uropathogenicity-specific protein) and negatively with hra (heat-resistant agglutinin) and iroN (salmochelin receptor). The consensus virulence gene profile (>90% prevalence) of the ST131 isolates included fimH, fyuA, malX, and usp (100% each), ompT and the F10 papA allele (95% each), and kpsM II and iutA (93% each). ST131 isolates were also positively associated with community acquisition, extraintestinal pathogenic E. coli (ExPEC) status, and the O25, K100, and H4 antigens. Thus, among ESBL E. coli isolates in Copenhagen, ST131 was the most prevalent clonal group, was community associated, and exhibited distinctive and comparatively extensive virulence profiles, plus a greater variety of capsular antigens than reported previously. PMID:23554186

  5. Prevalence and characteristics of the epidemic multiresistant Escherichia coli ST131 clonal group among extended-spectrum beta-lactamase-producing E. coli isolates in Copenhagen, Denmark.

    PubMed

    Olesen, Bente; Hansen, Dennis S; Nilsson, Frida; Frimodt-Møller, Jakob; Leihof, Rikke Fleron; Struve, Carsten; Scheutz, Flemming; Johnston, Brian; Krogfelt, Karen A; Johnson, James R

    2013-06-01

    We report the characteristics of 115 extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli clinical isolates, from 115 unique Danish patients, over a 1-year study interval (1 October 2008 to 30 September 2009). Forty-four (38%) of the ESBL isolates represented sequence type 131 (ST13)1, from phylogenetic group B2. The remaining 71 isolates were from phylogenetic groups D (27%), A (22%), B1 (10%), and B2 (3%). Serogroup O25 ST131 isolates (n = 42; 95% of ST131) comprised 7 different K antigens, whereas two ST131 isolates were O16:K100:H5. Compared to non-ST131 isolates, ST131 isolates were associated positively with CTX-M-15 and negatively with CTX-M-1 and CTX-M-14. They also were associated positively with 11 virulence genes, including afa and dra (Dr family adhesins), the F10 papA allele (P fimbria variant), fimH (type 1 fimbriae), fyuA (yersiniabactin receptor), iha (adhesin siderophore), iutA (aerobactin receptor), kpsM II (group 2 capsules), malX (pathogenicity island marker), ompT (outer membrane protease), sat (secreted autotransporter toxin), and usp (uropathogenicity-specific protein) and negatively with hra (heat-resistant agglutinin) and iroN (salmochelin receptor). The consensus virulence gene profile (>90% prevalence) of the ST131 isolates included fimH, fyuA, malX, and usp (100% each), ompT and the F10 papA allele (95% each), and kpsM II and iutA (93% each). ST131 isolates were also positively associated with community acquisition, extraintestinal pathogenic E. coli (ExPEC) status, and the O25, K100, and H4 antigens. Thus, among ESBL E. coli isolates in Copenhagen, ST131 was the most prevalent clonal group, was community associated, and exhibited distinctive and comparatively extensive virulence profiles, plus a greater variety of capsular antigens than reported previously.

  6. Recombinant L-Asparaginase from Zymomonas mobilis: A Potential New Antileukemic Agent Produced in Escherichia coli.

    PubMed

    Einsfeldt, Karen; Baptista, Isis Cavalcante; Pereira, Juliana Christina Castanheira Vicente; Costa-Amaral, Isabele Campos; Costa, Elaine Sobral da; Ribeiro, Maria Cecília Menks; Land, Marcelo Gerardin Poirot; Alves, Tito Lívio Moitinho; Larentis, Ariane Leites; Almeida, Rodrigo Volcan

    2016-01-01

    L-asparaginase is an enzyme used as a chemotherapeutic agent, mainly for treating acute lymphoblastic leukemia. In this study, the gene of L-asparaginase from Zymomonas mobilis was cloned in pET vectors, fused to a histidine tag, and had its codons optimized. The L-asparaginase was expressed extracellularly and intracellularly (cytoplasmically) in Escherichia coli in far larger quantities than obtained from the microorganism of origin, and sufficient for initial cytotoxicity tests on leukemic cells. The in silico analysis of the protein from Z. mobilis indicated the presence of a signal peptide in the sequence, as well as high identity to other sequences of L-asparaginases with antileukemic activity. The protein was expressed in a bioreactor with a complex culture medium, yielding 0.13 IU/mL extracellular L-asparaginase and 3.6 IU/mL intracellular L-asparaginase after 4 h of induction with IPTG. The cytotoxicity results suggest that recombinant L-asparaginase from Z. mobilis expressed extracellularly in E.coli has a cytotoxic and cytostatic effect on leukemic cells.

  7. Recombinant L-Asparaginase from Zymomonas mobilis: A Potential New Antileukemic Agent Produced in Escherichia coli

    PubMed Central

    Pereira, Juliana Christina Castanheira Vicente; Costa-Amaral, Isabele Campos; da Costa, Elaine Sobral; Ribeiro, Maria Cecília Menks; Land, Marcelo Gerardin Poirot; Alves, Tito Lívio Moitinho; Larentis, Ariane Leites; Almeida, Rodrigo Volcan

    2016-01-01

    L-asparaginase is an enzyme used as a chemotherapeutic agent, mainly for treating acute lymphoblastic leukemia. In this study, the gene of L-asparaginase from Zymomonas mobilis was cloned in pET vectors, fused to a histidine tag, and had its codons optimized. The L-asparaginase was expressed extracellularly and intracellularly (cytoplasmically) in Escherichia coli in far larger quantities than obtained from the microorganism of origin, and sufficient for initial cytotoxicity tests on leukemic cells. The in silico analysis of the protein from Z. mobilis indicated the presence of a signal peptide in the sequence, as well as high identity to other sequences of L-asparaginases with antileukemic activity. The protein was expressed in a bioreactor with a complex culture medium, yielding 0.13 IU/mL extracellular L-asparaginase and 3.6 IU/mL intracellular L-asparaginase after 4 h of induction with IPTG. The cytotoxicity results suggest that recombinant L-asparaginase from Z. mobilis expressed extracellularly in E.coli has a cytotoxic and cytostatic effect on leukemic cells. PMID:27253887

  8. Genetic characterization of Shiga toxin-producing Escherichia coli (STEC) and atypical enteropathogenic Escherichia coli (EPEC) isolates from goat's milk and goat farm environment.

    PubMed

    Álvarez-Suárez, María-Elena; Otero, Andrés; García-López, María-Luisa; Dahbi, Ghizlane; Blanco, Miguel; Mora, Azucena; Blanco, Jorge; Santos, Jesús A

    2016-11-01

    The aim of this study was to characterize a collection of 44 Shiga toxin-producing (STEC) and enteropathogenic Escherichia coli (EPEC) isolated from goat milk and goat farm environment. Of the 19 STEC isolates, five (26.3%) carried the stx1 gene, four (21.1%) the stx2 gene and 10 (52.6%) presented both stx genes. Six (31.6%) STEC strains were eae-positive and belonged to serotypes related to severe human disease (O157:H7 and O5:HNM). Another seven STEC strains were of serotype O146:H21 and three of serotype O166:H28, also linked to human disease. The STEC strains isolated from goat milk were of serotypes potentially pathogenic for humans. All the 25 EPEC isolates were considered atypical (aEPEC) and one aEPEC strain was of serotype O26:H11, a serotype frequently isolated in children with diarrhea. Multilocus sequence typing (MLST) was carried out with seven housekeeping genes and 23 sequence types (ST) were detected, 14 of them newly described. Twelve STs grouped STEC isolates and 11 STs grouped EPEC isolates. Genetic typing by pulsed field gel electrophoresis (PFGE) resulted in 38 patterns which grouped in 10 clusters. Well-defined groups were also observed for strains of pathogenic serotypes. In conclusion, strains of STEC and aEPEC belonging to serotypes related to severe human disease have been detected in goat milk and the goat farm environment. Ruminants are an important reservoir of STEC strains and the role of these animals as carriers of other pathogenic types of E. coli seems to be an emerging concern.

  9. Crystal Diagnostics Xpress™ E7 STEC Kit for the Rapid Multiplex Detection of E. coli O157 and non-O157 Shiga toxin-producing E. coli.

    PubMed

    Zhao, Weidong; Stumpf, Curtis H; Bullard, Brian; Kuzenko, Stephanie; Niehaus, Gary D

    2015-01-01

    The Crystal Diagnostics (CDx) Xpress E7 STEC kit is a rapid and sensitive detection assay for the detection of Escherichia coli O157 and six non-O157 Shiga toxin-producing E. coli (serogroups O26, O45, O1O3, O111, O121, and O145, collectively referred to as STEC) at 1 CFU/325 g of raw ground beef and raw beef trim, or 200 g of spinach. The system comprises an automatic Crystal Diagnostics Xpress System Reader that integrates immunochemical and optical processes for the liquid crystal-based detection of microorganisms, a CDx BioCassette that incorporates antibody-coupled microspheres and liquid crystal for selective identification of the intended microbe, and additional commercially available components. The Crystal Diagnostics Xpress System(TM) combines proprietary liquid crystal technology with antibody-coated paramagnetic microspheres to selectively capture and detect STEC from food matrixes. The Xpress System expeditiously (9.5 h enrichment) provides the sensitivity and specificity of the U. S. Department of Agriculture Food Safety and Inspection Service and the U. S. Food and Drug Administration reference methods in screening as low as 1 STEC CFU/test portion. The inclusivity validation demonstrated detection of 53 of 54 STEC test strains. Shelf life testing of the antibody-coated microspheres and other Crystal Diagnostic consumables indicated that all materials were stable for a minimum of 3 months (ongoing), and lot-to-lot testing demonstrated consistent results between lots (data not shown). The internal and independent laboratory tests demonstrate that the method is rapid and sensitive for screening of the target STEC. PMID:26651567

  10. Genetic characterization of Shiga toxin-producing Escherichia coli (STEC) and atypical enteropathogenic Escherichia coli (EPEC) isolates from goat's milk and goat farm environment.

    PubMed

    Álvarez-Suárez, María-Elena; Otero, Andrés; García-López, María-Luisa; Dahbi, Ghizlane; Blanco, Miguel; Mora, Azucena; Blanco, Jorge; Santos, Jesús A

    2016-11-01

    The aim of this study was to characterize a collection of 44 Shiga toxin-producing (STEC) and enteropathogenic Escherichia coli (EPEC) isolated from goat milk and goat farm environment. Of the 19 STEC isolates, five (26.3%) carried the stx1 gene, four (21.1%) the stx2 gene and 10 (52.6%) presented both stx genes. Six (31.6%) STEC strains were eae-positive and belonged to serotypes related to severe human disease (O157:H7 and O5:HNM). Another seven STEC strains were of serotype O146:H21 and three of serotype O166:H28, also linked to human disease. The STEC strains isolated from goat milk were of serotypes potentially pathogenic for humans. All the 25 EPEC isolates were considered atypical (aEPEC) and one aEPEC strain was of serotype O26:H11, a serotype frequently isolated in children with diarrhea. Multilocus sequence typing (MLST) was carried out with seven housekeeping genes and 23 sequence types (ST) were detected, 14 of them newly described. Twelve STs grouped STEC isolates and 11 STs grouped EPEC isolates. Genetic typing by pulsed field gel electrophoresis (PFGE) resulted in 38 patterns which grouped in 10 clusters. Well-defined groups were also observed for strains of pathogenic serotypes. In conclusion, strains of STEC and aEPEC belonging to serotypes related to severe human disease have been detected in goat milk and the goat farm environment. Ruminants are an important reservoir of STEC strains and the role of these animals as carriers of other pathogenic types of E. coli seems to be an emerging concern. PMID:27497630

  11. High Throughput Quantitative Expression Screening and Purification Applied to Recombinant Disulfide-rich Venom Proteins Produced in E. coli

    PubMed Central

    Saez, Natalie J.; Nozach, Hervé; Blemont, Marilyne; Vincentelli, Renaud

    2014-01-01

    Escherichia coli (E. coli) is the most widely used expression system for the production of recombinant proteins for structural and functional studies. However, purifying proteins is sometimes challenging since many proteins are expressed in an insoluble form. When working with difficult or multiple targets it is therefore recommended to use high throughput (HTP) protein expression screening on a small scale (1-4 ml cultures) to quickly identify conditions for soluble expression. To cope with the various structural genomics programs of the lab, a quantitative (within a range of 0.1-100 mg/L culture of recombinant protein) and HTP protein expression screening protocol was implemented and validated on thousands of proteins. The protocols were automated with the use of a liquid handling robot but can also be performed manually without specialized equipment. Disulfide-rich venom proteins are gaining increasing recognition for their potential as therapeutic drug leads. They can be highly potent and selective, but their complex disulfide bond networks make them challenging to produce. As a member of the FP7 European Venomics project (www.venomics.eu), our challenge is to develop successful production strategies with the aim of producing thousands of novel venom proteins for functional characterization. Aided by the redox properties of disulfide bond isomerase DsbC, we adapted our HTP production pipeline for the expression of oxidized, functional venom peptides in the E. coli cytoplasm. The protocols are also applicable to the production of diverse disulfide-rich proteins. Here we demonstrate our pipeline applied to the production of animal venom proteins. With the protocols described herein it is likely that soluble disulfide-rich proteins will be obtained in as little as a week. Even from a small scale, there is the potential to use the purified proteins for validating the oxidation state by mass spectrometry, for characterization in pilot studies, or for sensitive

  12. High throughput quantitative expression screening and purification applied to recombinant disulfide-rich venom proteins produced in E. coli.

    PubMed

    Saez, Natalie J; Nozach, Hervé; Blemont, Marilyne; Vincentelli, Renaud

    2014-07-30

    Escherichia coli (E. coli) is the most widely used expression system for the production of recombinant proteins for structural and functional studies. However, purifying proteins is sometimes challenging since many proteins are expressed in an insoluble form. When working with difficult or multiple targets it is therefore recommended to use high throughput (HTP) protein expression screening on a small scale (1-4 ml cultures) to quickly identify conditions for soluble expression. To cope with the various structural genomics programs of the lab, a quantitative (within a range of 0.1-100 mg/L culture of recombinant protein) and HTP protein expression screening protocol was implemented and validated on thousands of proteins. The protocols were automated with the use of a liquid handling robot but can also be performed manually without specialized equipment. Disulfide-rich venom proteins are gaining increasing recognition for their potential as therapeutic drug leads. They can be highly potent and selective, but their complex disulfide bond networks make them challenging to produce. As a member of the FP7 European Venomics project (www.venomics.eu), our challenge is to develop successful production strategies with the aim of producing thousands of novel venom proteins for functional characterization. Aided by the redox properties of disulfide bond isomerase DsbC, we adapted our HTP production pipeline for the expression of oxidized, functional venom peptides in the E. coli cytoplasm. The protocols are also applicable to the production of diverse disulfide-rich proteins. Here we demonstrate our pipeline applied to the production of animal venom proteins. With the protocols described herein it is likely that soluble disulfide-rich proteins will be obtained in as little as a week. Even from a small scale, there is the potential to use the purified proteins for validating the oxidation state by mass spectrometry, for characterization in pilot studies, or for sensitive

  13. Relative Fecal Abundance of Extended-Spectrum-β-Lactamase-Producing Escherichia coli Strains and Their Occurrence in Urinary Tract Infections in Women

    PubMed Central

    Lixandru, Brandusa; Cojocaru, Radu; Büke, Çağrı; Paramythiotou, Elisabeth; Angebault, Cécile; Visseaux, Claire; Djuikoue, Ingrid; Erdem, Esra; Burduniuc, Olga; El Mniai, Assiya; Marcel, Candice; Perrier, Marion; Kesteman, Thomas; Clermont, Olivier; Denamur, Erick; Armand-Lefèvre, Laurence; Andremont, Antoine

    2013-01-01

    Extended-spectrum-beta-lactamase (ESBL)-producing Escherichia coli (ESBL E. coli) strains are of major concern because few antibiotics remain active against these bacteria. We investigated the association between the fecal relative abundance (RA) of ESBL-producing E. coli (ESBL-RA) and the occurrence of ESBL E. coli urinary tract infections (UTIs). The first stool samples passed after suspicion of UTI from 310 women with subsequently confirmed E. coli UTIs were sampled and tested for ESBL-RA by culture on selective agar. Predictive values of ESBL-RA for ESBL E. coli UTI were analyzed for women who were not exposed to antibiotics when the stool was passed. ESBL E. coli isolates were characterized for ESBL type, phylogroup, relatedness, and virulence factors. The prevalence of ESBL E. coli fecal carriage was 20.3%, with ESBL E. coli UTIs being present in 12.3% of the women. The mean ESBL-RA (95% confidence interval [CI]) was 13-fold higher in women exposed to antibiotics at the time of sampling than in those not exposed (14.3% [range, 5.6% to 36.9%] versus 1.1% [range, 0.32% to 3.6%], respectively; P < 0.001) and 18-fold higher in women with ESBL E. coli UTI than in those with another E. coli UTI (10.0% [range, 0.54% to 100%] versus 0.56% [range, 0.15% to 2.1%[, respectively; P < 0.05). An ESBL-RA of <0.1% was 100% predictive of a non-ESBL E. coli UTI. ESBL type, phylogroup, relatedness, and virulence factors were not found to be associated with ESBL-RA. In conclusion, ESBL-RA was linked to the occurrence of ESBL E. coli UTI in women who were not exposed to antibiotics and who had the same clone of E. coli in urine samples and fecal samples. Especially, a low ESBL-RA appeared to be associated with a low risk of ESBL E. coli infection. PMID:23836184

  14. Predictors and Molecular Epidemiology of Community-Onset Extended-Spectrum β-Lactamase–Producing Escherichia coli Infection in a Midwestern Community

    PubMed Central

    Banerjee, Ritu; Strahilevitz, Jacob; Johnson, James R.; Nagwekar, Payal P.; Schora, Donna M.; Shevrin, Ilene; Du, Hongyan; Peterson, Lance R.; Robicsek, Ari

    2014-01-01

    OBJECTIVE To identify predictors of community-onset extended-spectrum β-lactamase (ESBL)–producing Escherichia coli infection. DESIGN Prospective case-control study. SETTING Acute care hospitals and ambulatory clinics in the Chicago, Illinois, region. PATIENTS Adults with E. coli clinical isolates cultured in ambulatory settings or within 48 hours of hospital admission. METHODS Cases were patients with ESBL-producing E. coli clinical isolates cultured in ambulatory settings or within 48 hours of admission, and controls were patients with non-ESBL-producing E. coli isolates, matched to cases by specimen, location, and date. Clinical variables were ascertained through interviews and medical record review. Molecular methods were used to identify ESBL types, sequence type ST131, and aac(6′)-Ib-cr. RESULTS We enrolled 94 cases and 158 controls. Multivariate risk factors for ESBL-producing E. coli infection included travel to India in the past year (odds ratio [OR], 14.40 [95% confidence interval (CI), 2.92–70.95]), ciprofloxacin use (OR, 3.92 [95% CI, 1.90–8.1]), and age (OR, 1.04 [95% CI, 1.02–1.06]). Case isolates exhibited high prevalence of CTX-M-15 (78%), ST131 (50%), and aac(6′)-Ib-cr (66% of isolates with CTX-M-15). CONCLUSIONS Providers should be aware of the increased risk of ESBL-producing E. coli infection among returned travelers, especially those from India. PMID:23917909

  15. Detection of Escherichia coli sequence type 131 clonal group among extended-spectrum β-lactamase-producing E. coli using VITEK MS Plus matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    PubMed

    Matsumura, Yasufumi; Yamamoto, Masaki; Nagao, Miki; Tanaka, Michio; Takakura, Shunji; Ichiyama, Satoshi

    2015-12-01

    We investigated the performance of the VITEK MS Plus system for the detection of Escherichia coli sequence type 131 (ST131) among extended-spectrum β-lactamase-producing E. coli isolates. The SARAMIS software could discriminate the 67 ST131 isolates from 82 non-ST131 isolates with a sensitivity of 86.6% and a specificity of 95.1%. PMID:26415529

  16. Properties of pertussis toxin B oligomer assembled in vitro from recombinant polypeptides produced by Escherichia coli.

    PubMed Central

    Burnette, W N; Arciniega, J L; Mar, V L; Burns, D L

    1992-01-01

    The subunits that make up the pentameric B oligomer of pertussis toxin (S2, S3, S4, and S5) were individually synthesized as recombinant polypeptides in Escherichia coli, isolated as insoluble inclusion bodies, and assembled into a multimeric form in vitro by spontaneous association following treatment with a chaotropic agent, reduction, and reoxidation. The recombinant B multimer, purified by fetuin-Sepharose affinity chromatography, contained all four of the individual subunits and possessed the mitogenic and hemagglutinating activities characteristic of the native B oligomer. Immunization of mice with the recombinant B oligomer elicited antibodies that neutralized pertussis toxin in vitro and, moreover, provided protection in vivo against the leukocytosis-promoting activity of the toxin. These results demonstrate the potential for assembly of complex multimeric proteins from recombinant DNA-derived polypeptides and provide a novel means for production of an acellular pertussis vaccine component. Images PMID:1587592

  17. A Shiga toxin-producing Escherichia coli O157 outbreak associated with consumption of rice cakes in 2011 in Japan.

    PubMed

    Nabae, K; Takahashi, M; Wakui, T; Kamiya, H; Nakashima, K; Taniguchi, K; Okabe, N

    2013-09-01

    In May 2011, an outbreak of Shiga toxin-producing Escherichia coli (STEC) O157 was reported from Yamagata Prefecture, Japan. Investigations, including a case-control study, revealed that the outbreak was linked to two varieties of rice cakes produced by a local manufacturer between 2 and 7 May. Active and passive surveillance identified 136 suspected cases, 142 confirmed cases, 26 asymptomatic cases, and 25 secondary cases. While no environmental samples taken from the manufacturing premises tested positive for STEC, other than a stool sample taken from one employee, on-site and epidemiological investigations indicated that STEC was introduced during the manufacturing process of rice cakes rather than through contamination of raw materials. This was the first reported outbreak of STEC associated with cakes and confectionery in Japan, which indicates that contamination and outbreaks of STEC can occur in any food unless proper precautions are taken.

  18. Reduction of extended-spectrum-β-lactamase- and AmpC-β-lactamase-producing Escherichia coli through processing in two broiler chicken slaughterhouses.

    PubMed

    Pacholewicz, Ewa; Liakopoulos, Apostolos; Swart, Arno; Gortemaker, Betty; Dierikx, Cindy; Havelaar, Arie; Schmitt, Heike

    2015-12-23

    Whilst broilers are recognised as a reservoir of extended-spectrum-β-lactamase (ESBL)- and AmpC-β-lactamase (AmpC)-producing Escherichia coli, there is currently limited knowledge on the effect of slaughtering on its concentrations on poultry meat. The aim of this study was to establish the concentration of ESBL/AmpC producing E. coli on broiler chicken carcasses through processing. In addition the changes in ESBL/AmpC producing E. coli concentrations were compared with generic E. coli and Campylobacter. In two slaughterhouses, the surface of the whole carcasses was sampled after 5 processing steps: bleeding, scalding, defeathering, evisceration and chilling. In total, 17 batches were sampled in two different slaughterhouses during the summers of 2012 and 2013. ESBL/AmpC producing E. coli was enumerated on MacConkey agar with 1mg/l cefotaxime, and the ESBL/AmpC phenotypes and genotypes were characterised. The ESBL/AmpC producing E. coli concentrations varied significantly between the incoming batches in both slaughterhouses. The concentrations on broiler chicken carcasses were significantly reduced during processing. In Slaughterhouse 1, all subsequent processing steps reduced the concentrations except evisceration which led to a slight increase that was statistically not significant. The changes in concentration between processing steps were relatively similar for all sampled batches in this slaughterhouse. In contrast, changes varied between batches in Slaughterhouse 2, and the overall reduction through processing was higher in Slaughterhouse 2. Changes in ESBL/AmpC producing E. coli along the processing line were similar to changes in generic E. coli in both slaughterhouses. The effect of defeathering differed between ESBL/AmpC producing E. coli and Campylobacter. ESBL/AmpC producing E. coli decreased after defeathering, whereas Campylobacter concentrations increased. The genotypes of ESBL/AmpC producing E. coli (blaCTX-M-1, blaSHV-12, blaCMY-2, blaTEM-52c

  19. Diversity of Escherichia coli strains producing extended-spectrum beta-lactamases in Spain: second nationwide study.

    PubMed

    Díaz, Miguel A; Hernández-Bello, José R; Rodríguez-Baño, Jesús; Martínez-Martínez, Luis; Calvo, Jorge; Blanco, Jorge; Pascual, Alvaro

    2010-08-01

    The prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli (ESBLEC) in Spain increased 8-fold from 2000 to 2006. ESBL type, clonal relationship, antimicrobial susceptibility, and clinical data about infections caused by ESBLEC are evaluated in a second nationwide study developed in 2006. From 1008 clinical isolates obtained over 2 months from 44 hospitals, 254 were used for further analysis. ESBL production was evaluated by synergy testing, PCR, and sequencing. Antimicrobial activity was evaluated by microdilution. The clonal relationship was evaluated by repetitive extragenic palindromic-PCR (REP-PCR). The O25b subtype and the new afa operon FM955459 were determined by triplex PCR in isolates producing CTX-M-15. Multilocus sequence typing was performed on these isolates. A total of 72% of all ESBLs were of the CTX-M type, 26.8% were of the SHV type, and 1.2% were of the TEM type. The most prevalent ESBLs were CTX-M-14 (119 isolates), SHV-12 (68 isolates), CTX-M-15 (37 isolates), and CTX-M-9 (21 isolates). By REP-PCR, 214 clones were detected. All but five CTX-M-15 ESBLEC isolates corresponded to the international O25b/ST131 clone. This clone had not been detected in the first study (published in 2000). Epidemiological and clinical features were studied in 304 representative patients. A total of 60% of the patients were older than 60 and had nonfatal underlying diseases, and 55% had recently received antibiotics. Urinary tract infections accounted for 71% of cases, and 9% were bacteremic. There has been a significant increase in the prevalence of ESBLEC in Spain, with most of these strains being CTX-M-producing isolates, including the pandemic O25b-ST131. SHV-12-producing E. coli remains an important cause of community-acquired infection.

  20. Diversity of Escherichia coli strains producing extended-spectrum beta-lactamases in Spain: second nationwide study.

    PubMed

    Díaz, Miguel A; Hernández-Bello, José R; Rodríguez-Baño, Jesús; Martínez-Martínez, Luis; Calvo, Jorge; Blanco, Jorge; Pascual, Alvaro

    2010-08-01

    The prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli (ESBLEC) in Spain increased 8-fold from 2000 to 2006. ESBL type, clonal relationship, antimicrobial susceptibility, and clinical data about infections caused by ESBLEC are evaluated in a second nationwide study developed in 2006. From 1008 clinical isolates obtained over 2 months from 44 hospitals, 254 were used for further analysis. ESBL production was evaluated by synergy testing, PCR, and sequencing. Antimicrobial activity was evaluated by microdilution. The clonal relationship was evaluated by repetitive extragenic palindromic-PCR (REP-PCR). The O25b subtype and the new afa operon FM955459 were determined by triplex PCR in isolates producing CTX-M-15. Multilocus sequence typing was performed on these isolates. A total of 72% of all ESBLs were of the CTX-M type, 26.8% were of the SHV type, and 1.2% were of the TEM type. The most prevalent ESBLs were CTX-M-14 (119 isolates), SHV-12 (68 isolates), CTX-M-15 (37 isolates), and CTX-M-9 (21 isolates). By REP-PCR, 214 clones were detected. All but five CTX-M-15 ESBLEC isolates corresponded to the international O25b/ST131 clone. This clone had not been detected in the first study (published in 2000). Epidemiological and clinical features were studied in 304 representative patients. A total of 60% of the patients were older than 60 and had nonfatal underlying diseases, and 55% had recently received antibiotics. Urinary tract infections accounted for 71% of cases, and 9% were bacteremic. There has been a significant increase in the prevalence of ESBLEC in Spain, with most of these strains being CTX-M-producing isolates, including the pandemic O25b-ST131. SHV-12-producing E. coli remains an important cause of community-acquired infection. PMID:20519460

  1. Molecular Characterization of Shiga Toxin-Producing Escherichia coli Isolated from Ruminant and Donkey Raw Milk Samples and Traditional Dairy Products in Iran

    PubMed Central

    Momtaz, Hassan; Farzan, Rahil; Rahimi, Ebrahim; Safarpoor Dehkordi, Farhad; Souod, Negar

    2012-01-01

    The aims of the current study were to detect the virulence factors and antibiotic resistance of Shiga toxin-producing E. coli, in animal milk and dairy products in Iran. After E. coli dentification with culture method, PCR assay were developed for detection of pathogenic genes, serotypes and antibiotic resistance genes of E. coli. Results showed that out of 719 samples, 102 (14.18%) were confirmed to be positive for E. coli and out of 102 positive samples, 17.64% were O26 and 13.72% were O157 and 1.96% were O91 and 1.96% were O145 serotypes. Totally, the prevalence of stx1 and papA genes were the highest while the prevalence of sfaS and fyuA were the lowest in the positive samples. PCR results showed that tetA, tetB were the highest (64.70%) and aac(3)-IV were the lowest (27.45%) antibiotic resistant genes in E. coli positive samples. Our study indicated that the isolated E. coli trains in these regions had a highest antibiotic resistance to tetracycline (58.82%) and the lowest to nitrofurantoin (3.92%). tetA gene and E. coli O157 serotype had highest and aac(3)-IV gene, and E. coli O145 serotype had a lowest frequency rates of antibiotics resistance genes, in the region. PMID:22919299

  2. Characterization of Fosfomycin Resistant Extended-Spectrum β-Lactamase-Producing Escherichia coli Isolates from Human and Pig in Taiwan

    PubMed Central

    Tseng, Sung-Pin; Wang, Sheng-Fan; Kuo, Cheng-Yu; Huang, Jun-Wei; Hung, Wei-Chun; Ke, Guan-Ming; Lu, Po-Liang

    2015-01-01

    To investigate the efficacy of fosfomycin against extended-spectrum β-lactamases (ESBL) producing Escherichia coli in Taiwan and the resistance mechanisms and characterization of human and pig isolates, we analyzed 145 ESBL-producing isolates collected from two hospitals (n = 123) and five farms (n = 22) in Taiwan from February to May, 2013. Antimicrobial susceptibilities were determined. Clonal relatedness was determined by PFGE and multi-locus sequence typing. ESBLs, ampC, and fosfomycin resistant genes were detected by PCR, and their flanking regions were determined by PCR mapping and sequencing. The fosfomycin resistant mechanisms, including modification of the antibiotic target (MurA), functionless transporters (GlpT and UhpT) and their regulating genes such as uhpA, cyaA, and ptsI, and antibiotic inactivation by enzymes (FosA and FosC), were examined. The size and replicon type of plasmids carrying fosfomycin resistant genes were analyzed. Our results revealed the susceptibility rates of fosfomycin were 94% for human ESBL-producing E. coli isolates and 77% for pig isolates. The PFGE analysis revealed 79 pulsotypes. No pulsotype was found existing in both human and pig isolates. Three pulsotypes were distributed among isolates from two hospitals. ISEcp1 carrying blaCTX-M-group 9 was the predominant transposable elements of the ESBL genes. Among the thirteen fosfomycin resistant isolates, functionless transporters were identified in 9 isolates. Three isolates contained novel amino acid substitutions (Asn67Ile, Phe151Ser and Trp164Ser, Val146Ala and His159Tyr, respectively) in MurA (the target of fosfomycin). Four isolates had fosfomycin modified enzyme (fosA3) in their plasmids. The fosA3 gene was harboured in an IncN-type plasmid (101 kbp) in the three pig isolates and an IncB/O-type plasmid (113 kbp) in the human isolate. In conclusion, we identified that 6% and 23% of the ESBL-producing E. coli from human and pigs were resistant to fosfomycin, respectively

  3. Evaluation of lactic acid as an initial and secondary subprimal intervention for Escherichia coli O157:H7, non-O157 Shiga toxin-producing E. coli, and a nonpathogenic E. coli surrogate for E. coli O157:H7.

    PubMed

    Pittman, C I; Geornaras, I; Woerner, D R; Nightingale, K K; Sofos, J N; Goodridge, L; Belk, K E

    2012-09-01

    Lactic acid can reduce microbial contamination on beef carcass surfaces when used as a food safety intervention, but effectiveness when applied to the surface of chilled beef subprimal sections is not well documented. Studies characterizing bacterial reduction on subprimals after lactic acid treatment would be useful for validations of hazard analysis critical control point (HACCP) systems. The objective of this study was to validate initial use of lactic acid as a subprimal intervention during beef fabrication followed by a secondary application to vacuum-packaged product that was applied at industry operating parameters. Chilled beef subprimal sections (100 cm(2)) were either left uninoculated or were inoculated with 6 log CFU/cm(2) of a 5-strain mixture of Escherichia coli O157:H7, a 12-strain mixture of non-O157 Shiga toxin-producing E. coli (STEC), or a 5-strain mixture of nonpathogenic (biotype I) E. coli that are considered surrogates for E. coli O157:H7. Uninoculated and inoculated subprimal sections received only an initial or an initial and a second "rework" application of lactic acid in a custombuilt spray cabinet at 1 of 16 application parameters. After the initial spray, total inoculum counts were reduced from 6.0 log CFU/cm(2) to 3.6, 4.4, and 4.4 log CFU/cm(2) for the E. coli surrogates, E. coli O157:H7, and non-O157 STEC inoculation groups, respectively. After the second (rework) application, total inoculum counts were 2.6, 3.2, and 3.6 log CFU/cm(2) for the E. coli surrogates, E. coli O157:H7, and non-O157 STEC inoculation groups, respectively. Both the initial and secondary lactic acid treatments effectively reduced counts of pathogenic and nonpathogenic strains of E. coli and natural microflora on beef subprimals. These data will be useful to the meat industry as part of the HACCP validation process.

  4. Usability and Performance of CHROMagar STEC Medium in Detection of Shiga Toxin-Producing Escherichia coli Strains

    PubMed Central

    Siitonen, Anja; Kaukoranta, Suvi-Sirkku

    2012-01-01

    The performance and usability of CHROMagar STEC medium (CHROMagar Microbiology, Paris, France) for routine detection of Shiga toxin-producing Escherichia coli (STEC) strains were examined. The ability of the medium to selectively propagate STEC strains differing by their serotypes and virulence genes was studied with a collection of diarrheagenic E. coli isolates (n = 365) consisting of 49 different serotypes and with non-STEC and other bacterial isolates (n = 264). A total of 272 diarrheagenic E. coli (75.0%) isolates covering 24 different serotypes grew on CHROMagar STEC. The highest detection sensitivities were observed within the STEC serogroups O26 (90.0%), O111 (100.0%), O121 (100.0%), O145 (100.0%), and O157 (84.9%), and growth on CHROMagar STEC was highly associated with the presence of the tellurite resistance gene (terD). The specificity of the medium was 98.9%. In addition, CHROMagar STEC was used in parallel with a Shiga toxin-detecting immunoassay (Ridaquick Verotoxin/O157 Combi; R-biopharm, Darmstadt, Germany) to screen fecal specimens (n = 47) collected from patients suffering from hemorrhagic diarrhea. Positive growth on CHROMagar STEC was confirmed by the Premier EHEC enzyme immunoassay (Meridian Bioscience, Inc., Cincinnati, OH), and discrepant results between the two screening methods were confirmed by stx gene-detecting PCR. All 16 of the 47 stool samples that showed positive growth on CHROMagar STEC were also positive in the confirmatory tests. CHROMagar STEC proved to be an interesting option for STEC screening, allowing good detection sensitivity and specificity and permitting strain isolation for further outbreak investigations when required. PMID:22933601

  5. Characterization of enteropathogenic and Shiga toxin-producing Escherichia coli in cattle and deer in a shared agroecosystem.

    PubMed

    Singh, Pallavi; Sha, Qiong; Lacher, David W; Del Valle, Jacquelyn; Mosci, Rebekah E; Moore, Jennifer A; Scribner, Kim T; Manning, Shannon D

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen. Cattle are suggested to be an important reservoir for STEC; however, these pathogens have also been isolated from other livestock and wildlife. In this study we sought to investigate transmission of STEC, enterohemorrhagic E. coli (EHEC) and enteropathogenic E. coli (EPEC) between cattle and white-tailed deer in a shared agroecosystem. Cattle feces were collected from 100 animals in a Michigan dairy farm in July 2012, while 163 deer fecal samples were collected during two sampling periods (March and June). The locations of deer fecal pellets were recorded via geographic information system mapping and microsatellite multi-locus genotyping was used to link the fecal samples to individual deer at both time points. Following subculture to sorbitol MacConkey agar and STEC CHROMagar, the pathogens were characterized by serotyping, stx profiling, and PCR-based fingerprinting; multilocus sequence typing (MLST) was performed on a subset. STEC and EHEC were cultured from 12 to 16% of cattle, respectively, and EPEC was found in 36%. Deer were significantly less likely to have a pathogen in March vs. June where the frequency of STEC, EHEC, and EPEC was 1, 6, and 22%, respectively. PCR fingerprinting and MLST clustered the cattle- and deer-derived strains together in a phylogenetic tree. Two STEC strains recovered from both animal species shared MLST and fingerprinting profiles, thereby providing evidence of interspecies transmission and highlighting the importance of wildlife species in pathogen shedding dynamics and persistence in the environment and cattle herds.

  6. Influence of selective media on successful detection of Shiga toxin-producing Escherichia coli in food, fecal, and environmental samples.

    PubMed

    Hussein, Hussein S; Bollinger, Laurie M

    2008-06-01

    Shiga toxin-producing Escherichia coli (STEC) strains have caused a large number of human illness outbreaks worldwide. In most cases, the infection was traced to consumption of meats or vegetables contaminated with cattle feces. To combat this public health problem, pre- and post-harvest control strategies are continuously implemented to assure food safety. Thus, rapid, reliable, and sensitive methods for STEC detection must be available to provide confidence not only in the meats or vegetables entering the food chain but also in testing humans with illnesses. As a result, enrichment for STEC has been a critical step in any successful protocol for their detection. The base media commonly used for STEC enrichment include sorbitol MacConkey agar, tryptic soy broth (TSB), E. coli broth, enterohemorrhagic E. coli broth, buffered peptone water (BPW), and brain heart infusion broth. In addition to bile salts, antibiotics (e.g., tellurite, cefixime, novobiocin, vancomycin, cefsulodin, and acriflavin) are used at different concentrations to enrich for STEC. In most published reports, however, the reasons for choosing the selective medium were not provided. Thus, this review was intended to evaluate the base media and antibiotics commonly used for STEC detection. The efficacy of a detection method will certainly depend on the choice of the base medium, selective agents, and their concentrations. The interactions among these factors are also expected to affect sensitivity of the detection method, especially when the test sample contains a small number of STEC cells. Because sensitivity of detection is expected to decline when testing for stressed or injured STEC cells, as is the case in environmental samples, a pre-enrichment step in TSB or BPW without antibiotics may be necessary. Future research should focus on identifying possible antibiotic combinations that effectively inhibit most background bacteria without affecting pathogenic STEC strains in the test sample.

  7. Shiga toxin-producing Escherichia coli (STEC): Zoonotic risks associated with psittacine pet birds in home environments.

    PubMed

    Gioia-Di Chiacchio, R M; Cunha, M P V; Sturn, R M; Moreno, L Z; Moreno, A M; Pereira, C B P; Martins, F H; Franzolin, M R; Piazza, R M F; Knöbl, T

    2016-02-29

    Psittacidae are frequently bred as pets worldwide, but little is known about the zoonotic risks of these animals. The objective of this study was to investigate the presence of Shiga toxin-producing Escherichia coli (STEC) in the feces of psittacine birds housed as pets. A total of 171 fecal samples (67 cockatiels, 59 budgerigars, and 45 agapornis) were cultured. Forty-two (E. coli) strains were identified, and the presence of the eae, stx1, and stx2 genes was determined using PCR. The antimicrobial resistance profiles of the STEC strains were determined using the disk diffusion method and phylogenetic analysis according to the new Clermont phylotyping method. Using these methods, 19.4% (8/42) of the STEC strains were determined to be positive for the eae and stx2 genes. The results revealed a STEC frequency of 4.6% in the birds (8/171), with a percentage of 8.47% in budgerigars (5/59), 4.47% in cockatiels (3/67), and 0% in agapornis (0/45). None of the STEC isolates belonged to the O157 serogroup. Most of the strains were classified as sensitive to the 18 antibiotics tested. None of the strains exhibited a multiresistance profile. In the phylogenetic analysis, two strains were classified as non-typeable, three were classified as B2, two were classified as F, and one was classified as Clade I. Seven of the eight STEC strains showed a clonal profile using AFLP. E. coli strains that are stx2(+) plus eae(+) are usually associated with severe human diseases such as hemorrhagic colitis and hemolytic-uremic syndrome. The STEC-positive results indicate the zoonotic risk of breeding psittacidae in home environments.

  8. Prevalence of Shiga toxin-producing Escherichia coli in food products of animal origin as determined by molecular methods.

    PubMed

    Rantsiou, Kalliopi; Alessandria, Valentina; Cocolin, Luca

    2012-03-01

    In this study we report on the prevalence and distribution of Shiga toxin-producing Escherichia coli (STEC) in food products of animal origin, collected in the Piedmont region of Italy, as determined by a combination of quantitative PCR (qPCR) protocols, applied directly to the samples, and of culture-dependent isolation and subsequent molecular identification and characterization of isolates. The qPCR protocols were developed and optimized in this study and targeted the rpoB gene (as a marker for total E. coli) and the stx₁, stx₂ and eaeA genes (as markers for potentially virulent E.coli). They were then used to test for STEC in 101 food samples, before and after enrichment. A STEC prevalence of 42% (21/50) for dairy products and 70% (36/51) for meat products was obtained. A total of 54 STEC isolates were recovered from dairy and meat samples, resulting in a prevalence of 36% and 27% in dairy and meat products, respectively, by the culture method. A large number of strains carried the stx₂ gene (39 out of the 54 STEC strains) compared to strains that carried stx₁ (30 out of 54); only 11 out of 54 strains contained the eaeA gene, while 14 strains contained both stx₁ and stx₂. Eight of the 54 isolates belonged to the O157 serogroup, and none belonged to serogroups O26, O145, O111 or O103. Strains isolated from meat products were diverse, as determined by Enterobacterial repetitive intergenic consensus-PCR (ERIC), while those isolated from dairy products were more similar and grouped together by cluster analysis. The results of the qPCR approach showed a high prevalence of STEC in dairy and meat based products, mainly fermented, indicating a possible safety risk for these types of food commodities.

  9. Characterization of enteropathogenic and Shiga toxin-producing Escherichia coli in cattle and deer in a shared agroecosystem

    PubMed Central

    Singh, Pallavi; Sha, Qiong; Lacher, David W.; Del Valle, Jacquelyn; Mosci, Rebekah E.; Moore, Jennifer A.; Scribner, Kim T.; Manning, Shannon D.

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen. Cattle are suggested to be an important reservoir for STEC; however, these pathogens have also been isolated from other livestock and wildlife. In this study we sought to investigate transmission of STEC, enterohemorrhagic E. coli (EHEC) and enteropathogenic E. coli (EPEC) between cattle and white-tailed deer in a shared agroecosystem. Cattle feces were collected from 100 animals in a Michigan dairy farm in July 2012, while 163 deer fecal samples were collected during two sampling periods (March and June). The locations of deer fecal pellets were recorded via geographic information system mapping and microsatellite multi-locus genotyping was used to link the fecal samples to individual deer at both time points. Following subculture to sorbitol MacConkey agar and STEC CHROMagar, the pathogens were characterized by serotyping, stx profiling, and PCR-based fingerprinting; multilocus sequence typing (MLST) was performed on a subset. STEC and EHEC were cultured from 12 to 16% of cattle, respectively, and EPEC was found in 36%. Deer were significantly less likely to have a pathogen in March vs. June where the frequency of STEC, EHEC, and EPEC was 1, 6, and 22%, respectively. PCR fingerprinting and MLST clustered the cattle- and deer-derived strains together in a phylogenetic tree. Two STEC strains recovered from both animal species shared MLST and fingerprinting profiles, thereby providing evidence of interspecies transmission and highlighting the importance of wildlife species in pathogen shedding dynamics and persistence in the environment and cattle herds. PMID:25883908

  10. Emergence of a Multidrug-Resistant Shiga Toxin-Producing Enterotoxigenic Escherichia coli Lineage in Diseased Swine in Japan.

    PubMed

    Kusumoto, Masahiro; Hikoda, Yuna; Fujii, Yuki; Murata, Misato; Miyoshi, Hirotsugu; Ogura, Yoshitoshi; Gotoh, Yasuhiro; Iwata, Taketoshi; Hayashi, Tetsuya; Akiba, Masato

    2016-04-01

    EnterotoxigenicEscherichia coli(ETEC) and Shiga toxin-producingE. coli(STEC) are important causes of diarrhea and edema disease in swine. The majority of swine-pathogenicE. colistrains belong to a limited range of O serogroups, including O8, O138, O139, O141, O147, O149, and O157, which are the most frequently reported strains worldwide. However, the circumstances of ETEC and STEC infections in Japan remain unknown; there have been few reports on the prevalence or characterization of swine-pathogenicE. coli In the present study, we determined the O serogroups of 967E. coliisolates collected between 1991 and 2014 from diseased swine in Japan, and we found that O139, O149, O116, and OSB9 (O serogroup ofShigella boydiitype 9) were the predominant serogroups. We further analyzed these four O serogroups using pulsed-field gel electrophoresis (PFGE), multilocus sequence typing, and virulence factor profiling. Most of the O139 and O149 strains formed serogroup-specific PFGE clusters (clusters I and II, respectively), whereas the O116 and OSB9 strains were grouped together in the same cluster (cluster III). All of the cluster III strains belonged to a single sequence type (ST88) and carried genes encoding both enterotoxin and Shiga toxin. This PFGE cluster III/ST88 lineage exhibited a high level of multidrug resistance (to a median of 10 antimicrobials). Notably, these bacteria were resistant to fluoroquinolones. Thus, this lineage should be considered a significant risk to animal production due to the toxigenicity and antimicrobial resistance of these bacteria. PMID:26865687

  11. Subtractive hybridization and identification of putative adhesins in a Shiga toxin-producing eae-negative Escherichia coli.

    PubMed

    Vidal, Maricel; Prado, Valeria; Whitlock, Gregory C; Solari, Aldo; Torres, Alfredo G; Vidal, Roberto M

    2008-12-01

    Adherence to epithelial cells by specific adhesins is a characteristic of Shiga toxin-producing Escherichia coli (STEC) strains. The eae-encoded protein intimin is the main adhesin implicated in intestinal colonization in vivo. We recently showed that STEC strains isolated in Chile displayed a wide variety of adhesins; here we demonstrate that some of these STEC strains are eae-negative and still adhere to epithelial cells at a level 100-fold higher than enterohaemorrhagic E. coli (EHEC) O157 : H7 prototype strain EDL933. This phenotype is associated with the presence of adherence factors different from the intimin protein. Subtractive hybridization between EHEC EDL933 and STEC eae-negative strain 472-1 was used to identify regions implicated in adhesion. In addition to the saa gene, we identified 18 specific genes in STEC 472-1, 16 of which had nucleotide identity to Salmonella ST46 phage genes; the two remaining ones shared identity to a gene encoding a hypothetical protein of uropathogenic E. coli. The DNA sequence of the STEC 472-1 psu-int region identified five open reading frames with homology to phage genes. We constructed mutant strains in the saa gene and the psu-int region to study the participation of these genes in the adherence to epithelial cells and our results demonstrated that STECDeltasaa and STECDeltapsu-int mutants displayed a 10-fold decrease in adherence as compared to the STEC 472-1 wild-type strain. Overall, our results suggest that STEC strain 472-1 adheres to epithelial cells in an eae-independent matter and that saa and psu-int participate in this adhesion process.

  12. Characterization of enteropathogenic and Shiga toxin-producing Escherichia coli in cattle and deer in a shared agroecosystem.

    PubMed

    Singh, Pallavi; Sha, Qiong; Lacher, David W; Del Valle, Jacquelyn; Mosci, Rebekah E; Moore, Jennifer A; Scribner, Kim T; Manning, Shannon D

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen. Cattle are suggested to be an important reservoir for STEC; however, these pathogens have also been isolated from other livestock and wildlife. In this study we sought to investigate transmission of STEC, enterohemorrhagic E. coli (EHEC) and enteropathogenic E. coli (EPEC) between cattle and white-tailed deer in a shared agroecosystem. Cattle feces were collected from 100 animals in a Michigan dairy farm in July 2012, while 163 deer fecal samples were collected during two sampling periods (March and June). The locations of deer fecal pellets were recorded via geographic information system mapping and microsatellite multi-locus genotyping was used to link the fecal samples to individual deer at both time points. Following subculture to sorbitol MacConkey agar and STEC CHROMagar, the pathogens were characterized by serotyping, stx profiling, and PCR-based fingerprinting; multilocus sequence typing (MLST) was performed on a subset. STEC and EHEC were cultured from 12 to 16% of cattle, respectively, and EPEC was found in 36%. Deer were significantly less likely to have a pathogen in March vs. June where the frequency of STEC, EHEC, and EPEC was 1, 6, and 22%, respectively. PCR fingerprinting and MLST clustered the cattle- and deer-derived strains together in a phylogenetic tree. Two STEC strains recovered from both animal species shared MLST and fingerprinting profiles, thereby providing evidence of interspecies transmission and highlighting the importance of wildlife species in pathogen shedding dynamics and persistence in the environment and cattle herds. PMID:25883908

  13. Internalization and thermal susceptibility of Shiga toxin-producing Escherichia coli (STEC) in marinated beef products.

    PubMed

    Pokharel, S; Brooks, J C; Martin, J N; Echeverry, A; Parks, A R; Corliss, B; Brashears, M M

    2016-06-01

    This study evaluated the internalization and cooking susceptibility of seven individual Escherichia coli (STEC) serogroups in surface-inoculated (10(5)log CFU/cm(2)) and vacuum tumbled marinated (30 or 60 min) bottom sirloin steaks. After storage for 14 days (0 to 2°C), flaps were cooked to various endpoint temperatures (55, 60, 65, and 71°C) for evaluation of pathogen survival by direct plating or rapid PCR based detection (BAX®). Direct plating of cooked samples yielded no enumerable plates. The data indicate varied internalization, translocation, and heat susceptibility patterns among serogroups. Using the rapid PCR based detection method O26, O103, and O111 were detected in flaps after cooking to 55 and 60°C, while O157:H7 survived in flaps cooked to 60 and 65°C. However, STEC O145 was the only serogroup that survived in all cooking temperatures. Serogroup O121 was not detected by plating or PCR in any cooked products. Intriguingly, STEC serogroups can be internalized during marination and the internalized pathogens vary in thermal susceptibility. PMID:26900979

  14. Activities of the feline immunodeficiency virus integrase protein produced in Escherichia coli.

    PubMed Central

    Vink, C; van der Linden, K H; Plasterk, R H

    1994-01-01

    Retroviral DNA integration requires the activity of at least one viral protein, the integrase (IN) protein. We cloned and expressed the integrase gene of feline immunodeficiency virus (FIV) in Escherichia coli as a fusion to the malE gene and purified the IN fusion protein by affinity chromatography. The protein is active in site-specific cleavage of the viral DNA ends, DNA strand transfer, and disintegration. FIV IN has a relaxed viral DNA substrate requirement: it cleaves and integrates FIV DNA termini, human immunodeficiency virus DNA ends, and Moloney murine leukemia virus DNA ends with high efficiencies. In the cleavage reaction, IN exposes a specific phosphodiester bond near the viral DNA end to nucleophilic attack. In vitro, either H2O, glycerol, or the 3' OH group of the viral DNA terminus can serve as nucleophile in this reaction. We found that FIV IN preferentially uses the 3' OH ends of the viral DNA as nucleophile, whereas HIV IN protein preferentially uses H2O and glycerol as nucleophiles. Images PMID:8107210

  15. Structural analysis of phage-borne stx genes and their flanking sequences in shiga toxin-producing Escherichia coli and Shigella dysenteriae type 1 strains.

    PubMed

    Unkmeir, A; Schmidt, H

    2000-09-01

    The stx-flanking regions of 49 Shiga toxin-producing Escherichia coli strains and nine Shigella dysenteriae serotype 1 strains containing either stx, stx(1), stx(2), or stx(2) variant genes, were examined. We analyzed these regions by PCR using a set of primers with one primer specific for the respective stx gene and a second primer complementary to sequences of Stx phages H-19B and 933W. We further characterized the amplification products by restriction endonuclease digestion and nucleotide sequencing. PCR products of stx(1)-containing E. coli strains of serogroups O157, O26, and 0103 showed the same lengths and similar restriction patterns. However, we failed to amplify the 3' stx-flanking region in stx(1)-harboring E. coli O111:H(-) strains. Stx2-producing E. coli strains revealed amplification products of different lengths and restriction patterns, suggesting greater heterogeneity than in stx(1)-positive strains. We also obtained specific PCR products for two Stx2c-producing and seven Stx2f-producing E. coli strains when they were subjected to PCR analysis. In nine S. dysenteriae type 1 strains, H-19B- and 933W-specific primers amplified only the 3' stx-flanking region. The results of our study demonstrate that the stx genes of all strains investigated are continuous with phage sequences. Whereas almost all strains except E. coli O111:H(-) strains were associated with a S-like gene, association with Q could not be demonstrated in nine S. dysenteriae type 1 strains and three E. coli strains. Furthermore, we showed that the organization of the stx-flanking regions is similar in all strains investigated, whereas fine-structure analysis showed subtle differences among the sequences examined. Our results support the hypothesis that stx genes in E. coli and S. dysenteriae are generally phage-borne.

  16. High Prevalence of Escherichia coli-Producing CTX-M-15 Extended-Spectrum Beta-Lactamases in Poultry and Human Clinical Isolates in Romania.

    PubMed

    Maciuca, Iuliana E; Williams, Nicola J; Tuchilus, Cristina; Dorneanu, Olivia; Guguianu, Eleonora; Carp-Carare, Catalin; Rimbu, Cristina; Timofte, Dorina

    2015-12-01

    Use of antibiotics in food animals may contribute to development and spread of resistant organisms, particularly so in some countries. The aim of this study was two-fold; first, to establish the prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in chicken production in a region within Romania. Second, to study the relatedness of ESBL-producing E. coli isolates recovered from broilers, abattoir workers where the chickens were slaughtered and from the human clinical specimens from two regional hospitals. The results indicated a very high (69%) rate of carriage of ESBL and AmpC-producing E. coli in chickens with 36% CTX-M producers. Sequencing showed that chickens in Romania have the highest worldwide prevalence (53%) of blaCTX-M-15 reported in poultry E. coli isolates. The majority (53%) of the extended-spectrum cephalosporin-resistant E. coli carried plasmid-mediated blaampC genes, mostly blaCMY-2 type, one of the highest prevalences reported in Europe. The predominant CTX-M type found in the human clinical E. coli isolates was blaCTX-M-15 and most isolates coharbored blaOXA-1, blaTEM, and aac(6')-ib-cr. The majority (60%) of the human clinical isolates belonged to the pandemic virulent clone B2-ST131. The clonal relationship between broiler and the human CTX-M-producing E. coli isolates was assessed by macrorestriction pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST), which indicated strain diversity with no common STs found between human and poultry isolates. Moreover, IncI1 was the most prevalent replicon found in broiler ESBL-producing E. coli isolates and also in transconjugants, indicating that plasmids and not clonal spread may play a role in the transfer of blaCTX-M genes. This study identifies a high prevalence of ESBL-producing E. coli from broiler chickens in Romania with a high occurrence incidence of blaCTX-M-15, which reflects the main ESBL type found in human E. coli infections in this

  17. Production of specific IgY antibody to the recombinant FanC protein produced in Escherichia coli

    PubMed Central

    Nasiri, Khadijeh; Zibaee, Saeed; Nassiri, Mohammadreza; Tahmoorespur, Mojtaba; Haghparast, Alireza

    2016-01-01

    Objective(s): Enterotoxigenic Escherichia coli (ETEC) strains are one of the primary causes of diarrhea in newborn calves and in humans, pigs, and sheep. IgY technology has been identified as a promising alternative to generating a mass amount of specific antibody for use in immunotherapy and immunodiagnostics. The purpose of this study was to produce specific antibody by egg yolk antibody (IgY) to recombinant FanC protein from ETEC. Materials and Methods: FanC (K99) gene was amplified from ETEC by specific primers and polymerase chain reaction. The gene was cloned and subcloned into pTZ57R/T and pET32a (+) vectors, respectively. Recombinant vector was transferred into E. coli BL21 CodonPlus (DE3). Protein expression was investigated by 1 mM IPTG induction. Hens were immunized by the purified recombinant FanC protein. The activity and specificity of the IgY antibody were detected by dot-blotting, Western blotting, and indirect ELISA. Results: We obtained FanC specific IgYs by immunizing the hens with the recombinant FanC protein. The anti-FanC IgY showed binding specifically to the FanC protein of ETEC. Conclusion: The results emphasize that specific IgY against the recombinant FanC protein could be recommended as a candidate for passive immunization against ETEC infection in animals and humans. PMID:27746871

  18. Detection and characterization of Shiga toxin-producing Escherichia coli in game meat and ready-to-eat meat products.

    PubMed

    Díaz-Sánchez, S; Sánchez, S; Sánchez, M; Herrera-León, S; Hanning, I; Vidal, D

    2012-11-15

    A total of 142 samples of game meat and ready-to-eat meat products from red deer and wild boar were analysed in order to assess the presence of Shiga toxin-producing Escherichia coli (STEC). Shiga-toxin encoding genes (stx genes) were detected by PCR in 36 (25.4%) of the samples and STEC was isolated from 8 (5.6%) of the same samples. None of the samples tested positive for E. coli O157:H7. Four different serotypes were found among the 8 STEC isolates, with serotype O27:H30 being predominant (62.5%, 5/8). The PCR assay indicated the presence of the stx2 gene in all of the STEC isolates and further subtyping resulted in detection of three different subtypes: stx2a, stx2b and stx2g. The only stx1-positive isolate was further subtyped as stx1c. The ehxA gene was detected in 3 (37.5%) of the isolates and none of them contained the eae gene. All STEC isolates were sensitive to the 13 antibiotics tested. Some isolates possessed serotypes and virulence gene profiles previously associated with STEC infections in humans. The isolation of a STEC strain carrying the stx2a subtype from a ready-to-eat meat product from deer suggests the role of these products as a potential source of STEC infections in humans.

  19. Defining pathogenic verocytotoxin-producing Escherichia coli (VTEC) from cases of human infection in the European Union, 2007-2010.

    PubMed

    Messens, W; Bolton, D; Frankel, G; Liebana, E; McLAUCHLIN, J; Morabito, S; Oswald, E; Threlfall, E J

    2015-06-01

    During 2007-2010, 13 545 confirmed human verocytotoxin (VT)-producing Escherichia coli (VTEC) infections were reported in the European Union, including 777 haemolytic uraemic syndrome (HUS) cases. Clinical manifestations were reported for 53% of cases, 64% of which presented with diarrhoea alone and 10% with HUS. Isolates from 85% of cases were not fully serotyped and could not be classified on the basis of the Karmali seropathotype concept. There is no single or combination of phenotypic or genetic marker(s) that fully define 'pathogenic' VTEC. Isolates which contain the vtx2 (verocytotoxin 2) gene in combination with the eae (intimin-encoding) gene or aaiC (secreted protein of enteroaggregative E. coli) and aggR (plasmid-encoded regulator) genes have been associated with a higher risk of more severe illness. A molecular approach targeting genes encoding VT and other virulence determinants is thus proposed to allow an assessment of the potential severity of disease that may be associated with a given VTEC isolate. PMID:25921781

  20. Approaches to treatment of emerging Shiga toxin-producing Escherichia coli infections highlighting the O104:H4 serotype

    PubMed Central

    Rahal, Elias A.; Fadlallah, Sukayna M.; Nassar, Farah J.; Kazzi, Natalie; Matar, Ghassan M.

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) are a group of diarrheagenic bacteria associated with foodborne outbreaks. Infection with these agents may result in grave sequelae that include fatality. A large number of STEC serotypes has been identified to date. E. coli serotype O104:H4 is an emerging pathogen responsible for a 2011 outbreak in Europe that resulted in over 4000 infections and 50 deaths. STEC pathogenicity is highly reliant on the production of one or more Shiga toxins that can inhibit protein synthesis in host cells resulting in a cytotoxicity that may affect various organ systems. Antimicrobials are usually avoided in the treatment of STEC infections since they are believed to induce bacterial cell lysis and the release of stored toxins. Some antimicrobials have also been reported to enhance toxin synthesis and production from these organisms. Various groups have attempted alternative treatment approaches including the administration of toxin-directed antibodies, toxin-adsorbing polymers, probiotic agents and natural remedies. The utility of antibiotics in treating STEC infections has also been reconsidered in recent years with certain modalities showing promise. PMID:25853096

  1. Shiga toxin-producing Escherichia coli: pre- and postharvest control measures to ensure safety of dairy cattle products.

    PubMed

    Hussein, Hussein S; Sakuma, Toshie

    2005-01-01

    The large number of cases of human illness caused by Shiga toxin-producing Escherichia coli (STEC) worldwide has raised safety concerns for foods of bovine origin. These human illnesses include diarrhea, hemorrhagic colitis, hemolytic uremic syndrome, and thrombotic thrombocytopenic purpura. Severe cases end with chronic renal failure, chronic nervous system deficiencies, and death. Over 100 STEC serotypes, including E. coli O157:H7, are known to cause these illnesses and to be shed in cattle feces. Thus, cattle are considered reservoirs of these foodborne pathogens. Because beef and dairy products were responsible for a large number of STEC outbreaks, efforts have been devoted to developing and implementing control measures that assure safety of foods derived from dairy cattle. These efforts should reduce consumers' safety concerns and support a competitive dairy industry at the production and processing levels. The efficacy of control measures both before harvest (i.e., on-farm management practices) and after harvest (i.e., milk processing and meat packing) for decreasing the risk of STEC contamination of dairy products was evaluated. The preharvest measures included sanitation during milking and management practices designed to decrease STEC prevalence in the dairy herd (i.e., animal factors, manure handling, drinking water, and both feeds and feeding). The postharvest measures included the practices or treatments that could be implemented during processing of milk, beef, or their products to eliminate or minimize STEC contamination.

  2. Identification of an NDM-5-producing Escherichia coli Sequence Type 167 in a Neonatal Patient in China

    PubMed Central

    Zhu, Yuan-qi; Zhao, Jing-yi; Xu, Cha; Zhao, Hui; Jia, Nan; Li, Yan-nian

    2016-01-01

    Emergence of New Delhi metallo-β-lactamase-producing Enterobacteriaceae has become a challenging threat to public health. Two carbapenem-resistant Escherichia coli, strain QD28 and QD29, were recovered from the aspirating sputum of a neonate and the urine of an adult in a Chinese hospital in 2013. Molecular typing revealed that both isolates belonged to the sequence type 167, but they were clonally diverse. Both isolates exhibited resistance to carbapenems, cephalosporins, ciprofloxacin, gentamicin, piperacillin-tazobactam and trimethoprim-sulfamethoxazole. In addition, strain QD28 was also resistant to aztreonam, and strain QD29 was resistant to amikacin, fosfomycin and minocycline. Antimicrobial resistance gene screening revealed that strain QD28 harbored aac(6′)-Ib, blaCTX-M-14, blaNDM-5, blaTEM-1 and sul1 genes, and strain QD29 harbored aac(6′)-Ib, blaCTX-M-3, blaNDM-5, blaTEM-1, rmtB, sul1 and sul2 genes. The blaNDM-5 gene was found to be located on a 46-kb plasmid in two isolates, and further sequence analysis showed that this plasmid was highly similar to the previously reported IncX3 plasmid pNDM-MGR194 in India. This is the first identification of blaNDM-5-carrying E. coli in the neonatal infection. PMID:27406405

  3. Shiga toxin-producing Escherichia coli O157:H7 in milk and milk products in Ogun State, Nigeria.

    PubMed

    Ivbade, Akhigbe; Ojo, Olufemi Ernest; Dipeolu, Morenike Atinuke

    2014-01-01

    Shiga toxin producing Escherichia coli (STEC) O157 is a major cause of food-borne illnesses in humans. This study investigated the presence of STEC O157 in milk and milk products in Ogun State, Nigeria. Of a total of 202 samples 10 (5%) were positive for STEC O157 including 1 (2%) of 50 raw milk samples, 3 (6%) of 50 samples of fresh local cheese, 1 (2%) of 50 samples of fried local cheese and 5 (9.6%) of 52 fermented milk samples. There was no significant difference (p>0.05) in the prevalence of STEC O157 among the sample types. Of 10 isolates, shiga toxin 1 gene (stx1) was detected only in 2 samples (20%), shiga toxin 2 (stx2) was extracted only in 6 samples (60%), stx1 /stx2 in 2 samples (20.0%), intimin gene (eaeA) in 5 samples (50%), and enterohaemolysin (E-hlyA) gene was isolated in 7 (70%) samples. Rates of resistance of the STEC O157 isolates were: amoxicillin/clavulanic acid 100%, ampicillin 100%, chloramphenicol 60%, nalidixic acid 20%, norfloxacin 10%, streptomycin 30%, sulphamethoxazole/trimethprim 20%, and tetracycline 90%. The isolates were all susceptible to ciprofloxacin and neomycin. The presence of virulent multidrug resistant E. coli O157 strains in milk and milk products as revealed by this study unveils a risk of human exposure to these potentially fatal pathogens following consumption of contaminated products. PMID:25273960

  4. Shiga toxin-producing Escherichia coli O157:H7 in milk and milk products in Ogun State, Nigeria.

    PubMed

    Ivbade, Akhigbe; Ojo, Olufemi Ernest; Dipeolu, Morenike Atinuke

    2014-01-01

    Shiga toxin producing Escherichia coli (STEC) O157 is a major cause of food-borne illnesses in humans. This study investigated the presence of STEC O157 in milk and milk products in Ogun State, Nigeria. Of a total of 202 samples 10 (5%) were positive for STEC O157 including 1 (2%) of 50 raw milk samples, 3 (6%) of 50 samples of fresh local cheese, 1 (2%) of 50 samples of fried local cheese and 5 (9.6%) of 52 fermented milk samples. There was no significant difference (p>0.05) in the prevalence of STEC O157 among the sample types. Of 10 isolates, shiga toxin 1 gene (stx1) was detected only in 2 samples (20%), shiga toxin 2 (stx2) was extracted only in 6 samples (60%), stx1 /stx2 in 2 samples (20.0%), intimin gene (eaeA) in 5 samples (50%), and enterohaemolysin (E-hlyA) gene was isolated in 7 (70%) samples. Rates of resistance of the STEC O157 isolates were: amoxicillin/clavulanic acid 100%, ampicillin 100%, chloramphenicol 60%, nalidixic acid 20%, norfloxacin 10%, streptomycin 30%, sulphamethoxazole/trimethprim 20%, and tetracycline 90%. The isolates were all susceptible to ciprofloxacin and neomycin. The presence of virulent multidrug resistant E. coli O157 strains in milk and milk products as revealed by this study unveils a risk of human exposure to these potentially fatal pathogens following consumption of contaminated products.

  5. Secondary transmissions during the outbreak of Shiga toxin-producing Escherichia coli O104 in Hesse, Germany, 2011.

    PubMed

    Hauri, Am; Gotsch, U; Strotmann, I; Krahn, J; Bettge-Weller, G; Westbrock, Hj; Bellinger, O; Uphoff, H

    2011-08-04

    During the recent outbreak of Shiga toxin-producing Escherichia coli (STEC) O104:H4 in Germany most cases notified in the State of Hesse (6 million inhabitants) were linked to satellite clusters or had travelled to the outbreak area in northern Germany. Intensified surveillance was introduced to rapidly identify cases not linked to known clusters or cases and thus to obtain timely information on possible further contaminated vehicles distributed in Hesse, as well to describe the risk of secondary transmission among known cases. As of 2 August 2011* [corrected], 56 cases of haemolytic uraemic syndrome (HUS) including two fatal cases, and 124 cases of STEC gastroenteritis meeting the national case definitions have been reported in Hesse. Among the 55 HUS and 81 STEC gastroenteritis cases thatmet the outbreak case definition, one HUS case and eight STEC gastroenteritis cases may have acquired their infection through secondary transmission. They include six possible transmissions within the family, two possible nosocomial and one possible laboratory transmission. Our results do not suggest an increased transmissibility of the outbreak strain compared to what is already known about E. coli O157 and other STEC serotypes.

  6. Defining pathogenic verocytotoxin-producing Escherichia coli (VTEC) from cases of human infection in the European Union, 2007-2010.

    PubMed

    Messens, W; Bolton, D; Frankel, G; Liebana, E; McLAUCHLIN, J; Morabito, S; Oswald, E; Threlfall, E J

    2015-06-01

    During 2007-2010, 13 545 confirmed human verocytotoxin (VT)-producing Escherichia coli (VTEC) infections were reported in the European Union, including 777 haemolytic uraemic syndrome (HUS) cases. Clinical manifestations were reported for 53% of cases, 64% of which presented with diarrhoea alone and 10% with HUS. Isolates from 85% of cases were not fully serotyped and could not be classified on the basis of the Karmali seropathotype concept. There is no single or combination of phenotypic or genetic marker(s) that fully define 'pathogenic' VTEC. Isolates which contain the vtx2 (verocytotoxin 2) gene in combination with the eae (intimin-encoding) gene or aaiC (secreted protein of enteroaggregative E. coli) and aggR (plasmid-encoded regulator) genes have been associated with a higher risk of more severe illness. A molecular approach targeting genes encoding VT and other virulence determinants is thus proposed to allow an assessment of the potential severity of disease that may be associated with a given VTEC isolate.

  7. Soft sensor control of metabolic fluxes in a recombinant Escherichia coli fed-batch cultivation producing green fluorescence protein.

    PubMed

    Gustavsson, Robert; Mandenius, Carl-Fredrik

    2013-10-01

    A soft sensor approach is described for controlling metabolic overflow from mixed-acid fermentation and glucose overflow metabolism in a fed-batch cultivation for production of recombinant green fluorescence protein (GFP) in Escherichia coli. The hardware part of the sensor consisted of a near-infrared in situ probe that monitored the E. coli biomass and an HPLC analyzer equipped with a filtration unit that measured the overflow metabolites. The computational part of the soft sensor used basic kinetic equations and summations for estimation of specific rates and total metabolite concentrations. Two control strategies for media feeding of the fed-batch cultivation were evaluated: (1) controlling the specific rates of overflow metabolism and mixed-acid fermentation metabolites at a fixed pre-set target values, and (2) controlling the concentration of the sum of these metabolites at a set level. The results indicate that the latter strategy was more efficient for maintaining a high titer and low variability of the produced recombinant GFP protein.

  8. Isolation and Characteristics of Shiga Toxin 2f-Producing Escherichia coli among Pigeons in Kyushu, Japan

    PubMed Central

    Murakami, Koichi; Etoh, Yoshiki; Ichihara, Sachiko; Maeda, Eriko; Takenaka, Shigeyuki; Horikawa, Kazumi; Narimatsu, Hiroshi; Kawano, Kimiko; Kawamura, Yoshiaki; Ito, Kenitiro

    2014-01-01

    An increasing number of Shiga toxin 2f-producing Escherichia coli (STEC2f) infections in humans are being reported in Europe, and pigeons have been suggested as a reservoir for the pathogen. In Japan, there is very little information regarding carriage of STEC2f by pigeons, prompting the need for further investigation. We collected 549 samples of pigeon droppings from 14 locations in Kyushu, Japan, to isolate STEC2f and to investigate characteristics of the isolates. Shiga toxin stx2f gene fragments were detected by PCR in 16 (2.9%) of the 549 dropping samples across four of the 14 locations. We obtained 23 STEC2f-isolates from seven of the original samples and from three pigeon dropping samples collected in an additional sampling experiment (from a total of seven locations across both sampling periods). Genotypic and phenotypic characteristics were then examined for selected isolates from each of 10 samples with pulsed-field gel electrophoresis profiles. Eight of the stx2f gene fragments sequenced in this study were homologous to others that were identified in Europe. Some isolates also contained virulence-related genes, including lpfAO26, irp2, and fyuA, and all of the 10 selected isolates maintained the eae, astA, and cdt genes. Moreover, five of the 10 selected isolates contained sfpA, a gene that is restricted to Shiga toxin-producing E. coli O165:H2 and sorbitol-fermenting Shiga toxin-producing E. coli O157:NM. We document serotypes O152:HNM, O128:HNM, and O145:H34 as STEC2f, which agrees with previous studies on pigeons and humans. Interestingly, O119:H21 was newly described as STEC2f. O145:H34, with sequence type 722, was described in a German study in humans and was also isolated in the current study. These results revealed that Japanese zoonotic STEC2f strains harboring several virulence-related factors may be of the same clonal complexes as some European strains. These findings provide useful information for public health-related disease management

  9. Risk of Escherichia coli O157:H7, Non-O157 Shiga Toxin-Producing Escherichia coli, and Campylobacter spp. in Food Animals and Their Products in Qatar.

    PubMed

    Mohammed, Hussni O; Stipetic, Korana; Salem, Ahmed; McDonough, Patrick; Chang, Yung Fu; Sultan, Ali

    2015-10-01

    Escherichia coli O157:H7, non-O157 E. coli, and Campylobacter spp. are among the top-ranked pathogens that threaten the safety of food supply systems around the world. The associated risks and predisposing factors were investigated in a dynamic animal population using a repeat-cross-sectional study design. Animal and environmental samples were collected from dairy and camel farms, chicken processing plants, and abattoirs and analyzed for the presence of these pathogens using a combination of bacterial enrichment and real-time PCR tests without culture confirmation. Data on putative risk factors were also collected and analyzed. E. coli O157:H7 was detected by PCR at higher levels in sheep and camel feces than in cattle feces (odds ratios [OR], 6.8 and 21.1, respectively). Although the genes indicating E. coli O157:H7 were detected at a relatively higher rate (4.3%) in fecal samples from dairy cattle, they were less common in milk and udder swabs from the same animals (1 and 2%, respectively). Among the food adulterants, E. coli O103 was more common in cattle fecal samples, whereas O26 was more common in sheep feces and O45 in camel feces compared with cattle (OR, 2.6 and 3.1, respectively). The occurrence of E. coli in the targeted populations differed by the type of sample and season of the year. Campylobacter jejuni and Campylobacter coli were more common in sheep and camel feces than in cattle feces. Most of the survey and surveillance of E. coli focused on serogroup O157 as a potential foodborne hazard; however, based on the PCR results, non-O157 Shiga toxin-producing E. coli serotypes appeared to be more common, and efforts should be made to include them in food safety programs.

  10. Risk of Escherichia coli O157:H7, Non-O157 Shiga Toxin-Producing Escherichia coli, and Campylobacter spp. in Food Animals and Their Products in Qatar.

    PubMed

    Mohammed, Hussni O; Stipetic, Korana; Salem, Ahmed; McDonough, Patrick; Chang, Yung Fu; Sultan, Ali

    2015-10-01

    Escherichia coli O157:H7, non-O157 E. coli, and Campylobacter spp. are among the top-ranked pathogens that threaten the safety of food supply systems around the world. The associated risks and predisposing factors were investigated in a dynamic animal population using a repeat-cross-sectional study design. Animal and environmental samples were collected from dairy and camel farms, chicken processing plants, and abattoirs and analyzed for the presence of these pathogens using a combination of bacterial enrichment and real-time PCR tests without culture confirmation. Data on putative risk factors were also collected and analyzed. E. coli O157:H7 was detected by PCR at higher levels in sheep and camel feces than in cattle feces (odds ratios [OR], 6.8 and 21.1, respectively). Although the genes indicating E. coli O157:H7 were detected at a relatively higher rate (4.3%) in fecal samples from dairy cattle, they were less common in milk and udder swabs from the same animals (1 and 2%, respectively). Among the food adulterants, E. coli O103 was more common in cattle fecal samples, whereas O26 was more common in sheep feces and O45 in camel feces compared with cattle (OR, 2.6 and 3.1, respectively). The occurrence of E. coli in the targeted populations differed by the type of sample and season of the year. Campylobacter jejuni and Campylobacter coli were more common in sheep and camel feces than in cattle feces. Most of the survey and surveillance of E. coli focused on serogroup O157 as a potential foodborne hazard; however, based on the PCR results, non-O157 Shiga toxin-producing E. coli serotypes appeared to be more common, and efforts should be made to include them in food safety programs. PMID:26408129

  11. Aggregative adherence fimbriae I (AAF/I) mediate colonization of fresh produce and abiotic surface by Shiga toxigenic enteroaggregative Escherichia coli O104:H4.

    PubMed

    Nagy, Attila; Xu, Yunfeng; Bauchan, Gary R; Shelton, Daniel R; Nou, Xiangwu

    2016-07-16

    The Shiga toxigenic Escherichia coli O104:H4 isolated during the 2011 European outbreak expresses Shiga toxin 2a and possess virulence genes associated with the enteroaggregative E. coli (EAEC) pathotype. It produces plasmid encoded aggregative adherence fimbriae I (AAF/I) which mediate cell aggregation and biofilm formation in human intestine and promote Shiga-toxin adsorption, but it is not clear whether the AAF/I fimbriae are involved in the colonization and biofilm formation on food and environmental matrices such as the surface of fresh produce. We deleted the gene encoding for the AAF/I fimbriae main subunit (AggA) from an outbreak associated E. coli O104:H4 strain, and evaluated the role of AAF/I fimbriae in the adherence and colonization of E. coli O104:H4 to spinach and abiotic surfaces. The deletion of aggA did not affect the adherence of E. coli O104:H4 to these surfaces. However, it severely diminished the colonization and biofilm formation of E. coli O104:H4 on these surfaces. Strong aggregation and biofilm formation on spinach and abiotic surfaces were observed with the wild type strain but not the isogenic aggA deletion mutant, suggesting that AAF/I fimbriae play a crucial role in persistence of O104:H4 cells outside of the intestines of host species, such as on the surface of fresh produce.

  12. Occurrence of non-O157 Shiga toxin-producing Escherichia coli in two commercial swine farms in the Eastern Cape Province, South Africa.

    PubMed

    Iwu, Chinwe Juliana; Iweriebor, Benson Chuks; Obi, Larry Chikwelu; Okoh, Anthony Ifeanyi

    2016-02-01

    Shiga toxin-producing Escherichia coli (STEC) is one of the most significant causes of food-borne infections capable of causing serious health complications in humans. Even though ruminants are known to be the major reservoirs of STEC, other non-ruminant food producing animals may also harbour pathogenic E. coli strains. In this study, we investigated the prevalence of E. coli serogroups O26, O111, O121, O145, and O157 and their associated virulence genes (stx1, stx2, eae, and ehxA) in swine faecal samples obtained from the two major commercial farms located in the Nkonkobe Municipality, Eastern Cape, South Africa. The proportions of serogroups detected were O26; 35 (7%), O145; 14 (2.8%), and O157:H7; 43 (8.6%) of the total animals sampled. Out of the 500 animals sampled, 22 isolates of E. coli (1.4%) tested positive for the stx2 gene, and 7 of these isolates belonged to E. coli O26 serogroup, while the remaining 15 most likely belonged to serogroups untargeted in this study. Other virulence genes (stx1, eae, and ehxA) that we screened for were not detected. These findings reveal that pigs within the Eastern Cape Province of South Africa can harbour Shiga toxin-producing E. coli.

  13. Occurrence of non-O157 Shiga toxin-producing Escherichia coli in two commercial swine farms in the Eastern Cape Province, South Africa.

    PubMed

    Iwu, Chinwe Juliana; Iweriebor, Benson Chuks; Obi, Larry Chikwelu; Okoh, Anthony Ifeanyi

    2016-02-01

    Shiga toxin-producing Escherichia coli (STEC) is one of the most significant causes of food-borne infections capable of causing serious health complications in humans. Even though ruminants are known to be the major reservoirs of STEC, other non-ruminant food producing animals may also harbour pathogenic E. coli strains. In this study, we investigated the prevalence of E. coli serogroups O26, O111, O121, O145, and O157 and their associated virulence genes (stx1, stx2, eae, and ehxA) in swine faecal samples obtained from the two major commercial farms located in the Nkonkobe Municipality, Eastern Cape, South Africa. The proportions of serogroups detected were O26; 35 (7%), O145; 14 (2.8%), and O157:H7; 43 (8.6%) of the total animals sampled. Out of the 500 animals sampled, 22 isolates of E. coli (1.4%) tested positive for the stx2 gene, and 7 of these isolates belonged to E. coli O26 serogroup, while the remaining 15 most likely belonged to serogroups untargeted in this study. Other virulence genes (stx1, eae, and ehxA) that we screened for were not detected. These findings reveal that pigs within the Eastern Cape Province of South Africa can harbour Shiga toxin-producing E. coli. PMID:26851595

  14. An Influenza A/H1N1/2009 Hemagglutinin Vaccine Produced in Escherichia coli

    PubMed Central

    Aguilar-Yáñez, José M.; Portillo-Lara, Roberto; Mendoza-Ochoa, Gonzalo I.; García-Echauri, Sergio A.; López-Pacheco, Felipe; Bulnes-Abundis, David; Salgado-Gallegos, Johari; Lara-Mayorga, Itzel M.; Webb-Vargas, Yenny; León-Angel, Felipe O.; Rivero-Aranda, Ramón E.; Oropeza-Almazán, Yuriana; Ruiz-Palacios, Guillermo M.; Zertuche-Guerra, Manuel I.; DuBois, Rebecca M.; White, Stephen W.; Schultz-Cherry, Stacey; Russell, Charles J.; Alvarez, Mario M.

    2010-01-01

    Background The A/H1N1/2009 influenza pandemic made evident the need for faster and higher-yield methods for the production of influenza vaccines. Platforms based on virus culture in mammalian or insect cells are currently under investigation. Alternatively, expression of fragments of the hemagglutinin (HA) protein in prokaryotic systems can potentially be the most efficacious strategy for the manufacture of large quantities of influenza vaccine in a short period of time. Despite experimental evidence on the immunogenic potential of HA protein constructs expressed in bacteria, it is still generally accepted that glycosylation should be a requirement for vaccine efficacy. Methodology/Principal Findings We expressed the globular HA receptor binding domain, referred to here as HA63–286-RBD, of the influenza A/H1N1/2009 virus in Escherichia coli using a simple, robust and scalable process. The recombinant protein was refolded and purified from the insoluble fraction of the cellular lysate as a single species. Recombinant HA63–286-RBD appears to be properly folded, as shown by analytical ultracentrifugation and bio-recognition assays. It binds specifically to serum antibodies from influenza A/H1N1/2009 patients and was found to be immunogenic, to be capable of triggering the production of neutralizing antibodies, and to have protective activity in the ferret model. Conclusions/Significance Projections based on our production/purification data indicate that this strategy could yield up to half a billion doses of vaccine per month in a medium-scale pharmaceutical production facility equipped for bacterial culture. Also, our findings demonstrate that glycosylation is not a mandatory requirement for influenza vaccine efficacy. PMID:20661476

  15. Genotype Cluster Analysis in Pathogenic Escherichia coli Isolates Producing Different CDT Types

    PubMed Central

    Javadi, Maryam; Oloomi, Mana; Bouzari, Saeid

    2016-01-01

    Diarrheagenic and uropathogenic E. coli types are mainly characterized by the expression of distinctive bacterial virulent factors. stx1, stx2 (Shiga toxins), and cdt (cytolethal distending toxin) genes have been acquired by horizontal gene transfer. Some virulent genes such as espP (serine protease), etpD (part of secretion pathway), and katP (catalase-peroxidase), or sfpA gene (Sfp fimbriae), are on plasmids and the others like fliC (flagellin) and the fimH gene (fimbriae type-I) are located on chromosome. Genomic pathogenicity islands (PAIs) carry some virulent genes such as hly gene. To determine the existence of virulence genes in cdt clinical isolates, genes including stx1, stx2, cdt, hly, espP, katP, sfpA, etpD, fliC, and fimH were assessed by Polymerase Chain Reaction (PCR). The most prevalent isolates for etpD and katP genes were 85.7% in cdtII. katP gene was also observed 83.3% in cdtI. However, in 42.85% of cdtIII isolates, espP gene was the most detected. Moreover, hly gene was also the most prominent gene in cdtIII (71.42%). sfpA gene was observed in 66.6% of cdtV. stx1 gene was detected in 100% of cdtII, cdtIV, and cdtV types. Presence and pattern of virulence genes were considered among cdt positive isotypes and used for their clustering and profiling. PMID:27042356

  16. Outcome of cephalosporin treatment of bacteremia due to CTX-M-type extended-spectrum beta-lactamase-producing Escherichia coli.

    PubMed

    Bin, Cao; Hui, Wang; Renyuan, Zhu; Yongzhong, Ning; Xiuli, Xie; Yingchun, Xu; Yuanjue, Zhu; Minjun, Chen

    2006-12-01

    The aim of the study was to analyze the outcome of different antibiotic treatments for bacteremia due to CTX-M-type extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli. In a prospective controlled clinical study from October 2002 to April 2005, 22 consecutive cases of bacteremia due to ESBL-producing E. coli with a ceftazidime-inhibition zone diameter of > or =18 mm were studied. The Etest method was used to determine the MIC values of cefotaxime, ceftazidime, imipenem, gentamicin, and ciprofloxacin against 22 isolates of E. coli. The polymerase chain reaction and sequencing analyses were used to determine the genotypes of the ESBLs. Of these 22 episodes, 7 were treated with ceftazidime, 8 were treated with imipenem/cilastatin, and 7 were treated with cefoperazone/sulbactam after detection of bacteremia. The demographic characteristics were comparable between the 3 groups. The treatment success ratio was similar (ceftazidime 85.7%, imipenem/cilastatin 87.5%, cefoperazone/sulbactam 71.4%, P = 0.637). Difficulties arose during treatment of peritonitis caused by CTX-M-producing E. coli bacteremia. Patients with bacteremia associated with urinary tract infection or biliary tract infection had a better chance of survival. All the 22 strains of E. coli produced CTX-M ESBLs (CTX-M-3, CTX-M-14, or CTX-M-27). The MICs of ceftazidime for 22 strains of E. coli were < or =8 microg/mL. All 7 patients who received ceftazidime survived, 6 of them were cured. Treatment in one patient with a ceftazidime MIC of 2 mug/mL failed because of abdominal abscess. Treatment with ceftazidime in vivo was effective against cases of CTX-M ESBL-producing E. coli bacteremia due to urinary tract infections and biliary tract infection when the MICs of ceftazidime were < or =8 microg/mL.

  17. Hemolytic uremic syndrome with mild renal involvement due to Shiga toxin-producing Escherichia coli (STEC) O145 strain.

    PubMed

    Pérez, Lucía; Apezteguía, Lucía; Piñeyrúa, Cecilia; Dabezies, Agustín; Bianco, María N; Schelotto, Felipe; Varela, Gustavo

    2014-01-01

    Hemolytic uremic syndrome (HUS) is a disorder characterized by the presence of the classic triad: microangiopathic hemolytic anemia, thrombocytopenia and acute renal injury. HUS without acute renal failure can be confused with other hematologic diseases. An infantile HUS caused by a Shiga-toxin-producing Escherichia coli (STEC) O145 strain carrying genotype stx2, ehxA, eae subtype β1 is herein reported. The infant did not require dialysis during the acute stage of HUS, evolved favorably, maintained normal blood pressure and normal renal function and had no recurrence until the last control. This could be due to several factors, such as the characteristics of infecting STEC strain and a reduction in host susceptibility to renal injury. This report highlights the regional participation of non-O157 STEC in childhood diseases and the importance of performing active surveillance for all forms of HUS.

  18. [Occurrence of Salmonella spp. and shigatoxin-producing escherichia coli (STEC) in horse faeces and horse meat products].

    PubMed

    Pichner, Rohtraud; Sander, Andrea; Steinrück, Hartmut; Gareis, Manfred

    2005-01-01

    In order to assess the relevance of horses as a possible reservoir of Salmonella and Shigatoxin-producing Escherichia coli (STEC), 400 samples of horse faeces and 100 samples of horse meat products were examined by PCR-screening methods. Salmonella enterica was not found in any of the samples. One faeces-sample and one horse meat product were proved to be STEC positive. The STEC-strain from faecal origin belonged to the serotype 0113:H21 and had the stx 2c gene and the enterohemolysin gene. The STEC-strain isolated from a horse meat product had the serotype O87:H16 and the stx 2d gene. The results indicate a very low risk for human to get a Salmonella- or EHEC- infection from horses in Germany.

  19. Binding of Soluble Natural Ligands to a Soluble Human T-Cell Receptor Fragment Produced in Escherichia coli

    NASA Astrophysics Data System (ADS)

    Hilyard, Katherine L.; Reyburn, Hugh; Chung, Shan; Bell, John I.; Strominger, Jack L.

    1994-09-01

    An Escherichia coli expression system has been developed to produce milligram quantities of the variable domains of a human T-cell receptor from a cytotoxic T cell that recognizes the HLA-A2-influenza matrix peptide complex as a single polypeptide chain. The recombinant protein was purified by metal-chelate chromatography and then refolded in a redox buffer system. The refolded protein was shown to directly bind both Staphylococcus aureus enterotoxin B and the major histocompatibility complex protein-peptide complex using a BIAcore biosensor. Thus this preparation of a single-chain, variable-domain, T-cell receptor fragment can bind both of its natural ligands and some of it is therefore a functional fragment of the receptor molecule.

  20. Ambler Class A Extended-Spectrum Beta-Lactamase-Producing Escherichia coli and Klebsiella spp. in Canadian Hospitals

    PubMed Central

    Mulvey, Michael R.; Bryce, Elizabeth; Boyd, David; Ofner-Agostini, Marianna; Christianson, Sara; Simor, Andrew E.; Paton, Shirley

    2004-01-01

    This report describes a study carried out to gain baseline information on the molecular characteristics of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella spp. in Canada. A total of 29,323 E. coli and 5,156 Klebsiella sp. isolates were screened at 12 participating sites. Of these, 505 clinically significant, nonrepeat isolates displaying reduced susceptibility to the NCCLS-recommended beta-lactams were submitted to a central laboratory over a 1-year period ending on 30 September 2000. A total of 116 isolates were confirmed to be ESBL producers. PCR and sequence analysis revealed the presence of TEM-11 (n = 1), TEM-12 (n = 1), TEM-29 (n = 1), TEM-52 (n = 4), CTX-M-13 (n = 1), CTX-M-14 (n = 15), CTX-M-15 (n = 11), SHV-2 (n = 2), SHV-2a (n = 12), SHV-5 (n = 6), SHV-12 (n = 45), and SHV-30 (n = 2). Five novel beta-lactamases were identified and designated TEM-115 (n = 2), TEM-120 (n = 1), SHV-40 (n = 2), SHV-41 (n = 4), and SHV-42 (n = 1). In addition, no molecular mechanism was identified for five isolates displaying an ESBL phenotype. Macrorestriction analysis of all ESBL isolates was conducted, as was restriction fragment length polymorphism analysis of plasmids harboring ESBLs. Although a “clonal” distribution of isolates was observed at some individual sites, there was very little evidence suggesting intrahospital spread. In addition, examples of identical or closely related plasmids that were identified at geographically distinct sites across Canada are given. However, there was considerable diversity with respect to plasmid types observed. PMID:15047521

  1. The Polymorphic Aggregative Phenotype of Shiga Toxin-Producing Escherichia coli O111 Depends on RpoS and Curli

    PubMed Central

    Diodati, M. E.; Bates, A. H.; Miller, W. G.; Carter, M. Q.; Zhou, Y.

    2015-01-01

    Escherichia coli O111 is an emerging non-O157:H7 serotype of Shiga toxin-producing E. coli (STEC). We previously reported that outbreak and environmental, but not sporadic-case, strains of STEC O111 share a distinct aggregation phenotype (M. E. Diodati, A. H. Bates, M. B. Cooley, S. Walker, R. E. Mandrell, and M. T. Brandl, Foodborne Pathog Dis 12:235−243, 2015, http://dx.doi.org/10.1089/fpd.2014.1887). We show here the natural occurrence of nonaggregative variants in single STEC O111 strains. These variants do not produce curli fimbriae and lack RpoS function but synthesize cellulose. The deletion of csgBAC or rpoS in an aggregative outbreak strain abolished aggregate formation, which was rescued when curli biogenesis or RpoS function, respectively, was restored. Complementation of a nonaggregative variant with RpoS also conferred curli production and aggregation. These observations were supported by Western blotting with an anti-CsgA antibody. Immunomicroscopy revealed that curli were undetectable on the cells of the nonaggregative variant and the RpoS mutant but were present in large quantities in the intercellular matrix of the assemblages formed by aggregative strains. Sequence analysis of rpoS in the aggregative strain and its variant showed a single substitution of threonine for asparagine at amino acid 124. Our results indicate that the multicellular behavior of STEC O111 is RpoS dependent via positive regulation of curli production. Aggregation may confer a fitness advantage in O111 outbreak strains under stressful conditions in hydrodynamic environments along the food production chain and in the host, while the occurrence of nonaggregative variants may allow the cell population to adapt to conditions benefiting a planktonic lifestyle. PMID:26712542

  2. Characterization of the Shiga toxin-producing Escherichia coli O26 isolated from human in Poland between 1996 and 2014.

    PubMed

    Januszkiewicz, A; Wołkowicz, T; Chróst, A; Szych, J

    2015-06-01

    Shiga toxin-producing Escherichia coli (STEC) O26 infections can be comparable with STEC O157 infections in severity of the acute haemolytic-uremic syndrome HUS and long-term sequelae. Among O26 STEC isolates, highly virulent clone O26:H11/H- Sequence Type 29 (ST 29) emerged in Germany in mid-1990s and spread to European countries. However, up to date, no STEC O26:H11/H- belonging to ST29 has been documented in Poland. In this study, we determined the relationship and clonal structure, stx genotypes, plasmid gene profiles and antimicrobial resistance of nine human STEC O26:H11/H- strains from human patients in Poland between 1996 and 2014. Of the 9 human STEC O26:H11/H- strains, two belonged to ST29 and were isolated from two children with HUS and renal failure with sepsis respectively. These strains showed the molecular characteristics of the emerging human-pathogenic ST29 clone (stx1-, stx2a+, eae+, ehxA+, etpD+, katP-, espP-). The remaining STEC O26:H11/H- strains examined in this study, belonged to ST21, with plasmid genes profiles frequently reported in ST21 strains in Europe. STEC O26 infections with serious human health consequences highlight the need of continuous surveillance of non-O157 STEC and implementation of the diagnostic approaches focused on their detection. Significance and impact of the study: These study provides the first data on the occurrence of emerging Shiga toxin-producing Escherichia coli O26:H11 ST 29 clone in human patients in Poland. Those strains show the molecular characteristics of highly virulent new ST29 pathotype (stx1-, stx2a+, eae+ ehxA+, etpD+, katP-, espP-). These results demonstrated prompt efforts to implement diagnostic approaches detection of those pathogen in the European countries.

  3. Novel sequence types of non-O157 Shiga toxin-producing Escherichia coli isolated from cattle.

    PubMed

    Isiko, J; Khaitsa, M; Bergholz, T M

    2015-06-01

    The objective of this study was to assess the genetic diversity of non-O157 Shiga toxin-producing Escherichia coli (STEC) isolates from cattle. Multi-locus sequence typing (MLST) was used to identify and compare the sequence types (STs) of 43 non-O157 STEC cattle isolates using the EcMLST database curated by the STEC Center at Michigan State University. For the 43 isolates, 19 STs were identified and 10 of those STs were novel compared to those in EcMLST. For the 43 isolates, 19 different serotypes were identified. STEC O22:H8, O174:H28 and O8:H19 were most common, and STEC O8 isolates were the most diverse, with seven different STs for isolates with that O group. STEC strains with O types identified in this study have been isolated from cattle by other researchers, as well as from cases of human gastroenteritis. Of the 10 novel STs identified, six were found to be closely related to previously identified STs, indicating that populations of non-O157 STEC in cattle are similar to those from other sources, including human clinical cases. Significance and impact of the study: The foodborne pathogen Shiga toxin-producing Escherichia coli (STEC) is a significant public health concern. One of the main reservoirs for STEC are cattle, which can directly or indirectly contribute to STEC in the food supply. The genetic subtype data presented here highlight the diversity of STEC that can be isolated from cattle. These results further our understanding of the ecology of STEC in the primary production environment, which is important for developing effective control measures to reduce this pathogen in the food supply.

  4. Adherence of curli producing Shiga-toxigenic Escherichia coli to baby spinach leaves

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cellular appendages, such as curli fibers have been suggested to be involved in STEC persistence in fresh produce as these curli are critical in biofilm formation and adherence to animal cells. We determined the role of curli in attachment of STEC on spinach leaves. The curli expression by wild-ty...

  5. Beef carcass contamination by Shiga toxin-producing Escherichia coli strains in an abattoir in Brazil: characterization and resistance to antimicrobial drugs.

    PubMed

    Rigobelo, Everlon Cid; Santo, Edilene; Marin, Jose Moacir

    2008-12-01

    A survey was performed to estimate the frequency of Escherichia coli and Shiga toxin-producing E. coli (STEC) in carcasses obtained from an abattoir in Brazil between February 2006 and June 2007. A total of 216 beef carcasses were sampled at three stages of the slaughter process--preevisceration, postevisceration, and postprocessing--during the rain and dry seasons, respectively. Of the carcasses sampled, 58% were preevisceration E. coli positive, 38% were postevisceration positive, and 32% postprocessing positive. At the postprocessing stage, the isolation of E. coli was twice as high in the rain season. E. coli was isolated from 85 carcasses of which only 3 (1.4%) were positive for stx-encoding genes. No E. coli O157 serogroup isolates were detected. No antimicrobial resistance was found in nine of the isolates (10% of the total). The most frequent resistances were seen against cephalothin (78%), streptomycin (38%), nalidixic acid (36%), and tetracycline (30%). Multidrug resistance (MDR) to three or more antimicrobial agents was determined in 28 (33%) E. coli isolates. The presence of STEC and MDR strains among the isolates in the beef carcasses emphasizes the importance of proper handling to prevent carcass contamination.

  6. Detection of Shiga toxin (Stx)-producing Escherichia coli (STEC) in bovine dairy herds in Northern Italy.

    PubMed

    Trevisani, M; Mancusi, R; Delle Donne, G; Bacci, C; Bassi, L; Bonardi, S

    2014-08-01

    The aim of this study was to monitor the presence of Shiga toxin (Stx)-producing Escherichia coli in dairy farms authorized to sell raw milk and other farms, located in the same area, which sell milk to industry or use it to produce Parmesan or Grana cheese. Our research was focused on the serogroups O157 and O26, which are the most common in human cases in Italy and genetic markers that characterize the strains that can cause hemorrhagic colitis and hemolytic uremic syndrome (EHEC) in humans. Overall, 255 bulk-milk and 225 milk filter samples were screened for the presence of Shiga toxin genes (stx1 and stx2), O157 and O26 serogroups by using PCR. The samples were collected in 193 bovine dairy farms located in Northern Italy, including 32 farms selling raw milk to consumers. According to the preliminary PCR screening test, 32 out of 255 (12.5%; CI95%, 8.7% to 17.3%) bulk milk samples and 68 out of 225 (30.2%; CI95%, 24.3% to 36.7%) milk filters were positive for stx genes. Of the 32 milk samples that were stx-positive, 4 (1.6%, CI95%, 0.4% to 4%) were also positive by PCR for the rfbEO157 gene and 6 (2.4%, CI95%, 0.9% to 5.1%) were positive for the wzxO26 gene. The culture detection method, which was based on the immunomagnetic separation, achieved isolation rates of E. coli serogroups O157 and O26 in 25-67% of the milk samples that tested positive by PCR for these serogroups. STEC O26 was detected in one milk filter (1.6%) from a farm that sells raw milk to consumers directly and one sample (1.4%) of bulk milk intended for pasteurization. The presence of STEC O157 was also detected in 2 milk filters (1.7%) from farms that use milk to produce Grana cheese. All the STEC stains O157 and O26 isolated carried the genes eae and espK and genes belonging to the pathogenicity island OI-122 (efa1/2, sen, pagC), which are markers suitable for screening the human virulent EHEC strains. These virulence markers were also detected in the three strains of stx-negative E. coli O

  7. First description of OXA-48-producing Escherichia coli and the pandemic clone ST131 from patients hospitalised at a military hospital in Algeria.

    PubMed

    Agabou, A; Pantel, A; Ouchenane, Z; Lezzar, N; Khemissi, S; Satta, D; Sotto, A; Lavigne, J-P

    2014-09-01

    The aim of the study was to assess the frequency and diversity of carbapenemases and extended-spectrum β-lactamases (ESBL) produced by Escherichia coli isolates from patients hospitalised in the Regional Military Hospital of Constantine (Algeria). E. coli isolates were collected over a 2-year period from patients presenting E. coli infections. Strains with reduced susceptibility to ertapenem and/or positive for ESBL were characterised with regard to antibiotic resistance, bla genes, phylogenetic groups, O25 serotyping, quinolone resistance, repetitive sequence-based polymerase chain reaction (rep-PCR) profiles and multi-locus sequence typing (MLST). Of the 448 isolated E. coli, 94 (20.9 %) were multidrug-resistant. One of them (1.1 %) produced a bla OXA-48 and was identified as a B1 ST5 strain. The transposon bearing this gene was Tn1999.2. This strain was isolated from a patient coming from a border province with Tunisia, where this carbapenemase is endemic. In addition, 84 (18.8 %) isolates among them produced an ESBL with predominance (97.6 %) of bla CTX-M-15, which was coupled with qnr genes in 10.9 %. ESBL-producing strains were mainly detected in phylogroups D and A. They displayed 20 rep-PCR profiles and all the clonally related isolates were of the same sequence type (ST). Ten strains (9.4 %) belonged to the pandemic clone ST131. This study describes for the first time the presence of OXA-48-producing E. coli and the emergence of the intercontinental ST131 bla CTX-15-producing E. coli strains in Algeria.

  8. [ESBL-producing E. coli and EHEC in dogs and cats in the Tyrol as possible source of human infection].

    PubMed

    Franiek, Natalie; Orth, Dorothea; Grif, Katharina; Ewers, Christa; Wieler, Lothar H; Thalhammer, Johann G; Würzner, Reinhard

    2012-01-01

    In contrast to infections with enterohaemorrhagic E. coli (EHEC), which are thought to be classical zoonosis, the zoonotic potential of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae is still widely unknown. The aim of our study was to determine the frequency of EHEC and ESBL-producing Enterobacteriaceae in domestic animals (dogs and cats) in the Tyrol. Among 228 fecal samples of dogs (n = 92) and cats (n = 136) three samples (1.3%) were positive in the EHEC-ELISA. In two of the three cases isolation of the organism was not possible, the third sample of a two-year-old crossbreed bitch yielded EHEC O103:H2. In twelve of 228 (5.3%) fecal samples 13 ESBL-producing Enterobacteriaceae (in ten cats and two dogs) were found.These animals mainly derived from homes for animals (ten animals, 83%). 75% of the isolates belonged to the CTX-M-1-group, 8% to the CTX-M-2-group and 17% to the CTX-M-9-group. One isolate was positive for CTX-M-1 and CTX-M-9. Typing of the 13 ESBL-producing isolates by multilocus sequence typing (MLST) showed ten different sequence types, which points out the importance of the horizontal transfer of mainly plasmid-coded ESBL genes. Transmission of EHEC and ESBL-producing Enterobacteriaceae from domestic animals to humans is possible, corroborated by the fact that the EHEC serotype found in one dog and the sequence types detected by MLST in several dogs and cats were previously reported to occur in severe human infection.

  9. High rate of per oral mecillinam treatment failure in community-acquired urinary tract infections caused by ESBL-producing Escherichia coli.

    PubMed

    Søraas, Arne; Sundsfjord, Arnfinn; Jørgensen, Silje Bakken; Liestøl, Knut; Jenum, Pål A

    2014-01-01

    A population-based study was performed to investigate the efficacy of mecillinam treatment of community-acquired urinary tract infections (CA-UTI) caused by extended-spectrum β-lactamase (ESBL) producing Escherichia coli. The study was conducted in South-Eastern Norway. Data from patients with CA-UTI caused by ESBL-producing and non-producing (random controls) E. coli were collected through interviews, questionnaires, medical records and the Norwegian Prescription Database. Treatment failure was defined as a new antibiotic prescription appropriate for UTI prescribed within two weeks after the initial antimicrobial therapy. Multivariable logistic regression analysis was performed to identify treatment agents and patient- or bacterial traits associated with treatment failure. A total of 343 patients (mean age 59) were included, of which 158 (46%) were treated with mecillinam. Eighty-one patients (24%, mean age 54) had infections caused by ESBL producing E. coli, and 41 of these patients (51%) received mecillinam as the primary treatment. Mecillinam treatment failure was observed in 18 (44%) of patients infected by ESBL-producing strains and in 16 (14%) of patients with a CA-UTI caused by ESBL non-producing strains. Multivariable analysis showed that ESBL status (odds ratio (OR) 3.2, 95% confidence interval (CI) 1.3-7.8, p = 0.009) and increased MIC of mecillinam (OR 2.0 for each doubling value of MIC, CI 1.4-3.0, p<0.001) were independently associated with mecillinam treatment failure. This study showed a high rate of mecillinam treatment failure in CA-UTIs caused by ESBL producing E. coli. The high failure rate could not be explained by the increased MIC of mecillinam alone. Further studies addressing the use of mecillinam against ESBL-producing E. coli, with emphasis on optimal dosing and combination therapy with β-lactamase inhibitors, are warranted.

  10. Epidemiology of Escherichia coli, Klebsiella species, and Proteus mirabilis strains producing extended-spectrum β-lactamases from clinical samples in the Kinki Region of Japan.

    PubMed

    Nakamura, Tatsuya; Komatsu, Masaru; Yamasaki, Katsutoshi; Fukuda, Saori; Miyamoto, Yugo; Higuchi, Takeshi; Ono, Tamotsu; Nishio, Hisaaki; Sueyoshi, Noriyuki; Kida, Kenji; Satoh, Kaori; Toda, Hirofumi; Toyokawa, Masahiro; Nishi, Isao; Sakamoto, Masako; Akagi, Masahiro; Nakai, Isako; Kofuku, Tomomi; Orita, Tamaki; Wada, Yasunao; Zikimoto, Takuya; Koike, Chihiro; Kinoshita, Shohiro; Hirai, Itaru; Takahashi, Hakuo; Matsuura, Nariaki; Yamamoto, Yoshimasa

    2012-04-01

    In the present study, nonduplicate, clinical isolates of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli, Klebsiella spp, and Proteus mirabilis were collected during a 10-year period from 2000 to 2009 at several hospitals in the Kinki region, Japan. The detection rate of E coli markedly increased from 0.24% to 7.25%. The detection rate of Klebsiella pneumoniae increased from 0% to 2.44% and that of P mirabilis from 6.97% to 12.85%. The most frequently detected genotypes were the CTX-M9 group for E coli, the CTX-M2 group for K pneumoniae, and the CTX-M2 group for P mirabilis. E coli clone O25:H4-ST131 producing CTX-M-15, which is spreading worldwide, was first detected in 2007. The most common replicon type of E coli was the IncF type, particularly FIB, detected in 466 strains (69.7%). Of the K pneumoniae strains, 47 (55.3%) were of the IncN type; 77 P mirabilis strains (96.3%) were of the IncT type. In the future, the surveillance of various resistant bacteria, mainly ESBL-producing Enterobacteriaceae, should be expanded to prevent their spread.

  11. Isolation of Shiga Toxin-Producing Escherichia coli from Ground Beef Using Multiple Combinations of Enrichment Broths and Selective Agars.

    PubMed

    Brusa, Victoria; Piñeyro, Pablo E; Galli, Lucía; Linares, Luciano H; Ortega, Emanuel E; Padola, Nora L; Leotta, Gerardo A

    2016-03-01

    Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens, and beef cattle are recognized as the principal reservoir. The aims of this study were (1) to identify the most sensitive combination of selective enrichment broths and agars for STEC isolation in artificially inoculated ground beef samples, and (2) to evaluate the most efficient combination(s) of methods for naturally contaminated ground beef samples. A total of 192 ground beef samples were artificially inoculated with STEC and non-stx bacterial strains. A combination of four enrichment broths and three agars were evaluated for sensitivity, specificity, and positive predictive value for STEC isolation from experimentally inoculated samples. Enrichments with either modified tryptic soy broth (mTSB) containing 8 mg/L novobiocin (mTSB-8) or modified Escherichia coli (mEC) broth followed by isolation in MacConkey agar were the most sensitive combinations for STEC isolation of artificially inoculated samples. Independently, both enrichments media followed by isolation in MacConkey were used to evaluate ground beef samples from 43 retail stores, yielding 65.1% and 58.1% stx-positive samples by RT-PCR, respectively. No difference was observed in the isolate proportions between these two methods (8/25 [32%] and 8/28 [28.6%]). Identical serotypes and stx genotypes were observed in STEC strains isolated from the same samples by either method. In this study, no single enrichment protocol was sufficient to detect all STEC in artificially inoculated samples and had considerable variation in detection ability with naturally contaminated samples. Moreover, none of the single or combinations of multiple isolation agars used were capable of identifying all STEC serogroups in either artificially inoculated or naturally occurring STEC-contaminated ground beef. Therefore, it may be prudent to conclude that there is no single method or combination of isolation methods capable of identifying all STEC serogroups

  12. Detection of Healthcare-Related Extended-Spectrum Beta-Lactamase-Producing Escherichia coli Transmission Events Using Combined Genetic and Phenotypic Epidemiology

    PubMed Central

    Boers, Stefan A.; Jansen, Ruud; Hays, John P.; Goessens, Wil H. F.; Vos, Margreet C.

    2016-01-01

    Background Since the year 2000 there has been a sharp increase in the prevalence of healthcare-related infections caused by extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli. However, the high community prevalence of ESBL-producing E. coli isolates means that many E. coli typing techniques may not be suitable for detecting E. coli transmission events. Therefore, we investigated if High-throughput MultiLocus Sequence Typing (HiMLST) and/or Raman spectroscopy were suitable techniques for detecting recent E. coli transmission events. Methods This study was conducted from January until December 2010 at Erasmus University Medical Center, Rotterdam, the Netherlands. Isolates were typed using HiMLST and Raman spectroscopy. A genetic cluster was defined as two or more patients carrying identical isolates. We used predefined definitions for epidemiological relatedness to assess healthcare-related transmission. Results We included 194 patients; strains of 112 patients were typed using HiMLST and strains of 194 patients were typed using Raman spectroscopy. Raman spectroscopy identified 16 clusters while HiMLST identified 10 clusters. However, no healthcare-related transmission events were detected. When combining data from both typing techniques, we identified eight clusters (n = 34 patients), as well as 78 patients with a non-cluster isolate. However, we could not detect any healthcare-related transmission in these 8 clusters. Conclusions Although clusters were genetically detected using HiMLST and Raman spectroscopy, no definite epidemiological relationships could be demonstrated which makes the possibility of healthcare-related transmission events highly unlikely. Our results suggest that typing of ESBL-producing E. coli using HiMLST and/or Raman spectroscopy is not helpful in detecting E. coli healthcare-related transmission events. PMID:27463231

  13. Molecular Epidemiology over an 11-Year Period (2000 to 2010) of Extended-Spectrum β-Lactamase-Producing Escherichia coli Causing Bacteremia in a Centralized Canadian Region

    PubMed Central

    Peirano, Gisele; van der Bij, Akke K.; Gregson, Daniel B.

    2012-01-01

    A study was designed to assess the importance of sequence types among extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates causing bacteremia over an 11-year period (2000 to 2010) in a centralized Canadian region. A total of 197 patients with incident infections were identified; the majority presented with community-onset urosepsis, with a significant increase in the prevalence of ESBL-producing E. coli during the later part of the study. The majority of E. coli isolates produced either CTX-M-15 or CTX-M-14. We identified 7 different major sequence types among 91% of isolates (i.e., the ST10 clonal complex, ST38, ST131, ST315, ST393, ST405, and ST648) and provided insight into their clinical and molecular characteristics. ST38 was the most antimicrobial-susceptible sequence type and predominated during 2000 to 2004 but disappeared after 2008. ST131 was the most antimicrobial-resistant sequence type, and the influx of a single pulsotype of this sequence type was responsible for the significant increase of ESBL-producing E. coli strains since 2007. During 2010, 49/63 (78%) of the ESBL-producing E. coli isolates belonged to ST131, and this sequence type had established itself as a major drug-resistant pathogen in Calgary, Alberta, Canada, posing an important new public health threat within our region. We urgently need well-designed epidemiological and molecular studies to understand the dynamics of transmission, risk factors, and reservoirs for E. coli ST131. This will provide insight into the emergence and spread of this multiresistant sequence type. PMID:22162555

  14. Anaerobic respiration in engineered Escherichia coli with an internal electron acceptor to produce fuel ethanol.

    PubMed

    Peterson, Joy Doran; Ingram, Lonnie O

    2008-03-01

    Environmental concerns and unease with U.S. dependence on foreign oil have renewed interest in converting biomass into fuel ethanol. The volume of plant matter available makes lignocellulose conversion to ethanol desirable, although no one isolated organism has been shown to break bonds in lignocellulose and efficiently metabolize resulting sugars into one product. This work reviews directed engineering coupled with metabolic evolution resulting in microbial biocatalysts that produce up to 45 g L(-1) ethanol in 48 hours in a simple mineral salts medium and that convert various compounds of lignocellulosic materials to ethanol. Mutations contributing to ethanologenesis are discussed along with adding enzymatic capabilities to existing biocatalysts in order to decrease the commercial enzymes required to reduce plant matter into fermentable sugars.

  15. Virulence Profiles of Bacteremic Extended-Spectrum β-Lactamase-Producing Escherichia coli: Association with Epidemiological and Clinical Features

    PubMed Central

    Rodríguez-Baño, Jesús; Mingorance, Jesús; Fernández-Romero, Natalia; Serrano, Lara; López-Cerero, Lorena; Pascual, Alvaro

    2012-01-01

    There is scarce data about the importance of phylogroups and virulence factors (VF) in bloodstream infections (BSI) caused by extended-spectrum β-lactamase-producing Escherichia coli (ESBLEC). A prospective multicenter Spanish cohort including 191 cases of BSI due to ESBLEC was studied. Phylogroups and 25 VF genes were investigated by PCR. ESBLEC were classified into clusters according to their virulence profiles. The association of phylogropus, VF, and clusters with epidemiological features were studied using multivariate analysis. Overall, 57.6%, 26.7%, and 15.7% of isolates belonged to A/B1, D and B2 phylogroups, respectively. By multivariate analysis (adjusted OR [95% CI]), virulence cluster C2 was independently associated with urinary tract source (5.05 [0.96–25.48]); cluster C4 with sources other than urinary of biliary tract (2.89 [1.05–7.93]), and cluster C5 with BSI in non-predisposed patients (2.80 [0.99–7.93]). Isolates producing CTX-M-9 group ESBLs and from phylogroup D predominated among cluster C2 and C5, while CTX-M-1 group of ESBL and phylogroup B2 predominantes among C4 isolates. These results suggest that host factors and previous antimicrobial use were more important than phylogroup or specific VF in the occurrence of BSI due to ESBLEC. However, some associations between virulence clusters and some specific epidemiological features were found. PMID:22970186

  16. In vitro adhesion properties of Shiga toxin-producing Escherichia coli isolated from cattle, food, and humans

    PubMed Central

    Pradel, Nathalie; Etienne-Mesmin, Lucie; Thévenot, Jonathan; Cordonnier, Charlotte; Blanquet-Diot, Stéphanie; Livrelli, Valérie

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) are able to cause serious illnesses ranging from diarrhea to hemorrhagic colitis and hemolytic-uremic syndrome (HUS). These bacteria colonize the digestive tract of humans and produce Shiga-toxins, which are considered to be essential for virulence and are crucial in lethal infection. Colon colonization is supposed to be a determinant step in the development of the infection, but the virulence traits that mediate this step are unclear. We analyzed the ability of 256 STEC strains belonging to seropathotype A (the most virulent O157:H7 serotype) to seropathotype E (not involved in human disease) to adhere to HEp-2, HCT-8, and T84 cell lines. Of the 256 STEC tested most (82%) were non-adherent in our assays. The adhesion levels were globally low and were not related to pathogenicity, although the highest levels were associated to O26:H11 and O103:H2 strains of seropathotype B (associated with HUS but less commonly than serotype O157:H7), possessing both the eae and toxB genes. PMID:25774152

  17. High-level expression and purification of recombinant human growth hormone produced in soluble form in Escherichia coli.

    PubMed

    Levarski, Zdenko; Šoltýsová, Andrea; Krahulec, Ján; Stuchlík, Stanislav; Turňa, Ján

    2014-08-01

    Human growth hormone (hGH) was one of the first recombinant proteins approved for the treatment of human growth disorders. Its small size (191 amino acids), possession of only 2 disulphide bonds and absence of posttranslational modifications make Escherichia coli the host of choice for its production on any scale. In this work, we have utilized an efficient T7 based expression system to produce high levels of soluble thioredoxin-hGH (Trx-hGH) fusion protein. We outline a relatively simple three step purification process employing two immobilized metal-affinity chromatography and one anion-exchange steps and removal of fusion partner by enterokinase cleavage yielding native hGH. The ability of cell populations to produce quantities of up to 1 g/L of the soluble Trx-hGH fusion protein has been tested in flask cultivations as well as in batch and fed-batch bioreactor runs. The sequence and structure of derived hGH were confirmed by mass spectrometry and circular dichroism and its native function, to induce cell proliferation, was confirmed by employing a Nb2 cell line proliferation assay.

  18. Production of hydroxy-fatty acid derivatives from waste oil by Escherichia coli cells producing fungal cytochrome P450foxy.

    PubMed

    Kitazume, Tatsuya; Yamazaki, Yuya; Matsuyama, Shigeru; Shoun, Hirofumi; Takaya, Naoki

    2008-07-01

    Cytochrome P450foxy (P450foxy) is a fatty acid (FA) monooxygenase that is characterized by self-sufficient catalysis and high turnover numbers due to the fused structure of cytochrome P450 and its reductase. Here we found that resting recombinant Escherichia coli cells producing P450foxy converted saturated FA with a chain length of 7-16 carbon atoms to their omega-1 to omega-3 hydroxy derivatives. Most products were recovered from the culture supernatant. Decanoic acid was most efficiently converted to omega-1 to omega-3 hydroxy decanoic acids in the order of omega-1>omega-2>omega-3, with a total product yield of 47%. We also found that P450foxy was more active against physiological fatty acyl esters such as monopalmitoyl glycerol, monopalmitoyl phospholipid, and palmitoyl CoA than free palmitic acid. The bacteria producing P450foxy were applicable as biocatalysts in the production of omega-1 hydroxy palmitic acid from lard, vegetable, and soy sauce oil wastes from the food industry.

  19. A New Immunoassay for Detecting All Subtypes of Shiga Toxins Produced by Shiga Toxin-Producing E. coli in Ground Beef

    PubMed Central

    He, Xiaohua; Kong, Qiulian; Patfield, Stephanie; Skinner, Craig; Rasooly, Reuven

    2016-01-01

    Background Shiga toxin (Stx) is a common virulence factor of all Shiga toxin producing E. coli (STEC) that cause a wide spectrum of disease, including hemorrhagic colitis and hemolytic uremic syndrome (HUS). Although several commercial kits are available for detection of Stx produced by STEC, none of them are capable of recognizing all subtypes of Stxs, which include three subtypes of Stx1 and seven subtypes of Stx2. Methods and Findings New monoclonal and polyclonal antibodies against Stx1 and Stx2 were developed. A universal sandwich ELISA capable of detecting all known subtypes of Stx1 and Stx2 was established using a pool of newly developed antibodies. To precisely monitor the sensitivity of the assay for each subtype of Stxs, recombinant toxoids were created and used as standards in ELISAs. Because of the high affinity of the antibodies incorporated, the ELISA assay is highly sensitive with a limit of detection for the different subtypes of Stx1a and Stx2a between 10 and 50 pg/mL in phosphate buffered saline (PBS). The assay was also able to identify STEC based on the production of Stxs using the supernatants of culture fluids or even single colonies on agar plates without lengthy enrichment in liquid medium. When applied to ground beef samples, this newly developed ELISA was capable of distinguishing beef samples spiked with a single bacterial cell. Conclusions A highly sensitive and universal assay for all subtypes of Stx1 and Stx2 was developed. It has significantly improved upon the current technologies by avoiding false negative results due to the narrow detection range of the assay. The assay developed in this study can be useful for prompt detection of new and emerging serotypes and screening ground beef samples for contamination of STEC at an early stage in the food supply chain, thus avoiding the need for possible recall. PMID:26824247

  20. Acid Resistance and Molecular Characterization of Escherichia coli O157:H7 and Different Non-O157 Shiga Toxin-Producing E. coli Serogroups.

    PubMed

    Kim, Gwang-Hee; Breidt, Frederick; Fratamico, Pina; Oh, Deog-Hwan

    2015-10-01

    The objective of this study was to compare the acid resistance (AR) of non-O157 Shiga toxin-producing Escherichia coli (STEC) strains belonging to serogroups O26, O45, O103, O104, O111, O121, and O145 with O157:H7 STEC isolated from various sources in 400 mM acetic acid solutions (AAS) at pH 3.2 and 30 °C for 25 min with or without glutamic acid. Furthermore, the molecular subgrouping of the STEC strains was analyzed with the repetitive sequence-based PCR (rep-PCR) method using a DiversiLab(TM) system. Results for a total of 52 strains ranged from 0.31 to 5.45 log reduction CFU/mL in the absence of glutamic acid and 0.02 to 0.33 CFU/mL in the presence of glutamic acid except for B447 (O26:H11), B452 (O45:H2), and B466 (O104:H4) strains. Strains belonging to serogroups O111, O121, and O103 showed higher AR than serotype O157:H7 strains in the absence of glutamic acid. All STEC O157:H7 strains exhibited a comparable DNA pattern with more than 95% similarity in the rep-PCR results, as did the strains belonging to serogroups O111 and O121. Surprisingly, the DNA pattern of B458 (O103:H2) was similar to that of O157:H7 strains with 82% similarity, and strain B458 strain showed the highest AR to AAS among the O103 strains with 0.44 log reduction CFU/mL without glutamic acid. In conclusion, STEC serotypes isolated from different sources exhibited diverse AR and genetic subtyping patterns. Results indicated that some non-O157 STEC strains may have higher AR than STEC O157:H7 strains under specific acidic conditions, and the addition of glutamic acid provided enhanced protection against exposure to AAS.

  1. Interactive effects of temperature, pH, and water activity on the growth kinetics of Shiga toxin-producing Escherichia coli O104:H4 3.

    PubMed

    Juneja, Vijay K; Mukhopadhyay, Sudarsan; Ukuku, Dike; Hwang, Cheng-An; Wu, Vivian C H; Thippareddi, Harshavardhan

    2014-05-01

    The risk of non-O157 Shiga toxin-producing Escherichia coli strains has become a growing public health concern. Several studies characterized the behavior of E. coli O157:H7; however, no reports on the influence of multiple factors on E. coli O104:H4 are available. This study examined the effects and interactions of temperature (7 to 46°C), pH (4.5 to 8.5), and water activity (aw ; 0.95 to 0.99) on the growth kinetics of E. coli O104:H4 and developed predictive models to estimate its growth potential in foods. Growth kinetics studies for each of the 23 variable combinations from a central composite design were performed. Growth data were used to obtain the lag phase duration (LPD), exponential growth rate, generation time, and maximum population density (MPD). These growth parameters as a function of temperature, pH, and aw as controlling factors were analyzed to generate second-order response surface models. The results indicate that the observed MPD was dependent on the pH, aw, and temperature of the growth medium. Increasing temperature resulted in a concomitant decrease in LPD. Regression analysis suggests that temperature, pH, and aw significantly affect the LPD, exponential growth rate, generation time, and MPD of E. coli O104:H4. A comparison between the observed values and those of E. coli O157:H7 predictions obtained by using the U. S. Department of Agriculture Pathogen Modeling Program indicated that E. coli O104:H4 grows faster than E. coli O157:H7. The developed models were validated with alfalfa and broccoli sprouts. These models will provide risk assessors and food safety managers a rapid means of estimating the likelihood that the pathogen, if present, would grow in response to the interaction of the three variables assessed. PMID:25198132

  2. Characterisation of Shiga toxin-producing Escherichia coli O157 strains isolated from humans in Argentina, Australia and New Zealand

    PubMed Central

    Leotta, Gerardo A; Miliwebsky, Elizabeth S; Chinen, Isabel; Espinosa, Estela M; Azzopardi, Kristy; Tennant, Sharon M; Robins-Browne, Roy M; Rivas, Marta

    2008-01-01

    Background Shiga toxin-producing Escherichia coli (STEC) is an important cause of bloody diarrhoea (BD), non-bloody diarrhoea (NBD) and the haemolytic uraemic syndrome (HUS). In Argentina and New Zealand, the most prevalent STEC serotype is O157:H7, which is responsible for the majority of HUS cases. In Australia, on the other hand, STEC O157:H7 is associated with a minority of HUS cases. The main aims of this study were to compare the phenotypic and genotypic characteristics of STEC O157 strains isolated between 1993 and 1996 from humans in Argentina, Australia and New Zealand, and to establish their clonal relatedness. Results Seventy-three O157 STEC strains, isolated from HUS (n = 36), BD (n = 20), NBD (n = 10), or unspecified conditions (n = 7) in Argentina, Australia and New Zealand, were analysed. The strains were confirmed to be E. coli O157 by biochemical tests and serotyping. A multiplex polymerase chain reaction (PCR) was used to amplify the stx1, stx2 and rfbO157 genes and a genotyping method based on PCR-RFLP was used to determine stx1 and stx2 variants. This analysis revealed that the most frequent stx genotypes were stx2/stx2c (vh-a) (91%) in Argentina, stx2 (89%) in New Zealand, and stx1/stx2 (30%) in Australia. No stx1-postive strains were identified in Argentina or New Zealand. All strains harboured the eae gene and 72 strains produced enterohaemolysin (EHEC-Hly). The clonal relatedness of strains was investigated by phage typing and pulsed-field gel electrophoresis (PFGE). The most frequent phage types (PT) identified in Argentinian, Australian, and New Zealand strains were PT49 (n = 12), PT14 (n = 9), and PT2 (n = 15), respectively. Forty-six different patterns were obtained by XbaI-PFGE; 37 strains were grouped in 10 clusters and 36 strains showed unique patterns. Most clusters could be further subdivided by BlnI-PFGE. Conclusion STEC O157 strains isolated in Argentina, Australia, and New Zealand differed from each other in terms of stx

  3. Antimicrobial resistance in faecal Escherichia coli isolates from farmed red deer and wild small mammals. Detection of a multiresistant E. coli producing extended-spectrum beta-lactamase.

    PubMed

    Alonso, C A; González-Barrio, D; Tenorio, Carmen; Ruiz-Fons, F; Torres, C

    2016-04-01

    Eighty-nine Escherichia coli isolates recovered from faeces of red deer and small mammals, cohabiting the same area, were analyzed to determine the prevalence and mechanisms of antimicrobial resistance and molecular typing. Antimicrobial resistance was detected in 6.7% of isolates, with resistances to tetracycline and quinolones being the most common. An E. coli strain carrying blaCTX-M-1 as well as other antibiotic resistant genes included in an unusual class 1 integron (Intl1-dfrA16-blaPSE-1-aadA2-cmlA1-aadA1-qacH-IS440-sul3-orf1-mef(B)Δ-IS26) was isolated from a deer. The blaCTX-M-1 gene was transferred by conjugation and transconjugants also acquired an IncN plasmid. This strain was typed as ST224, which seems to be well adapted to both clinical and environmental settings. The phylogenetic distribution of the 89 strains varied depending on the animal host. This work reveals low antimicrobial resistance levels among faecal E. coli from wild mammals, which reflects a lower selective pressure affecting these bacteria, compared to livestock. However, it is remarkable the detection of a multi-resistant ESBL-E. coli with an integron carrying clinically relevant antibiotic-resistance genes, which can contribute to the dissemination of resistance determinants among different ecosystems. PMID:27012919

  4. Risk Factors for Salmonella, Shiga Toxin-Producing Escherichia coli and Campylobacter Occurrence in Primary Production of Leafy Greens and Strawberries.

    PubMed

    Ceuppens, Siele; Johannessen, Gro S; Allende, Ana; Tondo, Eduardo César; El-Tahan, Fouad; Sampers, Imca; Jacxsens, Liesbeth; Uyttendaele, Mieke

    2015-08-18

    The microbiological sanitary quality and safety of leafy greens and strawberries were assessed in the primary production in Belgium, Brazil, Egypt, Norway and Spain by enumeration of Escherichia coli and detection of Salmonella, Shiga toxin-producing E. coli (STEC) and Campylobacter. Water samples were more prone to containing pathogens (54 positives out of 950 analyses) than soil (16/1186) and produce on the field (18/977 for leafy greens and 5/402 for strawberries). The prevalence of pathogens also varied markedly according to the sampling region. Flooding of fields increased the risk considerably, with odds ratio (OR) 10.9 for Salmonella and 7.0 for STEC. A significant association between elevated numbers of generic E. coli and detection of pathogens (OR of 2.3 for STEC and 2.7 for Salmonella) was established. Generic E. coli was found to be a suitable index organism for Salmonella and STEC, but to a lesser extent for Campylobacter. Guidelines on frequency of sampling and threshold values for E. coli in irrigation water may differ from region to region.

  5. Risk Factors for Salmonella, Shiga Toxin-Producing Escherichia coli and Campylobacter Occurrence in Primary Production of Leafy Greens and Strawberries

    PubMed Central

    Ceuppens, Siele; Johannessen, Gro S.; Allende, Ana; Tondo, Eduardo César; El-Tahan, Fouad; Sampers, Imca; Jacxsens, Liesbeth; Uyttendaele, Mieke

    2015-01-01

    The microbiological sanitary quality and safety of leafy greens and strawberries were assessed in the primary production in Belgium, Brazil, Egypt, Norway and Spain by enumeration of Escherichia coli and detection of Salmonella, Shiga toxin-producing E. coli (STEC) and Campylobacter. Water samples were more prone to containing pathogens (54 positives out of 950 analyses) than soil (16/1186) and produce on the field (18/977 for leafy greens and 5/402 for strawberries). The prevalence of pathogens also varied markedly according to the sampling region. Flooding of fields increased the risk considerably, with odds ratio (OR) 10.9 for Salmonella and 7.0 for STEC. A significant association between elevated numbers of generic E. coli and detection of pathogens (OR of 2.3 for STEC and 2.7 for Salmonella) was established. Generic E. coli was found to be a suitable index organism for Salmonella and STEC, but to a lesser extent for Campylobacter. Guidelines on frequency of sampling and threshold values for E. coli in irrigation water may differ from region to region. PMID:26295251

  6. Contact with farming environment as a major risk factor for Shiga toxin (Vero cytotoxin)-producing Escherichia coli O157 infection in humans.

    PubMed Central

    O'Brien, S. J.; Adak, G. K.; Gilham, C.

    2001-01-01

    In a prospective, unmatched case-control study of sporadic Shiga toxin (Vero cytotoxin)-producing Escherichia coli O157 (STEC O157) infection in England, exposure to the farming environment emerged strongly as a risk factor (adjusted odds ratio = 2.45; 95% confidence intervals = 1.49-4.02; p=0.0004) posing further challenges and opportunities for prevention. PMID:11747741

  7. An environmental shiga toxin-producing Escherichia coli O145 clonal population exhibits high-level phenotypic variation that includes virulence traits

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing Escherichia coli (STEC) serotype O145 is one of the major non-O157 serotypes associated with severe human disease. Here we examined the genetic diversity, population structure, virulence potential, and antibiotic resistance profile of environmental O145 strains isolated from a ...

  8. Mixed biofilm formation by Shiga toxin-producing Escherichia coli and Salmonella enterica serovar typhimurium enhanced bacterial resistance to sanitization due to extracellular polymeric substances

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin–producing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium are important foodborne pathogens capable of forming single-species biofilms or coexisting in multispecies biofilm communities. Bacterial biofilm cells are usually more resistant to sanitization than their pla...

  9. Mathematical modeling and numerical analysis of the growth of Non-O157 shiga toxin-producing Escherichia coli in spinach leaves

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study was conducted to investigate the growth of non-O157 Shiga toxin-producing Escherichia coli (STEC) in spinach leaves and to develop kinetic models to describe the bacterial growth. Six serogroups of non-O157 STEC, including O26, O45, O103, O111, O121, and O145, were used in the growth stu...

  10. A polyclonal antibody based immunoassay detects seven subtypes of Shiga toxin 2 produced by escherichia coli in human and environmental samples

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The increase of outbreaks and illnesses linked to Shiga toxin-producing Escherichia coli (STEC) has necessitated the development of effective detection methods for these pathogens in various matrices. The best way to determine if a bacterial strain is a STEC is to examine the production of Shiga tox...

  11. Characterization of shiga toxin-producing Escherichia coli recovered from domestic animals to determine stx variants, virulence genes, and cytotoxicity in mammalian cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shiga toxin-producing Escherichia coli (STEC) can cause foodborne illnesses ranging from diarrhea to severe diseases such as hemorrhagic colitis (HC), and hemolytic uremic syndrome (HUS) in humans. In this study, we determined virulence genes, stx subtypes and we evaluated the cytotoxicity in mammal...

  12. Canonical single nucleotide polymorphisms (SNPs) for high-resolution subtyping of Shiga-toxin producing Escherichia coli (STEC) O157:H7

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to develop a canonical SNP panel for subtyping of Shiga-toxin producing Escherichia coli (STEC). To this purpose, 906 putative SNPs were identified using resequencing tiling arrays. A subset of 391 SNPs was further screened using high-throughput TaqMan PCR against a d...

  13. The effect of deep frying or conventional oven cooking on inactivation of Shiga toxin-producing cells of Escherichia coli (STEC) in meatballs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We investigated the effects deep frying or oven cooking on inactivation of Shiga toxin-producing cells of Escherichia coli (STEC) in meatballs. A finely-ground veal and/or a beef-pork-veal mixture were inoculated (ca. 7.0 log CFU/g) with an eight-strain, genetically-marked cocktail of rifampicin-res...

  14. What is the best method? Recovery of methicillin-resistant Staphylococcus aureus and extended-spectrum β-lactamase-producing Escherichia coli from inanimate hospi